Introduction
Obesity is the most prevalent nutritional disorder among children and adolescents in the United States and many other resource-rich countries and transitional economies [1]. Childhood obesity has more than doubled in children and quadrupled in adolescents in the past 30 years [2] The percentage of children aged 6–11 years in the U.S. who were obese increased from 7 % in 1980 to nearly 18 % in 2012. Similarly, the percentage of adolescents aged 12–19 years who were obese increased from 5 % to nearly 21 % over the same period [2, 3]. In this country main factors contributing to the weight gain are widespread net of fast food restaurants and hypodynamic habits.  
In Europe last surveys exstimate up to 27 % of 13-year-olds and 33 % of 11-year olds are overweight. Among 11-year-old boys and girls, the prevalence of overweight was highest in Greece (33 %), Portugal (32 %), Ireland (30 %) and Spain (30 %) and lowest in the Netherlands (13 %) and Switzerland (11 %) [4]. Obesity is associated with multiorgan (namely pancreatic, adipose, hepatic, cardiac and muscle tissue) chronic metabolic and inflammatory alterations that together are termed ‘metabolic syndrome’. Childhood obesity predisposes to insulin resistance and type 2 diabetes, hypertension, hyperlipidemia, liver and renal disease, and reproductive dysfunction. This condition also increases the risk of adult-onset obesity and cardiovascular disease. Unequivocal experimental and clinical evidence causally link IL-1β and IL-18 to the development of these metabolic pathologies and their complications.
During nutritional surplus, in addition to adipocyte hypertrophy owing to increased lipid storage, adipose tissue is infiltrated by classically activated, M1, macrophages that secrete pro-inflammatory cytokines. NLRP3, ASC and caspase-1 are preferentially expressed in adipose-tissue-infiltrating macrophages, in which the saturated fatty acid palmitate and lipotoxic ceramides trigger NLRP3 inflammasome activation through a mechanism that involves defective autophagy and the accumulation of mitochondrial ROS.
Chronic hyperglycaemia as a result of peripheral insulin resistance is compensated for by increased insulin output by pancreatic β cells. Local inflammatory processes coupled with the toxic effects of glucose lead to accelerated mass loss of β cells and decreased insulin secretion over time, which prompts the progression from obesity and insulin resistance to overt T2DM. IL-1β, preferentially expressed by pancreatic infiltrating macrophages and to a smaller extent by β cells, has been implicated as a critical driver of β-cell death in conditions of chronic exposure to elevated concentrations of glucose [5].
No single definition of obesity in childhood and adolescence has gained universal approval. Some investigators have used the terms overweight, obese, and morbidly obese to refer to children and adolescents whose weights exceed those expected for heights by 20%, 50%, and 80-100%, respectively. The body mass index (BMI) has not been consistently used or validated in children younger than 2 years.  Many factors, including genetics, environment, metabolism, lifestyle, and eating habits, are believed to play a role in the development of obesity. However, more than 90% of cases are idiopathic; less than 10% are associated with hormonal or genetic causes. Recently, the gut microbiota has emerged as an important contributor to the development of obesity and metabolic disorders, through its interactions with environmental (e.g. diet) and genetic factors. Human intestinal microflora represents a complex ecosystem consisting of trillions of microorganisms and thousands of bacterial species that are deeply involved in different functions of host metabolism, modulation of inflammation, and body weight homeostasis.
 Nutrition is a driving factor in shaping gut microbiota composition and its functional maturation from the early stages of life. The gut microbiota acquired in early life have long term implications for host metabolism and gastrointestinal (GI), immune and neurological function [6]. Reduced diversity or dysbiosis are linked to childhood and later life disorders, including necrotizing enterocolitis [7], eczema [8], asthma[9], inflammatory bowel diseases [10], irritable bowel syndrome [11], obesity [12], diabetes [13] and autism [14].

Definition
The microbiome refers to the entire habitat, including the microorganisms (bacteria, archaea, lower and higher eurkaryotes, and viruses), their genomes (i.e., genes), and the surrounding environmental conditions. The microbiome is characterized by the application of one or combinations of metagenomics, metabonomics, metatranscriptomics, and metaproteomics combined with clinical or environmental metadata. Different parts of the body have different microbiomes, for example, the skin microbiome is different to the gut microbiome, but they are all part of the human microbiome [15].
The microbiota, includes all the microorganisms present in a defined environment identified using molecular methods relying predominantly on the analysis of 16S rRNA genes, 18S rRNA genes, or other marker genes and genomic regions, amplified and sequenced from given biological samples.
The normal gut microbiome comprises 100 trillion diverse microbes, whose collective genomes contain at least 100 times as many genes as our own eukaryote genome, mostly bacteria, encompassing over 1100 prevalent species, with at least 160 species in each individual (e.g., about up to 1.5 kg of bacteria in the human gut) with Firmicutes (35–80%) and Bacteriodetes (17–60%)  and Actinobacteria constituting the dominant phyla. Generally, Firmicutes and Bacteroidetes are most abundant, followed by Proteobacteria and Actinobacteria with minor contributors like Verrucomicrobia and Fusobacteria ."e;€¨It is believed that the ratio of potentially pathogenic to beneficial commensal microbes, rather than the presence of a specific organism or a group, is more crucial for disease development.
Faecalibacterium prausnitzii (F. prausnitzii) is the most abundant bacterium in the human intestinal microbiota of healthy adults. It represents more than 5% of the total bacterial population and it is a major representative of Firmicutes phylum, Clostridium class, Ruminococcaceae family [16].  
While the Bacteroidetes phylum mainly produces acetate and propionate, the Firmicutes phylum has butyrate as its primary metabolic end product.
The intestinal flora could be grouped in 3 main distinct enterotypes (Bacteroides, Prevotella or Ruminococcus) that are influenced by dietary intake:  Bacteroides enterotype is associated with a “western” protein rich diet as opposed to the Prevotella enterotype which was associated with a carbohydrate rich diet [17] (Fig. 1).

Fig.1 Taxonomic of microrganisms. (Taylor K. Soderborg & Sarah J. Borengasser & Linda A. Barbour & Jacob E. Friedman;  Microbial transmission from mothers with obesity or diabetes to infants: an innovative opportunity to interrupt a vicious cycle; Diabetologia (2016) 59:895–906 DOI 10.1007/s00125-016-3880-0).

Gut microbiome development
The intestinal flora of the human body is established in infancy and gradually stabilizes with age. By approximately 2 years of age, it is similar to the adult intestinal flora (Fig. 2).  Host genetics and environmental factors such as gestational age, delivery mode, diet, pre- and probiotics, antibiotics, maternal weight and stress influence the process. Several factors, such as diet and genetic background of the host and immune status, affect the composition of the microbiota (Fig. 3). Antibiotic use in human infancy, before age 6 months, was significantly associated with obesity development [18]. In contrast, perinatal administration of a Lactobacillus rhamnosus GG-based probiotic decreased excessive weight gain during childhood [19].
Caesarian section instead of vaginal delivery is an obvious example of the potential impact of medical practice on microbiota composition, with substantial differences in founding population, that may persist for months. Immediately after vaginal delivery, founding microbial populations in the baby closely resemble that of their mother’s vagina and mother’s milk, with lactobacilli predominating. [20, 21].
Early life nutrition with breast or formula feeding of newborns impact differently on the gut microbiome composition in the early stages of life. Bacterial composition and alpha-diversity differ between breastfed (BF) and formula-fed (FF) infants, and solid food introduction has been associated with rapid and sustained alterations in the fecal microbiota.  Initially after birth, the newborn fecal microbiota is composed primarily of facultative anaerobes, such as Staphylococcus, Streptococcus, Lactobacillus, and Enterobacteriaceae, which are thought to consume oxygen and prime the GI tract for colonization with obligate anaerobes such as Bifidobacterium, Bacteroides, and Clostridium [22]. From the first week of life, different colonization patterns are present in BF and FF infants. The fecal microbiota of BF infants is more stable over time, and characterized by a lower alpha-diversity compared to FF infants. Bifidobacteria have been shown to account for 70% of the sequences of exclusively breastfed (EBF) infants and appear earlier in the feces of EBF than in FF infants. For example, B. longum and B. breve are the dominant species in BF infants, whereas FF infants also harbor adult associated bifidobacterial species, such as B. adolescentis. The FF infant microbiome was enriched in functions characteristic of a more mature microbiota (e.g. more similar to that of an adult) such as bile acid synthesis, methanogenesis, and the phosphotransferase system, while the fecal microbiome of BF infants was enriched in patterns associated with synthesis of B vitamins and oxidative phosphorylation. A greater prevalence and higher proportion and total counts  of C. difficile in FF compared to BF infants, as well as, significantly more Peptostreptococcaceae, Akkermansia, Veillonella, and Enterococcus have been described. Escherichia abundance was also higher in FF than BF infants [23].  
Breastfeeding may compensate for factors shown to negatively impact the infant’s GI microbiota. Certain strains of bacteria previously isolated from HM produce bacteriocins and others prevent growth of various GI-associated pathogens, which could contribute to the decreased prevalence of illness and other health problems observed in BF vs. FF infants.
 For example, infants delivered by Cesarean section (CS) had different fecal microbiota depending on if they had been breast- or formula-fed. The BF infants achieved numbers of B. bifidum comparable to those of vaginally-delivered (VD) infants earlier than FF or MF infants. Moreover it seems that feeding likely moderates the effects of intrapartum antibiotic prophylaxis (IAP) exposure on microbial colonization. Regardless of feeding mode, IAP-exposed infants, delivered by emergency CS, presented with dybiosis at 3 months. Compared to infants who were not exposed to IAP, these infants had a lower abundance of Bacteroidetes and increased abundance of Firmicutes. These differences, between IAP exposed and non-exposed, remained at 1 yr, but only in infants who were not exclusively breastfed at 3 mos. Moreover, available data suggest that early antibiotic administration is strongly associated with later overweight or obesity. In particular, a significant risk of increased body mass index (BMI) was shown in a large number of children who were prescribed antibiotics in the first 6 months of life or were given these drugs repeatedly during the first 2 years of life. Interestingly, several experimental studies have shown that although animal exposure to antibiotics was limited to infancy, weight gain could emerge later, even in adult age and the metabolic perturbation initially induced by microbiota alteration persisted after the cessation of antibiotic administration, despite the microbiota [24, 25, 26].
These findings further support the notion that breastfeeding may help to reconcile imbalances in an infant’s microbial composition resulting from adverse life events. Differences in the colonization patterns and microbial composition in BF vs. FF infants are proposed to be guided by complex sugars present in HM of each mother, known as human milk oligosaccharides (HMO). These sugars are the third largest component of HM and have been identified in over 200 distinct forms. Abundance of HMOs decreases throughout lactation and maternal genetic differences determine both the composition and orientations of the 5 monosaccharides known to comprise these glycans – Lfucose, D-glucose, D-galactose, N-acetylglucosamine, and N-acetylneuraminic acid. Being resistant to enzymatic hydrolysis, the HMOs pass intact through the infant stomach and upper GI tract to the distal small intestine and colon. HMOs exert prebiotic effects and inhibit the binding of pathogenic bacteria, thereby modulating the infant immune system and shaping the composition of the resident GI microbes. Of particular interest to researchers is the ability of Bifidobacterium to metabolize HMOs. Certain HMO show bifidogenic effects in vitro, selectively stimulating the growth of certain commensal bacteria often identified in BF infants, such as B. infantis, B. bifidum, B. breve and B. longum [27].
Additional studies investigating the long-term impacts of early feeding regimes on GI profiles beyond infancy are also necessary, as existing evidence has demonstrated that infants who were fed with HM for at least 50% of feedings during the first 3 months of life maintained a different fecal microbial composition between 12 and 24 mos than those who had received less than 50% of feedings from HM. The transition toward a more adult-like microbiota is observed during the introduction of solids foods; however, more recent studies suggest that this shift toward a more mature microbiota is more likely attributed to the cessation of breastfeeding, rather than the introduction of complementary foods. For example, continued breastfeeding during the introduction of solid foods seems to support consistent levels of Lactobacillus spp. and Bifidobacterium spp [28].

Fig.2 Sketch of the development of the microbiota from the first inoculum as an infant through continued change, modified by diet, genetics and the environment, throughout life.  (Dominguez-Bello; Development of the Human Gastrointestinal Microbiota and Insights From High-Throughput Sequencing; Gastroenterology 2011;140:1713–1719).

Fig.3 Proposed pathways for the transgenerational cycle of obesity. GWG, pre-pregnancy BMI, development of GDM and/or HFD/WSD can result in maternal gut dysbiosis. This dysbiosis may be directly transmitted to the infant and may cause dysbiosis in the infant gut by causing alterations to SCFA metabolite production, a proinflammatory state, epigenetic alterations and increased energy extraction from ingested nutrients. External influences such as early-life nutrition (breastfeeding vs formula-feeding), mode of delivery and antibiotic treatment may additionally influence the composition of the infant gut microbiome. These changes in gut microbiome function may result in infants born large for gestational age and with excess adiposity, both of which place the child at increased risk of obesity, immune dysfunction and NAFLD later in life. Adulthood obesity during childbearing years then perpetuates the cycle of obesity. DM, type 1 or 2 diabetes. (Taylor K. Soderborg & Sarah J. Borengasser & Linda A. Barbour & Jacob E. Friedman;  Microbial transmission from mothers with obesity or diabetes to infants: an innovative opportunity to interrupt a vicious cycle; Diabetologia (2016) 59:895–906 DOI 10.1007/s00125-016-3880-0).

The impact of diet on gut microbiome
Dietary habits are considered one of the main factors contributing to the diversity of human gut microbiota (Fig.4).  Long-term and short-term dietary intake influences the structure and activity of the trillions of microorganisms residing in the human gut. Gary D. Wu [29] proved that on 10 subjects, the microbiome composition changed detectably within 24 hours of initiating a high-fat/low-fiber or low-fat/high-fiber diet, but that enterotype identity remained stable during the 10-day study.  
Lawrence A . David [30] showed that the short-term consumption of diets composed entirely of animal or plant products, alters microbial community structure and overwhelms inter-individual differences in microbial gene expression. The animal-based diet increased the abundance of bile-tolerant microorganisms (Alistipes, Bilophila and Bacteroides) and decreased the levels of Firmicutes that metabolize dietary plant polysaccharides (Roseburia, Eubacterium rectale and Ruminococcus bromii).
Microbial activity mirrored differences between herbivorous and carnivorous mammals, reflecting trade-offs between carbohydrate and protein fermentation. The increase in the abundance and activity of Bilophila Wadsworthia on the animal-based diet support a link between dietary fat, bile acids and the outgrowth of microorganisms capable of triggering inflammatory bowel disease. In concert, these results demonstrate that the gut microbiome can rapidly respond to altered diet, potentially facilitating the diversity of human dietary lifestyles [31].
Due to overwhelming material abundance, high fat, high sugar and high protein diets are common. Numerous studies have determined that diet and its impact on gut microbiota are closely related to obesity and metabolic diseases. Different dietary components affect gut microbiota, thus impacting gastrointestinal disease occurrence and development.
We do not yet completely understand how the different environments and wide range of diets that modern humans around the world experience has affected the microbial ecology of the human gut, but several recent reviews provide a comprehensive treatment of the subject.
De Filippo et al. [32] analyzed  the fecal microbiota of European children (EU) and that of children from a rural African village of Burkina Faso (BF) where the diet, is rich in fiber (10.0 g/d in 1- to 2-y-old children and 14.2 g/d in 2- to 6-y-old children) , carbohydrate, and nonanimal protein and includes mostly cereals (millet grain, sorghum), legumes (black-eyed peas, called Niébé), and vegetables. Burkina Faso children showed a significant enrichment in Bacteroidetes and depletion in Firmicutes, with a unique abundance of bacteria from the genus Prevotella and Xylanibacter, known to contain a set of bacterial genes for cellulose and xylan hydrolysis, completely lacking in the EU children.  In addition, short-chain fatty acids (P < 0.001) in BF were more abundant than in EU children.  Also, Enterobacteriaceae (Shigella and Escherichia) were significantly underrepresented in BF than in EU children (P < 0.05).  This study enlighted the hypothesis that gut microbiota coevolved with the polysaccharide-rich diet of BF individuals, allowing them to maximize energy intake from fibers while also protecting them from inflammations and noninfectious colonic diseases.
A recent study [33] compared the omnivore diet to the Mediterranean diet, that includes high-level consumption of cereals, fruit, vegetables and legumes. It was detected significant associations between consumption of vegetable-based diets and increased levels of faecal short-chain fatty acids, Prevotella and some fibre-degrading Firmicutes, whose role in human gut warrants further research.  
Conversely, they detected higher urinary trimethylamine oxide levels, risk factor for promoting atherosclerosis in individuals with lower adherence to the MD.  High-level consumption of plant foodstuffs consistent with a Mediterranean diet is associated with beneficial microbiome-related metabolomic profiles [34]. It has been demonstrated that a diet high in saturated fatty acids led to an increased proportion of intestinal Firmicutes and decreased intestinal flora diversity [35]. One study found that converting a low sugar, low fat diet to a high sugar, high fat diet caused a rapid decline in the number of Bacteroidetes in the intestines. Another study also suggested that the number of Bacillus bifidus was reduced in mice fed a high fa diet [36].
In summary, gut microbiota may be an important intermediate link, causing gastrointestinal diseases under the influence of changes in diet and genetic predisposition. A diet that is high in fat, especially high in saturated and transfat, is closely related to obesity, metabolic syndrome and gastrointestinal diseases.

Fig.4 Interaction between diet and gut microbiota affects host metabolism. Dietary manipulation with probiotics and prebiotics alters the composition and metabolic capacity of gut microbiota. Dietary  anipulation in obesity with prebiotics and probiotics changes gut microbiota by favouring bacteria beneficial to the host and enhances the production of short chain fatty acids (SCFAs) – acetate, propionate and butyrate. These result in decreased lipogenesis, reduced inflammation and oxidative stress in liver;decreased adipogenesis, and reduced adipocyte size and number in adipose tissue; increased production of gut hormones and intestinal transit in the large intestine; reduced appetite in the brain. GLP-1: Glucagon like peptide-1, PYY: Peptide YY. (Petia Kovatcheva-Datchary, Tulika Arora;  Nutrition, the gut microbiome and the metabolic syndrome;  Best Practice & Research Clinical Gastroenterology 27 (2013) 59–72).

The role of the gut microbiome
There are broadly three enterotypes, namely: Enterotype 1, which has a high abundance of Bacteroides; Enterotype 2, which has high abundance of Prevotella; and Enterotype 3 which has high abundance of Ruminococcus. The bacteria belonging to Enterotype 1 have a wide saccharolytic potential, as evidenced by the presence of genes that code for enzymes such as proteases, hexoaminidases and galactosidases. In view of these set of enzymatic potential, it appears likely that these organisms derive energy from dietary carbohydrates and proteins. Enterotype 2 behave predominantly as a degrader of the mucin glycoproteins that line the gut mucosal layer. Enterotype 3 also is associated with mucin degradation, in addition to membrane transport of sugars. The enterotypes also possess other specific metabolic functions. For instance, biotin, riboflavin, pantothenate and ascorbate synthesis are more abundantly seen in enterotype 1 while thiamine and folate synthesis are more predominant in enterotype 2 [37, 38].
Nutrients metabolism : The gut microbiota functions as a metabolically active organ and digests dietary components that are indigestible for human cells which can then be absorbed and metabolized by the human body. The interaction between microbiome and the host can be realized by the production of short chain fatty acids (SCFAs) (acetate, propionate and butyrate), hydrogen and methane. Short-chain fatty acids (SCFAs) are derived from the microbial fermentation of undigested dietary fibers in the colon (polysaccharides and oligosaccharides), proteins, peptides, and glycoprotein precursors by the microbiota in the colon and distal small intestine [39].
The SCFAs absorbed through the gut epithelial cells, have strong effects on the energy regulation and the immune system of the host.  The relative absorption of SCFAs by the colon varies between 60–90%, and oxidization of SCFAs can provide energy for colonic mucosa and may contribute up to 5–10% of the total energy in a healthy body. Butyrate and propionate can regulate intestinal physiology and immune function, while acetate acts as a substrate for lipogenesis and gluconeogenesis. As carbohydrate becomes depleted as digesta moves distally, the gut microbiota switches to other substrates, notably  protein or amino acids. Fermentation of amino acids, besides liberating beneficial SCFAs, produces a range of potentially harmful compounds, like ammonia, phenols, p-cresol, certain amines and hydrogen sulfide, play important roles in the initiation or progression of a leaky gut, inflammation, DNA damage and cancer progression [40].
Synthesis of vitamin K and several components of vitamin B is another major metabolic function of the gut microbiota. Members of genus Bacteroides have been shown to synthesize conjugated linoleic acid (CLA) that is known to be antidiabetic, antiatherogenic, antiobesogenic, hypolipidemic and have immunomodulatory properties. The gut microbiota, especially Bacteroides intestinalis, and to a certain extent Bacteroides fragilis and E. coli, also has the capacity to deconjugate and dehydrate the primary bile acids and convert them into the secondary bile acids deoxycholic and lithocolic acids in the human colon [41, 42].
Recent studies have shown that human gut microbiota is also involved in breakdown of various polyphenols (phenolic compounds) that are consumed in the diet, found in a variety of plants, fruits and plant derived products (tea, cocoa, wine). Polyphenols exist as glycosylated derivatives bounded with sugars such as glucose, galactose, rhamnose, ribulose, arabinopyrinose and arabinofuranose and the can’t be adsorbed unless they are biotransformed to active compounds after removal of the sugar moiety by the gut microbiota [43, 44].
The gut microbiota could promote storage of triglyceride in adipocytes through suppression of intestinal expression of a circulating lipoprotein lipase (LPL) inhibitor, the angiopoietin-like 4. Another potential mechanism by which probiotics can counteract the negative effect of obesogenic diet is by interaction with commensal bacteria and altering expressions of microbial enzymes, especially those involved in carbohydrate metabolism or butyrate synthesis pathways [45].
Antimicrobial  function and immunomodulation : The gut microbiota, via its structural components and metabolites, has been shown to induce synthesis of antimicrobial proteins (AMP) such as cathelicidins, C-type lectins, and (pro) defensins by the host Paneth cells. Bacteroides thetaiotaomicron and Lactobacillus innocua appear to be among the key individual species that drive this production. The organism Bacteroides thetaiotaomicron has also been shown to induce expression of the matrix metalloproteinase matrilysin from the Paneth cells, which subsequently cleaves prodefensin to form active defensin. Another example of microbiota-host interaction in providing antimicrobial protection is the capability of Lactobacillus sp. to produce lactic acid, which can augment the antimicrobial activity of host lysozyme by disrupting the outer membrane of the bacterial cell wall [46, 47].The gut microbiota is also involved in the stimulation of the immune system and in providing resistance to pathogens. The gut microbiota, especially Gram-negative organisms like Bacteroides are shown to activate intestinal dendritic cells (DCs), which induces plasma cells in the intestinal mucosa to express secretory IgA (sIgA).  The sIgA that coats the microbiota are predominantly of sIgA2 subclass, which is more resistant to degradation by bacterial proteases. These mechanisms restrict the translocation of the microbiota from the intestinal lumen to the circulation, thereby preventing a systemic immune response. Intestinal microbiota is also essential for the normal development and function of Foxp3+ T regulatory (Treg) cells. Mucosal plasma cells produce secretory IgA upon induction by DCs [48, 49, 50, 51].
Integrity of the gut barrier and structure of the gastrointestinal tract:
The gut microbiota contributes to structural development of the gut mucosa by inducing the transcription factor angiogenin-3, which has been implicated in the development of intestinal microvasculature [52, 53].
Bacteroides thetaiotaomicron is reported to induce expression of the small proline-rich protein 2A (sprr2A), which is required for maintenance of desmosomes at the epithelial villus. Another mechanism that maintains the tight junctions is by TLR2 mediated signaling that is stimulated by the microbial cell wall peptidoglycan. Furthermore, the Lactobacillus rhamnosus GG strain produces two soluble proteins namely p40 and p75 that can prevent cytokine induced apoptosis of the intestinal epithelial cells in an epithelial growth factor receptor (EGFR) and protein kinase C (PKC) pathway dependent manner [54, 55].
The endocannabinoid system is yet another entity that regulates gut microbiota mediated maintenance of the gut barrier function. E.g., the Gram negative bacteria Akkermansia muciniphilia can increase the levels of endocannabinoids that control gut barrier functions by decreasing metabolic endotoxemia [56].
Recent studies have suggested a bacterial role in the development of autoimmune disorders including type 1 diabetes  (T1D). Metagenomics analysis realized on stool sample showed that autoimmune subjects have a functionally aberrant microbiome and altrered gut.  Those data suggest that a consortium of lactate- and butyrate-producing bacteria in a healthy gut induce a sufficient amount of mucin synthesis to maintain gut integrity. In contrast, non-butyrate-producing lactate-utilizing bacteria prevent optimal mucin synthesis, as identified in autoimmune subjects [57].

Childhood Obesity: A Role for Gut Microbiota?
Besides the well known and established causes of obesity like genetic predisposition, excessive intake of high calorigenic diet (fatty food) and lack of exercise which favours storage of calories in the form of fat in adipocytes, recently researchers in the field have shown the contribution and involvement of several other factors like hormonal imbalance; inflammatory cytokines of adipocyte and nonadipocyte origin; adipocytokines like adiponectin, leptin, and resistin, etc., toll-like receptors (TLR) and many others in the genesis of obesity [58]. Several metabolic studies have suggested that imbalances in the intestinal bacterial population may result in obesity, systemic inflammation and metabolic dysfunction (Fig. 5). Gut microflora are involved in obesity through some of their constitutive structural materials and through some of their metabolic end products (SCFAs; butyrate, acetate and propionate) that have been shown to be definitely related with obesity. It is important to say at the beginning that none of the most important enterotypes is purely obesogenic or antiobesogenic since they individually they produce more than one SCFA, each of which possessing opposite actions as metabolites, but the crucial factor to take in consideration  is the  the population ratio [59]. It has been demonstrated that obesity is associated with a reduction in the relative abundance of Bacteroidetes, and that the obese microflora has lower bacterial diversity than lean microflora [60]. Moreover, a high-fat diet increases the intestinal gram-negative to gram-positive bacterial ratio and thus the plasma lipopolysaccharide concentration, which sets the tone for metabolic endotoxemia and thus triggers the low-grade inflammatory state affecting insulin sensitivity. In overweight/obese humans, low faecal bacterial gene richness is associated with more marked overall adiposity and dyslipidaemia, impaired glucose homeostasis and higher low-grade inflammation [61, 62, 63].
High fat diet, both in animals and humans, has been found to alter the gut microbiota composition (more in favour of Gram negative phylum), which in turn increases the production and intestinal permeability of LPS, resulting in its high plasma concentration and development of “metabolic endotoxemia”. Recent observations show that obese person’s microbiota are rich in Prevotellaceae (a subgroup of Bacteroidetes), which is a good source of LPS [64, 65]. Chronic low-grade inflammation found in endotoxemia has been demonstrated to be due to activation of TLR-4 by LPS and dietary saturated fatty acids. TLR-4 activation induces upregulation of common intracellular inflammatory pathways like c-Jun N-terminal kinase and nuclear factor-kappa B in adipocytes and macrophages resulting in development of insulin resistance and increased adiposity [66].
SCFA act in this scenario in different ways. Some important actions of these three SCFAs have been found to be mediated through activation of endogenous free fatty acid receptor (FFAR) like FFAR2 and FFAR3, both these receptors has been demonstrated in adipocytes, epithelial cells and enteroendocrine cells. Activation of these two receptors leads to an increase in expression of satiety hormone polypeptide YY (PYY) and increase in intestinal motility. In addition to the above effect, their activation also increases the expression of leptin in adipocytes. SCFAs, like butyrate and propionate, increase the formation of the gut hormone glucagon-like peptide-1 (GLP-1). It reduces food intake by decreasing appetite. Maximal induction of GLP-1 requires activation of Gpr41, but is not essential [67, 68].
Butyrate has been shown to possess some mixed metabolic effects which include an increase in mitochondrial activity, prevention of metabolic endotoxemia and activation of intestinal gluconeogenesis. These actions are mediated through gene expression and regulation of hormonal activity. Some studies have indicated the antiinflammatory potential of butyrate which may contribute towards a decrease in obesity-associated metabolic complication, because of its capability to increase intestinal barrier function. For instance, F. prausnitzii, a butyrate producer from Clostridium cluster IV, was increased in the nonobese subjects. Butyrate and other short chain fatty acids are known to inhibit inflammation by limiting immune cell migration, adhesion, and cytokine production . In line with this, F. prausnitzii has been found to negatively correlate with inflammatory markers in obese subjects , suggesting that this microbe belonging to the Firmicutes may protect non-obese subjects from inflammation [69, 70].
Butyrate and propionate (beneficial SCFAs) cause weight regulation at least partially by controlling food intake; the action appears to be mediated through their stimulatory effect on the anorexigenic gut hormones. Like butyrate, propionate also possesses favourable some effects in obesity [71, 72]. They are as follows: (1) The SCFA has been found to reduce food intake and regulate body weight, similar to butyrate; (2) It decreases cholesterol synthesis by inhibiting the activity of the enzyme acetyl-CoA synthetase (the enzyme converts acetate to acetyl-CoA), thereby antagonizing the cholesterol increasing action of acetate; (3)  Moreover, propionate has been found to be a precursor for gluconeogenesis in the live [73, 74].
This may decrease the hepatic synthesis of cholesterol because fatty acids necessary for cholesterol synthesis are diverted towards synthesis of glucose (gluconeogenesis; (4) It has been shown that like butyrate, propionate also stimulates the formation of the anorexigenic hormone leptin. Of all the three SCFAs, acetate seems to be more obesogenic than butyrate and propionate [75, 76 , 77].

Fig.5 The role of inflammasomes in metabolic syndrome. During obesity, the NLRP3 inflammasome is activated by obesity-associated DAMPs in multiple tissues and cell types; the resultant pro-inflammatory-induced state often leads to a deterioration in metabolic functions. In adipose tissue, palmitate and ceramides activate the NLRP3 inflammasome in infiltrating macrophages, which leads to an enhancement of insulin resistance. In addition, caspase-1 activation through an unknown sensor protein regulates adipocyte differentiation and fatty acid oxidation. In the pancreas, IAPP and increased mitochondrial ROS production activate the NLRP3 inflammasome in mmLDL-primed macrophages and β cells, respectively. The increased levels of IL-1β in pancreatic islets result in increased β-cell death and decreased insulin production. Minute cholesterol crystals in early atherosclerotic lesions activate the NLRP3 inflammasome in mmLDL-primed macrophages, promoting inflammatory-cell infiltration and increased atherosclerosis progression. (Till Strowig, Jorge Henao-Mejia, Eran Elinav & Richard Flavell ; Inflammasomes in health and disease;  278; Nature, 481; 2012).

Dietary manipulation in obesity: Probiotics and Prebiotics
Probiotics are defined as the live microorganisms that, when administered in adequate amounts, confer health benefits to the host. Probiotic supplementation results in enrichment of the probiotic species in intestinal contents. It also results in alterations in the composition of gut microbiota and microbial metabolites, like SCFAs, in both murine models and humans [78].
Prebiotics are defined as dietary ingredients that promote ‘the selective stimulation of growth and/or activity (ies) of one or a limited number of microbial genus (era)/species in the gut microbiota that confer (s) health benefits to the host [79, 80]. Dietary fibres are known to exhibit prebiotic effects as they are utilized by specific bacteria, resulting in their proliferation, which are considered beneficial to the host.
Most currently used probiotics belong to bifidobacteria, lactic acid bacteria (LAB), dairy propionibacteria, yeasts (Saccharomyces boulardii), Bacillus, and the gram-negative Escherichia coli strain Nissle 1917. LAB represent a heterogeneous group of microorganisms broadly present in the diet, particularly by the use of non-human strains in the fermentation of dairy products being also normal inhabitants of the gastrointestinal and urogenital tract. Most of them are members of the phylum Firmicutes, while Bifidobacterium, also considered as lactic-producing bacteria, belong to Actinobacteria phylum.
Probiotic administration is frequently associated with important shifts in gut bacterial composition, along with beneficial effects on metabolism and inflammatory tone. Probiotic administration has been shown to stimulate the immune response, improve lactose tolerance, help prevent diarrhea, have an anti-inflammatory effect and even restore obesity-linked gut dysbiosis [81, 82, 83]. Fecal microbiota transplantation showed good results for treating Clostridium difficile infection (CDI), and  the three most relevant studies published in the last years on patients affected by Ulcerative colitis, not responding to conventional therapy,   and treated with retention enema, showed a remission ranging from 3 months to several years [84, 85, 86].
Treatment with VSL#3 in patients with active UC, not responding to mesalamine therapy, was proved to a trend toward a remission/response rate of 77% without presenting any adverse events [87].
Given the relationship between obesity-related disorders and gut homeostasis, probiotics may be of interest to supplement the limited arsenal of therapies against the metabolic syndrome.
In the context of obesity and metabolic disorders, probiotic supplementation may help to reduce hyperphagia [88], improving control of weight gain, fat mass loss and glucose tolerance. On the contrary, such positive effects could also be obtained without modulation of caloric intake, as demonstrated by most of the reported studies [89, 90].
Probiotics have been shown to reduce adipocyte size in different adipose depots, which is considered an important parameter in assessing their anti-obesity potential [91].
The putative mechanisms put forth are increased fecal excretion of neutral sterols and bile acids, decreased lymphatic absorption of triglycerides, phospholipids and cholesterol, or increased lipolysis. VSL#3 was demonstrated to promote the release of the hormone GLP-1, resulting in reduced food intake and improved glucose tolerance. By activating the G-protein-coupled receptors GPR41 and GPR43 on intestinal epithelial cells, SCFAs stimulate peptide YY (PYY) and glucagon-like peptide (GLP)-1 secretion. In turn, these hormones may suppress gut motility and retard intestinal transit, allowing for greater nutrient absorption [92].
Prevention and management of obesity is proposed to begin in childhood when environmental factors exert a long-term effect on the risk for obesity in adulthood. Thus, identifying modifiable factors may help to reduce this risk. With advancing knowledge of how probiotics interact with the gut microbiome, there is an increasing interest in exploring the anti-obesity potential of probiotics [93].  
One double-blind study focused on the impact of perinatal probiotic intervention , 4 weeks before expected delivery and 6 months postnatally (Lactobacillus rhamnosus GG ATCC 53103 vs palcebo) in 113 children, observed for 10 years, regards the development of overweight, obesity and child’s BMI. "e;€¨The probiotic intervention showed to modify the growth pattern of the child by restraining excessive weight gain during the first years of life [94].
The genus Bifidobacterium, affecting both the quantity and quality of the microbiota during the first year of life, was shown to be higher in number in children who remained normal weight at 7 y than in children developing overweight as reported in the prospective follow up study of Kalliomaki et al. [95, 96].
As regard prebiotics, several in vitro studies have shown that HMOs serve as the primary substrate for the growth of Bifidobacterium and other bacteria. Furthermore, several studies have demonstrated correlations between several bacterial genera present in BF infant feces, with the HMO in their mother’s milk or present in the infant feces. The most studied prebiotic addition to infant formula is a 9:1 mixture of short-chain galactooligosaccharides (scGOS) and long-chain fructooligosaccharides (lcFOS) [97]. Investigators have identified significant clinical impacts of these prebiotics on the immune and metabolic development of infants. Infants who consume formula supplemented with this mixture have a lower fecal pH and exhibit a fecal SCFA profile and stool characteristics comparable to that of BF infants. Findings have also shown that infants exposed to scGOS and lcFOS to require fewer doses of antibiotics and have a decreased incidence of infection likely due to an increased colonization resistance to enteric pathogens. The other prebiotics added to infant formula individually or in combination include, GOS, FOS, oligofructose and inulin, polydextrose (PDX), lactulose (LOS) and acidic oligosaccharides (AOS). Prebiotics are resistant to gastric acidity and enzymatic hydrolysis in the upper gastrointestinal tract and enter the colon intact, where they are metabolized by colonic microbiota. Shortchain prebiotics, such as FOS and GOS, are mainly fermented in the ascending colon, while longer-chain prebiotics, like PDX and inulin, are fermented along the entire colon [98, 99]. Most studies showed that supplementation of prebiotics to infant formula increased the numbers of beneficial bacteria, mainly Bifidobacterium and sometimes Lactobacillus. In the context of obesity, the use of relatively new prebiotics such as arabinoxylan (AX) and rabinoxylan oligo-saccharides (AXOS) may be promising candidates to counteract related metabolic disorders, since AX and AXOS have been linked to adiposity reduction  and lower metabolic endotoxemia  in obese mice, respectively [100].
Another growing concept is to genetically engineer bacterial strains in order to reinforce a pre-existing probiotic capacity or to increase their effectiveness. Duan et al. [101] recently reported the successful application of an engineered probiotic that secretes the inactive full-length form of GLP-1 to reprogram intestinal cells into glucose-responsive insulin-secreting cells for the treatment of type 1 diabetes.
Another interesting potential strategy is the genetic modification of the probiotic E. coli Nissle 1917 to produce N-acylphosphatidylethanolamines, which is converted quickly after meals into potent appetite-suppressing lipids, know as N-acylethanolamines [102].

Conclusions
Diet and lifestyle are crucial factors influencing the development and progression of obesity. Recent insights have examined obesity aetiology with a new perspective and found that our own microbiota might be involved in the development of these disorders. The infant GI microbiota undergoes rapid and profound changes during the first year of life. During this process, diet plays a predominant role over other environmental factors in shaping the microbial composition.
Most of medications for treatment of obesity are taken out the production because of their adverse. One of the potential ideal strategy for obesity treatment may be manipulation with gut microbiota. Probiotics are food supplements that confer beneficial effects under various clinical conditions inclusive of atherogenesis, allergy, and inflammatory bowel diseases. However, the widespread use of probiotics against obesity and
diabetes is lacking , primarily because of insufficient mechanistic insight and a paucity of efficacy data in small animal models. More controlled human and animal studies are necessary to clarify these complex interactions.


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geneticapediatrica.it trimestrale di divulgazione scientifica dell'Euromediterranean Paediatric Foundation
Legge 7 marzo 2001, n. 62 - Registro della Stampa Tribunale di Messina n. 3/09 - 11 maggio 2009

Abstract
Proteus syndrome is a very rare mosaic condition characterized by progressive and asymmetric overgrowth of tissues due to a somatic activating mutation of the akt1 gene. Clinical manifestations include distinct cutaneous features, like cerebriform connective tissue nevi, epidermal nevi, vascular malformations, and adipose abnormalities. Other unusual features include bullous lung alterations; specific neoplasms; a facial phenotype associated with intellectual disability and/or seizures, and/or brain malformations; and deep vein thrombosis, resulting in premature death. Prognosis of affected patients may be complicated by premature death, mostly due to pulmonary embolism and respiratory failure.

Key words: proteus syndrome (ps); disproportionate overgrowth; hemihyperplasia; asymmetry; hyperostoses; cerebriform connective tissue nevus.

Introduction
Proteus syndrome (ps) is part of overgrowth syndromes and it is due to a mosaic, activating akt1 mutation (c.49g>a, p.glu17lys)1.  PS is a rare complex disorder with multisystem involvement and great clinical variability, first described in 19792. In 1983, rudolf wiedemann, a german pediatrician, named it proteus syndrome after a greek sea-god, who could change his shape at will assuming many forms to escape capture. This name was given to represent the variable clinical manifestations seen in the first patients identified with this syndrome and the morphological changes of its presentation and evolution3, 4.  It is an extremely rare syndrome with an estimated prevalence of approximately 1:1.000.000, being more common among males at a ratio of 1:9:13, 4. This disease is characterized by a mosaic distribution, sporadic occurrence, and progressive course. Clinical features may be present at birth but typically develop over time, starting between 1 and 18 months of age. The postnatal, progressive, and asymmetrical overgrowth occurs in a mosaic pattern. Bone, connective tissue, fat, central nervous system, eye, spleen, thymus, and colon are commonly involved tissues5. The complications of ps include hyperostosis, cerebriform connective tissue progressive, skeletal deformities, benign and malignant tumors, capillary vascular malformations and deep venous thrombosis with pulmonary embolism. The syndrome has an incidence of less than 1 per 1, 000.000 live births and is estimated that 120 individuals with ps are currently alive worldwide6, 4.
Newborns with proteus syndrome have few or no signs of the condition, and overgrowth becomes apparent between 6 and 18 months of life, getting more severe with age

Clinical manifestations
Dermatologic findings
Cerebriform connective tissue nevus: the cerebriform connective tissue nevus found in some patients is not obligatory. They occur most frequently on the plantar surfaces of the feet (fig. 1), but may also occur on the hands, abdomen, and nose. They are characterized by highly collagenized connective tissue. Epidermal nevi and other skin lesions: In Proteus syndrome, such nevi are evident in early life and may occur on the neck (Fig. 2), trunk, or extremities. Histopathological findings consist of acanthosis and hyperkeratosis. Patchy areas of dermal hypoplasia and hypopigmentation have also been observed 8.

Fig.1 New England J Med 2011; 365:611-619

Fig.2 From Cohen and Hayden

Dysregulated adipose tissue
Overgrowth of adipose tissue is common in infancy but it can continue to appear in novel locations throughout childhood and into young adulthood. Similarly, many individuals with ps experience marked regional lipoatrophy, and many manifest both regional lipomatous overgrowth and lipoatrophy.

Pulmonary abnormalities
Bullous lung abnormalities have been discussed by several authors. It is known also pulmonary venous dilatation 4, 9.

Disproportionate asymmetric overgrowth and skeletal abnormalities (table 2)
Overgrowth in proteus syndrome is disproportionate, asymmetric, distorting, and relentless11. Skeletal abnormalities include kyphoscoliosis abnormal vertebral bodies (asymmetrical vertebral body overgrowth), segmentation defects, focal calvarial thickening, limb overgrowth, premature degenerative changes, macrodactyly and sometimes nondevelopment of peripheral bones11, 3. In proteus syndrome, the long bones are overgrown with abnormally thin cortices and deficiency of the overlying soft tissue 12. (fig.3)

Tab. 2. Skeletal abnormalities (Adapted from Jamis-Dow et al. and Turner et al.)

Fig.3 Asymmetric overgrowth of legs with thin cortices and deficiency of the overlying soft tissue. From Cohen.

Neoplasms
Only lipomas are common. Other reported neoplasms include monomorphic adenoma of the parotid gland; cystadenomas of the ovary; testicular tumors; meningiomas and mesothelioma 13.

Facial phenotype
The facial phenotype is present in only a minority of patients and may be associated with cognitive deficits and, in some cases, seizures and brain malformations. Manifestations include dolichocephaly, long face, minor downslanting palpebral fissures and/or minor ptosis, low nasal bridge, wide or anteverted nares, and an open mouth at rest3, 8. (fig.4)

Fig.4  Evolution of facial abnormalities in a patient with Proteus syndrome. Left: Age 1 month. Center: Age 5 months. Right: Age 5½ years. Note connective tissue nevus on left side of nose. From Cohen

Vascular malformations
Vascular malformations can be profound and include vascular ectasia, haemangiomata, lymphangiomata and varicosities14. The vascular ectasia may be progressive (figures 9–14). Deep-vein thrombosis leading to pulmonary embolism is the commonest cause of death in this group of patients, possibly related to venous stasis in grossly ectatic vessels 15.

Neurological manifestations
Cerebral arteriovenous malformations, abnormal grey–white matter differentiation and hydrocephalus are commonly seen. Schizencephaly, spinal lipomatosis and perineural cysts have also been described 16. (fig.5, 6)

Fig. 5, 6. (5) Asymmetry of the cerebral hemispheres and lateral ventricles with abnormal grey–white matter differentiation (arrow) and areas of macrogyria in the left cerebral hemisphere. MRI image, axial T2, repetition time 5700 ms, echo time 98 ms. M J Kaduthodil et al. (6)  Asymmetrical calvarial thickening (vertical arrow) and arteriovenous malformation (horizontal arrow). MRI image, axial T2, repetition time 5700 ms, echo time 98 ms. M J Kaduthodil et al.

Diagnosis and differential diagnosis
The diagnostic criteria for proteus syndrome are shown in table 117. To meet diagnostic criteria, the patient must meet the single criterion from category a, two criteria from category b, or all three criteria from category c3. Biesecker’s group demonstrated that a mosaic activating akt1 mutation, c.49g>a, p.glu17lys was the cause of proteus syndrome. They examined 29 patients, all of whom met the clinical diagnostic criteria (table 1). Of these, 26 had the akt1 mutation. In the remaining three patients the mutation has not been identified. Lindhurst et al. Suggest that the clinical criteria for proteus syndrome alone may be sufficient for the diagnosis (table 1) without identifying the mutation, except in early cases when the clinical findings are not definitive1. Molecular genetic testing may be useful to confirm the diagnosis in individuals who meet clinical criteria and to establish the diagnosis in individuals in whom the clinical findings are ambiguous or mild. The differential diagnosis of  ps must be made with other hamartomatous disorders such as klippel-trenaunay-weber (ktws; asymmetry in one limb and hemangioma), maffucci disease (enchondromatosis and hemangioma)18, ollier's disease (enchondromatosis), neurofibromatosis type i (macrocephaly, café-au-lait spots, subcutaneous neurofibromas), bannayan-zonana syndrome (macrocephaly, craniofacial abnormalities), hemihyperplasia and multiple lipomatosis syndrome (hhml) and other disorders that present with hemihyperplasia19. In ps, the abnormal growth is asymmetric, distorting, relentless, and occurred at a faster rate compared to the rest of the body. Furthermore, ps is associated with irregular and disorganized bone, including hyperostoses, hyperproliferation of osteoid with variable calcification, calcified connective tissue, and elongation of long bones with abnormal thinning. In contrast, non-proteus cases displayed overgrowth that is asymmetric but grew at a rate similar to the growth found in unaffected areas of the body. Also, the overgrowth in non-proteus cases is associated with normal or enlarged bones together with ballooning of the overlying soft tissues20. The two conditions which must be primarily distinguished from the proteus syndrome are klippel–trenaunay–weber syndrome (ktws) and neurofibromatosis type 1. In ktws, the changes of soft tissue growth and hypertrophy are present at birth and are usually severe. The key distinction between the two conditions is presence of bone involvement and progressive hypertrophy in ps, which is absent in ktws11. Neurofibromatosis is usually distinguished from ps by the presence of the clinical triad of pigmented skin nodules (cafe´-au-lait spots), lisch nodules and axillary freckling15. The proteus syndrome must also be differentiated from cloves syndrome. Cloves is composed of congenital lipomatous overgrowth, vascular malformations, epidermal nevi, and skeletal anomalies. While some of these abnormalities may develop over time, a significant number of them may be present at birth21.  It is caused by activating mutations in pik3ca22.

Evaluations following initial diagnosis
To establish the extent of disease in an individual diagnosed with proteus syndrome, the following evaluations are recommended [tosi et al 2011]:
•    Detailed and comprehensive (general, spine, and hand) orthopedic evaluation
•    Skeletal survey as a baseline study of the extent and severity of overgrowth
•    Ct imaging, possibly with three-dimensional reconstruction for patients with significant scoliosis. As the vertebral bodies are commonly progressively deformed, this study can be very helpful for surgical planning.
•    Pulmonology consultation, pulmonary function testing, and high-resolution computed tomography of the chest for patients with signs or symptoms compatible with bullous pulmonary disease

Management
As with any complex and multisystem disorder, patients with ps benefit from a coordinated and multidisciplinary clinical approach. Overgrowth is an ongoing issue for many patients with proteus syndrome. The management is complex and highly dependent on the nature of the overgrowth, which can vary substantially among patients. The approaches are diverse and include various orthopedic procedures to delay or halt linear bone growth; correction of skeletal deformities such as scoliosis22, 8. For overgrowth of tubular bones, epiphysiostasis and epiphysiodesis should be the mainstays of management. The skeletal overgrowth of ps can result in significant biomechanical and functional compromise. Because of this, ongoing and comprehensive rehabilitation medicine care, including physical and occupational therapy, is important for many patients. In addition, many patients with ps develop substantial needs for custom-designed footwear or orthotics due to leg-length inequality or plantar cctns. Management of the overgrowth of adipose tissue is challenging because the areas of adipose overgrowth are not encapsulated and discrete (in contrast to lipomas) and, therefore, can be difficult to resect and commonly regrow after surgical debulking. The authors generally recommend open surgical approaches over liposuction because the highly vascularized lipomatous overgrowth in some patients can result in hemorrhaging that is difficult to control and/or chronically weeping lymphatics. (deep vein thrombosis (dvt) and pulmonary embolism (pe). The most urgent and life-threatening complication of proteus syndrome can be dvt and pe [slavotinek et al 2000]. The rarity of this problem in the general pediatric population can result in a delay in diagnosis.
Any organ or tissue can be affected and, thus, the secondary complications are highly variable.
Surveillance: monitoring should be tailored to individual patients’ presentations; routine monitoring for evidence of tumor development is by medical history and physical examination; periodic imaging is not indicated 22.

References
1.    Lindhurst mj, sapp jc, teer jk, johnston jj, finn em, peters k, et al. A mosaic activating mutation in akt1 associated with the proteus syndrome. N engl j med. 2011;365 (7):611–9.
2.    Cohen mm jr, hayden pw. A newly recognized hamartomatous syndrome. Birth defects orig artic ser 1979;15:291–6.
3.    Biesecker lg, happle r, mulliken jb, weksberg r, graham jm, jr, viljoen dl, et al. Proteus syndrome: diagnostic criteria, differential diagnosis, and patient evaluation. Am j med genet. 1999;84:389–95.
4.    Biesecker l. The challenges of proteus syndrome: diagnosis and management. Eur j hum genet. 2006;14:1151–7.
5.    Cohen mm., jr proteus syndrome: an update. Am j med genet c semin med genet. 2005;137c:38–52.
6.    Cohen jr mm. Proteus syndrome review: molecular, clinical and pathologic features. Clin genet. 2014;85 (2):111–9.
7.    Thomason jl, abramowsky cr, rickets rr, culbertson jh, clifton ms, shehata bm. Proteus syndrome: three case reports with a review of the literature. Fetal pediatr pathol. 2012;31 (3):145–53.
8.    Cohen m. Michael jr. Proteus syndrome review: molecular, clinical, and pathologic features. Clin genet 2014: 85: 111–119. John wiley & sons a/s. Published by john wiley & sons ltd, 2013.
9.    Lim gy, kim oh, kim hw et al. Pulmonary manifestations in proteus syndrome: pulmonary varicosities and bullous lung disease. Am j med genet 2011: 155a: 865–869.
10.    Cohen mm jr. Proteus syndrome. In: cohen mm jr, neri g, weksberg r, eds. Overgrowth syndromes, vol. Chapter 9. New york: oxforduniversity press, 2002: 75–110.
11.    Jamis-dowca, turner j, biesecker lg, choyke pl. Radiologic manifestations of proteus syndrome. Radiographics 2004;24:1051–68.    
12.    Biesecker lg, peters kf, darling tn et al. Clinical differentiation between proteus syndrome and hemihyperplasia: description of a distinct form of hemihyperplasia. Am j med genet 1998: 79: 311–318.
13.    Gordon pl, wilroy rs, lasater oe, cohen mm jr. Neoplasms in proteus syndrome. Am j med genet 1995: 57: 74–78
14.    Hoeger ph, martinez a, maerker j, harper ji. Vascular anomalies in proteus syndrome. Clin exp dermatol 2004;29:222–30.
15.    M j kaduthodil, d s prasad, a s lowe, a s punekar, s yeung, and c l kay. Imaging manifestations in proteus syndrome: an unusual multisystem developmental disorder. The british journal of radiology, 85 (2012), e793–e799.
16.    Dietrich rb, glidden de, roth gm, martin ra, demo ds. The proteus syndrome: cns manifestations. Ajnr am j neuroradiol 1998;19:987–90.
17.    Turner jt, cohen mm jr, biesecker lg. A reassessment of the proteus syndrome literature: application of diagnostic criteria on published cases. Am j med genet 2004: 130a: 111–122  .
18.    Elsayes km, menias co, dillman jr, platt jf, willatt jm, heiken jp. Vascular malformations and hemangiomatosis syndromes: spectrum of imaging manifestations. Ajr am j roentgenol. 2008;190:1291–99
19.    cresio alves, angelina x. Acosta and maria betânia p. Toralles. Proteus syndrome: clinical diagnosis of a series of cases. Indian j endocrinol metab. 2013 nov-dec; 17 (6): 1053–1056.
20.    Joyce t. Turner, m. Michael cohen, jr., and leslie g. Biesecker. Reassessment of the proteus syndrome literature: application of diagnostic criteria to published cases. American journal of medical genetics 130a:111–122 (2004)
21.    Kurek kc, luks vl, ayturk um et al. Somatic mosaic activating mutations in pik3ca cause cloves syndrome. Am j hum genet 2012: 90: 1108–1115.
22.    Leslie g biesecker, md and julie c sapp, scm, cgc. Proteus syndrome. Genereviews, august 9, 2012

geneticapediatrica.it trimestrale di divulgazione scientifica dell'Euromediterranean Paediatric Foundation
Legge 7 marzo 2001, n. 62 - Registro della Stampa Tribunale di Messina n. 3/09 - 11 maggio 2009

Abstract

Isolated vascular anomalies and syndromes associated with vascular anomalies   vary in location, type, and clinical severity of the phenotype. In recent years, the identification of the genetic factors contributing to these anomalies have been elucidated.
Vascular anomalies have an incidence estimated around 1/10 000. (Boon, L.M., 1994) and subdivided into vascular tumors (mainly the hemangiomas, of unknown etiology) and vascular malformations (CVM) are thought to be due to defects in these pathways (Mulliken, J.B. et al., 1982). The two entities are diseases completely different, even though both are grouped under the common classification of vascular abnormalities. The CVM constitute errors of morphogenesis, as well as structural abnormalities, while hemangiomas are vascular tumors. Most malformations are present at birth and grow proportionately with the child.The etiopathological genetic defects have been elucidated for some of these, and they are discussed here with relevant functional data and development of small animal models. The clinical history allows to differentiate in most cases between cancer and the CVM, although not always be available history clear and appropriate.

Keywords: Klippel-Trenaunay syndrome; Weber syndrome; Sturge-Weber syndrome; capillary malformation-arteriovenous malformation; hereditary haemorrhagic telangiectasia; venous malformations

Childhood hemangiomas and congenital hemangiomas 
The childhood hemangiomas (HOI)  are the most common form of pediatric vascular proliferative tumors usually not present at birth, appearing at a later stage in childhood. These tumors have a two-step process of proliferation and regression: grow rapidly and disproportionately compared to the growth of the child, then reach a phase of maximum growth to pass then to a phase of involution. The majority of HOI reaches a stage of atrophy by the age of 10 years. It is likely that the HOI will originate from placental tissue. From the histological point of view, these lesions have a positive staining for glucose transporter-1 (GLUT-1) during all stages of growth and involution. (North PE et al., 2001)
The diagnosis of HOI is usually clinical, but ultrasound is necessary. Most hemangiomas are hypoechoic, although according to studies up to 18% of hemangiomas is hyperechoic. It is important to differentiate between HOI and congenital hemangiomas, with the latter constituting a small group of pediatric vascular tumors. The congenital hemangiomas grow completely utero, they are present at birth and therefore do not go through a phase of postnatal proliferation, as it occurs in the HOI. Histologically sight congenital hemangiomas do not stain with GLUT-1.
Normally it is possible to identify two subtypes of congenital hemangiomas: hemangiomas RICH (Rapidly involuting congenital hemangioma - fast congenital hemangiomas involution) and hemangiomas NICH (non-involuting congenital hemangioma - congenital hemangiomas not regressive).
The first pass through a phase of spontaneous involution in the first year of life, while the latter persist for a lifetime. Calcifications have not reported in infantile hemangioma but have been identified in a subset of congenital hemangiomas. (Gorincour G et al., 2005).

Table 1. Hemangiomas versus Vascular Malformations. (From Mulliken, J.B. and Glowacki, J. 1982)

Congenital vascular malformations (CVM)
The CVM are structural abnormalities of the vascular system and are NOT cancer. These injuries are result of an interruption of development in different stages of embryogenesis. They are always present from birth and constantly growing in line with the systemic growth of the child. In contrast, the HOI grow disproportionately compared to the growth of the child. The CVM does not regress spontaneously and remain throughout the course of the patient's life, such as residues of embryological tissue.
The CVM can affect vessels of any type and in general can be classified as follows (Table 1- Figure 2):
• arteriovenous malformations (AVM)
• venous malformations (VM)
• lymphatic malformations (LM)
• capillary malformations (CM)

Table 2. Classification of vascular anomalies (from Blei F et al., 2016)

Fig. 2. Mechanism of development of vascular malformations from fetal capillary plexus. (From Blei F et al., 2016)

Arteriovenous malformations (AVM)
The AVM are structural abnormalities involving a high flow interfacial communication abnormal between the arterial system and venous system and its full maturity can manifest itself in the form of mass extended button and hot. The increase in size is due the increase in abnormal vessels, in addition to hypertrophy associated soft tissue. The presence of hemorrhage can be a typical element of the presentation of an occult AVM. In infants the AVM may manifest in the form of pale pink skin color and can be difficult to differentiate with respect to a CM. The bone hypertrophy and / or of the normally associated soft tissue may entail an increase of the dimensions (length and width) of the involved side.
Venous malformations (VM)
Venous malformations (VM) are the most common abnormality of development dependent on the venous system. It is a fault caused by an interruption of the development of the venous system in different stages of embryogenesis.
VMs can be classified as follows:
1) VM extratronculari: lesions detectable in most tissues that may arise in the form of lumps of dilated veins or venous injury.
2) VM truncal: lesions in the form of aplasia, hypoplasia, obstruction, dilation, duplication or aneurysm.
Lymphatic malformations (LM)
The nomenclature for LM is still confusing and still are widely used obsolete terms like lymphangiomatosis, lymphangiectasia or lymphatic dysplasia to describe several related diseases.  When LM only involving the skin and subcutaneous tissue, the prognosis is usually favorable. When the lesions involving the upper airways, viscera or bone, the prognosis is generally poor, with a significant mortality rate.
Capillary malformation (CM)
The CM represent the most common type of VCM. Descriptive terminology used in past with reference to the CM was that of "Port wine stain" or "nevus flammeus". As congenital malformations of superficial blood vessels of the dermis, the CM are present at birth and grow in size in line with the child's development. The lesions persist for life and do not have any tendency to involution. The clinical progress of the CM. It varies depending on the anatomical location of the lesion. A significant number of dysmorphic syndromes was associated with CM.

Syndromic vascular malformations
Syndromic vascular malformations (Wassef M et al., 2015)include Klippel-Trenaunay (Capillary malformations, lymphatic or venous with hypertrophy of the limbs), Parkes-Weber syndrome (Similar to Klippel-Trenaunay with arterio-venous shunt), the Sturge-Weber syndrome (Malformation of the facial capillaries in triplets distribution, leptomeningeal angiomatosis, glaucoma and seizures), the Blue-Rubber-Bleb-Nevus (BRBNS syndrome, characterized by small generalized venous malformations), the Proteus syndrome (vascular malformations, lipoipoplasia trunk, scoliosis, cerebriform plantar surfaces, snow, partial gigantism and abnormalities of the fingers), the disease Ollier / Maffucci syndrome (hemangioendothelioma, enchondromatosis), the HHT (AVM multifocal), Gorham's syndrome (with lymphangiomatosis osteolysis), as well as several anomalies and syndromes or lymphatic linfedematose. Syndromes related to PTEN vascular anomalies include Cowden syndrome and syndrome Bannayan-Riley-Ruvalcaba; these syndromes have clinical features and radiological specifications. Patients with PTEN syndromes are at greater risk of malignant lesions (For example, thyroid, breast, colon, brain, genitourinary tract), which may occur earlier than normal in the general population. About 30% of patients with facial segmental hemangiomas may be suffering from syndrome PHACES (MIM 606519) [Injury to the posterior fossa or other structural lesions SNC, segmental hemangiomas, arterial anomalies, cardiac abnormalities, eye abnormalities or dependent deformation of the sternum or the median line]. The assessment provides: Cerebral magnetic resonance imaging with or without contrast and MRA brain, neck and upper chest, ophthalmologic examination, heart and chest, as well as thyroid function tests.

Table 3: Vascular malformation syndromes
 

Hereditary hemorrhagic telangiectasia
Ereditary hemorrhagic telangiectasia (HHT) (MIM 187300 and 600376) also known as Osler-Weber-Rendu syndrome, is an autosomal dominant disorder with an incidence around 1/10 000 (Abdalla, S.A.2006)  that is characterized by multiple arteriovenous malformations (AVMs) involving the skin, mucosal surfaces, and internal organs. HHT has an age-dependent penetrance and usually initially presents with recurrent epistaxis followed by the characteristic telangiectasias of the face, oropharynx, and hands over time. Patients often have vascular malformations that involve their lungs, brain, spinal cord, and gastrointestinal tract as well, which are the main causes of morbidity in patients with HHT (Guttmacher Ar et al., 1995). The sequelae of visceral organ involvement include ischemic stroke, cerebral bacterial abscesses, intracranial hemorrhage, chronic hypoxia, dyspnea with exertion, pulmonary hypertension, high output heart failure, gastrointestinal bleeds and liver failure.
At least four loci have been associated with HHT: HHT1 on 9q33–34, with mutations in endoglin (ENG) (McAllister, K.A., 1994 ), HHT2 on 12q11–14, with mutations in the activin receptor-like kinase 1 (ALK1) (Johnson, D.W., 1996), HHT3 on 5q (Cole et al., 2005) and HHT4 on 7p14 (Bayrak-Toydemir, 2006).Several genetic mutations (most commonly affecting one of two genes in the transforming growth factor-beta/bone morphogenetic protein [BMP] signaling family: endoglin or activin receptor-like kinase-1 [ACVRL1], or less commonly SMAD4 [Mothers Against Decapentaplegic, Drosophila Homolog of 4] or growth/differentiation factor [GDF2]; BMP9]) have been identified in this disorder, which has been recently reviewed by McDonald et al (McDonald et al., 2015).

Sturge–Weber syndrome
Sturge–Weber syndrome is a rare sporadic neurocutaneous syndrome the hallmark of which is a facial port-wine stain involving the first division of the trigeminal nerve, ipsilateral leptomeningeal angiomata and angioma involving the ipsilateral eye. The incidence of SWS is ~1:50, 000 infants, with no significant difference between males and females. (Di Rocco et al., 2006).The characteristic facial port-wine stain, involving the first branch of the trigeminal nerve and the embryonic vasculature distribution in this area, leads to several ocular complications of the anterior segment and can involve the eyelids and conjunctiva. The posterior segment of the eyes is also affected with diffuse choroidal hemangiomas. Our understanding of the disease process has vastly improved since it was first described in 1879, with recent identification of an activating somatic mutation in the GNAQ gene. A somatic mutation in the GNAQ gene in port-wine stains of patients with and without SWS has been identified. (Shirley MD et al., 2013).Sturge–Weber syndrome is marked by a variable but usually progressive course in early childhood characterised by seizures, stroke-like episodes, headaches, neurological and cognitive deterioration, hemiparesis, glaucoma and visual field defects. More recently, the increased prevalance of otolaryngological, endocrine and emotional–behavioural issues have been established. According to the clinical manifestation, SWS is classified into four types: 1) presence of brain and facial angioma, with or without glaucoma, 2) PWS without brain involvement, with or without glaucoma, 3) isolated brain angioma, usually without glaucoma, and 4) type 1 associated with systemic manifestation such as tuberous sclerosis (Table 2).Roach ES., 1992)
Neurophysiology and neuroimaging studies provide information regarding the evolution of changes in Sturge–Weber syndrome over time. Early recognition and aggressive management of symptoms remains cornerstone in the management of this syndrome. Two recent studies have challenged the previously accepted notion that CMs over the distribution of the trigeminal nerve places patients at highest risk of SWS (Dutkiewicz AS et al 2015, Waelchli R et al., 2014)
 

Table 4  Classification of Sturge–Weber syndrome

Maffucci syndrome
Maffucci syndrome, a rare sporadic form of enchondromatosis, is characterized by hemangiomas and multiple enchondromas, benign cartilaginous tumors that arise near growth plates. First described by Angelo Maffucci in 1881, Maffucci syndrome is an exceedingly rare disease with less than 250 cases reported. (Gao H et al., 2013) Maffucci syndrome affects both sexes and has no geographical or ethnic predisposition. The disease usually begins between the 1st and the 5th year of life. In 25% of cases, the clinical symptoms are present at birth or appear in the first year.  Considered a subtype of enchondromatosis, Maffuccisyndrome presents with multiple enchondromas that are associated with soft tissue hemangiomas. (Fanburg JC et al., 1995) Enchondromas are benign growths commonly in the small bones of the fingers and toes, long tubular bones, and flat bones such as the pelvis. They grow in close proximity to the growth plate cartilages and are therefore thought to result from abnormal regulation of proliferation and terminal differentiation of chondrocytes. (Hopyan S, et al 2002) Patients with enchondromatoses have an increased risk of chondrosarcomatous transformation, seen in up to 15%–30% of cases. ( Lewis et al., 1973, Kaplan et al., 1993)
The capillary malformations usually present on the distal end in the form of subcutaneous nodules prominent blue-dark, irregularly shaped, but can also be located elsewhere. They may be associated lymphatic and venous malformations. The skeletal and vascular lesions are usually asymmetrical and may be progressive. About 30-40% of enchondromas evolves into chondrosarcoma. The syndrome is associated with other benign or malignant tumors (goiter, parathyroid adenoma, pituitary adenoma, adrenal cancer, ovarian cancer, breast cancer, or astrocytoma; see these terms). The etiology of Maffucci syndrome is not completely known. The disease appears to be associated with mesodermal dysplasia early in life. No family transmission was observed. The diagnosis is based on clinical and radiographic findings. The main differential diagnosis with Ollier disease

Klippel-Trénaunay Syndrome and Parkes Weber Syndrome    
Klippel-Trénaunay syndrome is a congenital disorder classically characterized by three findings: a port-wine stain (nevus flammeus), abnormal venous structures (such as varicosities and venous malformations), and osseous and soft-tissue hypertrophy. This syndrome was initially described in 1900 by Klippel and Trénaunay (Klippel M, Trénaunay P 1900) and was originally called naevus vasculosus osteohypertrophicus. In 1907, Frederick Parkes Weber (Weber FP.1907) noted similar findings in association with arteriovenous malformations. This entity is referred to as Parkes Weber or Klippel-Trénaunay-Weber syndrome.
The affected limb can grow excessively, compared to the contralateral and the extent of the discrepancy in length between the limbs (LLD) can be mild to reach or exceed 10 centimeters. The effects of the pathological growth can only be evident at the level of a single bone (in particular the femur or tibia) or, in some cases, can affect an entire limb. (Jacob AG et al., 1998)The LLD may become evident in infancy, childhood or adolescence and is clearly diagnosable by comparing the folds of the buttocks or the back folds of the knees. The side signs are skin nevus, dilated superficial veins, limb enlargement, skin hyperthermia, dermatitis, ulcers and bleeding. These signs are not always present. Although the AOH syndrome is usually sporadic, in a few families it was noted autosomal dominant inheritance.

Conclusion
The evaluation, diagnosis, and management of vascular anomalies is a multidisciplinary process.  Some congenital vascular anomalies require no or minimal intervention, while others require a cohesive multidisciplinary approach.  It is important to recognize the clinical presentation and to establish an appropriate diagnosis to most appropriately evaluate and manage the patients. The rate of discovery continues to increase, expanding our understanding of the underlying interconnected molecular pathways.


References
1)    Abdalla, S.A. and Letarte, M. (2006) Hereditary haemorrhagic telangiectasia: current views on genetics and mechanisms of disease. J. Med. Genet., 43, 97–110.
2)    Bayrak-Toydemir, P., McDonald, J., Akarsu, N., Toydemir, R.M., Calderon, F., Tuncali, T., Tang, W., Miller, F. and Mao, R. (2006) A fourth locus for hereditary hemorrhagic telangiectasia maps to chromosome 7. Am J Med Genet A, 140, 2155–2162.
3)    Blei Francine, Bittman Mark E Congenital vascular anomalies: current perspectives on diagnosis, classification, and management Journal of Vascular Diagnostics and Interventions 2016
4)    Boon, L.M., Mulliken, J.B., Vikkula, M., Watkins, H., Seidman, J., Olsen, B.R. and Warman, M.L. (1994) Assignment of a locus for dominantly inherited venous malformations to chromosome 9p Hum. Mol. Genet., 3, 1583– 1587.
5)    Cole, S.G., Begbie, M.E., Wallace, G.M. and Shovlin, C.L. (2005) A new locus for hereditary haemorrhagic telangiectasia (HHT3) maps to chromosome 5. J. Med. Genet., 42, 577–582.
6)    Di Rocco C, Tamburrini G. Sturge-Weber syndrome. Childs Nerv Syst. 2006;22:909–921.
7)    Dutkiewicz AS, Ezzedine K, Mazereeuw-Hautier J, et al. A prospective study of risk for Sturge-Weber syndrome in children with upper facial port-wine stain. J Am Acad Dermatol. 2015;72 (3): 473–480.
8)    Fanburg JC, Meis-Kindblom JM, Rosenberg AE. Multiple enchondromas associated with spindle-cell hemangioendotheliomas. An overlooked variant of Maffucci’s syndrome. Am J Surg Pathol. 1995; 19 (9):1029–1038.
9)    Gao H, Wang B, Zhang X, et al. Maffucci syndrome with unilateral limb: a case report and review of the literature. Chin J Cancer Res. 2013 Apr;25 (2);254-8.
10)    Gorincour G, Kokta V, Rypens F, et al. Imaging characteristics of two subtypes of congenital hemangiomas: rapidly involuting congenital hemangiomas and non-involuting congenital hemangiomas. Pediatr Radiol. 2005;35 (12):1178–1185.
11)    Guttmacher, A.E., Marchuk, D.A. and White, R.I., Jr (1995) Hereditary hemorrhagic telangiectasia. N. Engl. J. Med., 333, 918–924.
12)    Hopyan S, Gokgoz N, Poon R, et al. A mutant PTH/PTHrP type I receptor in enchondromatosis. Nat Genet. 2002 Mar;30 (3):306-10.    
13)    Jacob AG, Driscoll DJ, Shaughnessy WJ, et al. Klippel-Trenaunay syndrome: spectrum and management. Mayo Clin Proc. 1998; 73 (1):28–36.
14)    Johnson, D.W., Berg, J.N., Baldwin, M.A., Gallione, C.J., Marondel, I., Yoon, S.J., Stenzel, T.T., Speer, M., Pericak-Vance, M.A., Diamond, A. et al. (1996) Mutations in the activin receptor-like kinase 1 gene in hereditary haemorrhagic telangiectasia type 2. Nat. Genet., 13, 189–195.
15)    Kaplan RP, Wang JT, Amron DM, et al. Maffucci’s syndrome: two case reports with a literature review. J Am Acad Dermatol. 1993;29 (5 pt 2):894–899.
16)    Klippel M, Trénaunay P. Du naevus variqueux osteohypertrophique. Arch Genet Med Paris 1900; 3:611 –672
17)    Lewis RJ, Ketcham AS. Maffucci’s syndrome: functional and neoplastic significance. Case report and review of the literature. J Bone Joint Surg Am. 1973;55 (7):1465–1479.
18)    McAllister, K.A., Grogg, K.M., Johnson, D.W., Gallione, C.J., Baldwin, M.A., Jackson, C.E., Helmbold, E.A., Markel, D.S., McKinnon, W.C., Murrell, J. et al. (1994) Endoglin, a TGF-beta binding protein of endothelial cells, is the gene for hereditary haemorrhagic telangiectasia type 1. Nat. Genet., 8, 345–351.
19)    McDonald J, Wooderchak-Donahue W, VanSant Webb C, et al. Hereditary hemorrhagic telangiectasia: genetics and molecular diagnostics in a new era. Front Genet. 2015;6:1–8.
20)    Mulliken, J.B. and Glowacki, J. (1982) Hemangiomas and vascular malformations in infants and children: a classification based on endothelial characteristics. Plast. Reconstr. Surg., 69, 412 –422.
21)    Mulliken JB, Glowacki J. Classification of pediatric vascular lesions. Plast Reconstr Surg. 1982;70 (1):120–121.
22)    North PE, Waner M, James CA, et al. Congenital nonprogressive hemangioma: a distinct clinicopathologic entity unlike infantile hemangioma. Arch Dermatol. 2001;137 (12):1607–1620.        
23)    Roach ES. Neurocutaneous syndromes. Pediatr Clin North Am. 1992;39 (4):591–620.
24)    Shirley MD, Tang H, Gallione CJ, et al. Sturge-Weber syndrome and port-wine stains caused by somatic mutation in GNAQ. N Engl J Med. 2013;368 (21):1971–1979.
25)    Waelchli R, Aylett SE, Robinson K, et al. New vascular classification of port wine stains: improving prediction of Sturge-Weber risk. Br J Dermatol. 2014;171 (4):861–867.
26)    Wassef M, Blei F, Adams D, et al. Vascular anomalies classification: recommendations from the international society for the study of vascular anomalies. Pediatrics. 2015;136 (1):e203–e214.
27)    Weber FP. Angioma formation in connection with hypertrophy of limbs and hemihypertrophy. Br J Dermatol 1907; 19:231–235

 

geneticapediatrica.it trimestrale di divulgazione scientifica dell'Euromediterranean Paediatric Foundation
Legge 7 marzo 2001, n. 62 - Registro della Stampa Tribunale di Messina n. 3/09 - 11 maggio 2009

Introduction
Cow’s milk protein allergy (CMPA) is the leading cause of food allergy in infants and young children younger than 3 years [1, 2], with diverse clinical presentation (e.g., cutaneous, gastrointestinal, and respiratory) of variable intensity in infants [1-3]. Although CMPA is thought to result from an immunological reaction to one or more milk proteins (e.g., immunoglobulin (Ig)E mediated, non-IgE mediated, or mixed) its aetiology is still unclear [1-3].
However, it is becoming evident that multiple factors, through modulation of immune response, can influence the development of CMPA [4]. On this regard, numerous studies have examined the possible benefits of breast-feeding in comparison to cow’s milk formula-fed on food allergy in children [5, fig. 1].

Fig.1 Julia V, Macia L, Dombrowicz D. The impact of diet on asthma and allergic diseases. Nat Rev Immunol. 2015 May;15 (5):308-22

In fact, breast milk contains a multitude of immunologically active components, including interleukin (IL)-10, a major regulatory cytokine of inflammatory responses, which, by promoting the induction of Th-1 and the suppression of Th-2 function, helps infants in the essential immune adaptations [1, 3, fig. 2, 3].

Fig.2 Breastmilk composition (http://www.pregworld.org/breast-milk-composition/)

Fig.3 Kim AR, Kim HS, Kim do K, et al. Mesenteric IL-10-producing CD5+ regulatory B cells suppress cow's milk casein-induced allergic responses in mice. Sci Rep. 2016 Jan 20;6:19685.

IL-10, mainly produced by Th-reg, is a suppressive cytokine of T-cell proliferation in both Th1 and Th2 cells, and it seems to be an important immune factor to regulate the immune response, and it favors the immune tolerance [6]. Herein, we evaluated if breast-feeding can represent a protective factor in CMPA-related atopic eczema/dermatitis syndrome (AEDS) children. Moreover, we measured serum IL-10 levels from breastfed and not breastfed infants with CMPA, during symptomatic and asymptomatic periods to evaluate its utility as marker of disease evolution.

Patients and Methods
32 breast-feeding children with CMPA-related AEDS (17 males and 15 females; mean age 5.56±2.41 months; 7 mild AD; 12 moderate AEDS; 13 severe AEDS) and 30 artificial feeding babies (16 males and 14 females; mean age 6.01±2.08 months; 14 mild AEDS; 5 moderate AEDS; 11 severe AEDS) were evaluated. 73 healthy children (39 males and 34 females; mean age was 5.51±2.93 months) were enrolled as control group.
The severity of AEDS was evaluated according to Score Atopic Dermatitis (SCORAD) index [7].
Subjects were eligible if they were affected by CMPA, assessed according to the NICE [8], positive cow’s milk challenge test and skin prick tests (SPT) or determination of serum total IgE levels.
Exclusion criteria were the following: associated diseases, any infection within one month before the start of the study, other skin diseases, positive SPT for additional food allergens, and use of any treatment, capable of interfering with the results in the previous month.
In all patients serum total IgE and IL-10 levels were detected.

Data analysis
Data were tabulated using Excel 2003 (Microsoft Corp., Redmond, WA, USA). The data collected were statistically analyzed by the statistical computer software SPSS, version 15.0. A p value of less than 0.05 was considered to be statistically significant.

Results
We found significant Score Atopic Dermatitis (SCORAD) index point differences between breastfed and not breastfed children (p<0.001). The serum IL-10 levels were lower in children with CMPA as compared to the healthy control group (p<0.001). Moreover, we also detected a significant inverse correlation between serum IL-10 levels and SCORAD in both enrolled groups. In particular, IL-10 levels, in both groups, were significantly lower in children with severe symptoms. Conversely, serum IL-10 levels were significantly increased in children with mild-severe symptoms in both groups. Furthermore, breast-fed children, with lower severe symptoms, had higher serum IL-10 levels.
Finally, serum total IgE levels were negatively correlated with serum IL-10 levels in both breastfed and non-breastfed children with CMPA (p<0.001).

Conclusions
In the present study, we compared a group of breastfed patients with not-breastfed children in order to evaluate the role of breast-feeding on CMPA-related AEDS.
In our study, we have shown that breast-feeding is associated to a minor severity disease in CMPA-related AEDS children and it also promotes the systemic release of IL-10, reaching similar values to those found in control group.
These data are also in agreement with a recent observation that IL-10 has a central role in down-regulating inflammatory cascades [9, 10]. It is well established that IL-10 has also an important role in the regulation of Th2 and allergic responses. Our data confirmed this issue because artificial- feeding infants, with more severe symptoms, had lower IL-10 levels. Instead, breast-fed children, with lower severe symptoms, had higher serum IL-10 levels (4.66±0.38 pg/ml).
Our findings also confirm the hypothesis that AEDS severity, in subjects affected by CMPA, might be associated with an impaired regulation of IL-10. During follow-up, we observed a statistically significant increase of IL-10 serum levels in breastfed children that underwent a positive open oral challenge test. Our results suggest that breast-feeding, with the participation of suppressor cytokines, such as IL-10, may induce hyposensitization in children affected by CMPA. These findings indicate also that IL-10 may become a useful marker of disease improvement monitoring.

References

1. Fiocchi A, Schünemann HJ, Brozek J, Restani P, Beyer K, Troncone R, Martelli A, Terracciano L, Bahna SL, Rancé F, Ebisawa M, Heine RG, Assa'ad A, Sampson H, Verduci E, Bouygue GR, Baena-Cagnani C, Canonica W, Lockey RF: Diagnosis and Rationale for Action Against Cow's Milk Allergy (DRACMA): a summary report. J Allergy Clin Immunol 2010;126 (6):1119-28.e12
2. S. Koletzko, B. Niggemann, A. Arato, J.A. Dias, R. Heuschkel, S. Husby, M.L. Mearin, A. Papadopoulou, F.M. Ruemmele, A. Staiano, M.G. Scha¨ppi, and Y. Vandenplas: Diagnostic Approach and Management of Cow’s-Milk Protein Allergy in Infants and Children: ESPGHAN GI Committee Practical Guidelines. JPGN 2012;55 (2):221-9.
3. Katz Y, Rajuan N, Goldberg MR, Eisenberg E, Heyman E, Cohen A, Leshno M: Early exposure to cow's milk protein is protective against IgE-mediated cow's milk protein allergy. J Allergy Clin Immunol 2010;126 (1):77-82.e1.
4. Yavuz ST, Buyuktiryaki B, Sahiner UM, Birben E, Tuncer A, Yakarisik S, Karabulut E, Kalayci O, Sackesen C: Factors that predict the clinical reactivity and tolerance in children with cow's milk allergy. Ann Allergy Asthma Immunol 2013;110 (4):284-9.
5. Greer FR, Sicherer SH, Burks AW; American Academy of Pediatrics Committee on Nutrition; American Academy of Pediatrics Section on Allergy and Immunology: Effects of early nutritional interventions on the development of atopic disease in infants and children: the role of maternal dietary restriction, breastfeeding, timing of introduction of complementary foods, and hydrolyzed formulas. Pediatrics 2008;121 (1):183-91.
6. Couper KN, Blount DG, and Riley EM. IL-10: the master regulator of immunity to infection. J Immunol 180:5771–5777, 2008.
7. Severity scoring of atopic dermatitis: the SCORAD index. Consensus Report of the European Task Force on Atopic Dermatitis. Dermatology 1993;186:23–31.
8. Baker M: NICE guidance points the way to tackling eczema in children. Community Pract 2013;86 (9):40. 42
9. Tiemessen MM, Van Ieperen-Van Dijk AG, Bruijnzeel-Koomen CA, Garssen J, Knol EF, Van Hoffen E: Cow's milk-specific T-cell reactivity of children with and without persistent cow's milkallergy: key role for IL-10. J Allergy Clin Immunol 2004;113 (5):932-9.
10. Ma S, Yin J: Imbalance of serum IL-10 and TGF- in patients with pollen food syndrome. Allergol Immunopathol (Madr) 2013; 42 (3):198-205
     

geneticapediatrica.it trimestrale di divulgazione scientifica dell'Euromediterranean Paediatric Foundation
Legge 7 marzo 2001, n. 62 - Registro della Stampa Tribunale di Messina n. 3/09 - 11 maggio 2009

Introduction

"e;€¨Abnormal changes of skin colour are observed in a vast number of diverse disorders with different underlying mechanisms. Essentially, any change of the components that contribute to normal skin colour, resulting in their excess or deficiency within the skin will produce an altered cutaneous colouration. Some authors use the term ‘pigmentation’ disorders as synonymous to ‘melanotic’ disorders to include entities that are characterized by a pathological change in melanin or melanocytes and differentiate these from the ‘non-melanotic’ disorders which are due to alterations of other cutaneous chromophores (1). Melanotic disorders are broadly divided into hypermelanotic (due to excess melanin but normal melanocytic population) and hypermelanocytic (due to normal melanin and increased melanocytic proliferation) as well as hypomelanotic/amelanotic and hypomelanocytic/amelanocytic which are due to melanin deficiency and reduction or absence of melanocyte number respectively. (2). Further categorization distinguishes these groups into congenital and acquired, circumscribed, mixed and generalized, epidermal, dermal and mixed groups."e;€¨We present a patient who had hypopigmentation disorders likely secondary to hereditary disease, describing probable causes and its differential diagnosis.

Case report "e;۬

A.C., the  third child born of non consanguineous Caucasians parents, was born by cesarean section, at 33 weeks of gestational age. The pregnancy was complicated by placenta previa. The birth weight was 1750 g. At birth she was in good health, exept transitory hypotonia. Although prematurity she has not needed intensive cure during the first days of life. At the age of 7 years she was admitted to the emergency room of our hospital, AOU G. Martino of Messina, for traumatic head injury. She was submitted to cranial TC, which showed lesions typical of previous cerebral ischemia; therefore she was hospitalized in Pediatric Neurology Department for further investigations. During the hospitalization she was referred to our observation for multiple, rounded and ovalar, asymptomatic, well marginated, hypomelanotic skin macules on the trunk, upper and lower limbs, associated to skin xerosis (Fig. 1 a, b, c). These lesions had been present since she was 1 years. No other phenotic features were present. Familiar dermatological history was positive since mother, older brother, maternal uncle, two cousins (affected uncle’s sons) had presented, when they were children, the same skin lesions which were regressed at the pubertal age. On the suspicion of hereditary disease with autosomal dominant inheritance, genetic investigations as cromosome karyotyping, array comparative genomic hybridization were performed and they are still ongoing.
 

(a)

(b)

(c)

Fig. 1 Multiple, rounded and ovalar, asymptomatic, well marginated, hypomelanotic skin macules on the trunk (a), upper and lower limbs, associated to skin xerosis

Discussion"e;۬

Hypo- and hyper-pigmentation disorders are the most severe dermatological diseases observed in patients from all over the world. These disorders can be divided into melanoses connected with disorders of melanocyte function, which could be secondary to immunological, infectious or inflammatory causes, and hereditary melanocytoses connected with melanocyte development. Hereditary hypomelanocytoses are caused by abnormal melanoblast development, migration and proliferation as well as by abnormal melanocyte viability and proliferation. These disorders are represented by piebaldism, Waardenburg syndrome, and Tietz syndrome, and are caused by different mutations of various or the same genes. The types of mutations comprise missense and nonsense mutations, frameshifts (in-frame insertions or deletions), truncating variations, splice alterations and non-stop mutations (3).
Piebaldism is a rare autosomal-dominant disorder of melanocyte development characterized by congenital poliosis, and stable patches of leukoderma (4). Piebaldism is caused by the congenital absence of melanocytes in affected areas of the skin and hair due to mutations of the c-kit gene, located on chromosome 4q12, which affects the differentiation and migration of melanoblasts from the neural crest during the embryonic life (5). Characteristically, lesions of leukoderma are well-circumscribed, irregular, white patches that are often accompanied by hyperpigmented macules noted on both depigmented and unaffected adjacent skin (6). Poliosis circumscripta, traditionally known as white forelock, may be the only manifestation in 80% to 90% of cases and is present at birth (7). The white forelock typically appears in a triangular shape and the underlying skin of the scalp also is amelanotic. Pigmentary anomalies in piebaldism are typically restricted to the hair and skin. Rare associations have been reported with piebaldism, in particular Hirschprung's disease or aganglionic megacolon, supporting evidence of a network of interacting genes and proteins for the regulation of melanocytes and enteric plexus neurons during their development at the time of embryogenesis (8). "e;۬Waardenburg syndrome is a genetic syndrome consisting primarily of anomalies of the skin, hair, eyes, and ears and affects 1 in 40, 000 according to population studies (9). There are four major types of Waardenburg syndrome, generally involving the PAX3 (Paired box 3), MITF
(microophthalmia-associated transcription factor), SOX10 (Sry box 10), EDN3 (endothelin 3), and EDNRB (endothelin receptor type B) genes (10, 11). As established at the Waardenburg consortium, diagnosis requires fulfillment of either two major criteria or one major and two minor criteria. (12) Major criteria include the characteristic white forelock (hair depigmentation), pigmentary anomalies of the iris, congenital sensorineural deafness, dystopia canthorum, or an affected first degree relative. Minor criteria are depigmented macules or patches, synophrys, broad nasal root, nose hypoplasia, or early graying of the hair by age 35. The musculoskeletal system and gastrointestinal tract can also be affected in certain subtypes.
Tietz syndrome is a hypopigmentation-deafness syndrome resulting, like WS2, from mutations in the region of the MITF gene encoding the DNA-binding domain, resulting in early embryionic loss of all melanocyte precursors. Tietz syndrome individuals exhibit generalized cutaneous hypopigmentation similar to that found in albinism, rather than distinct depigmented patches (13). Affected individuals could also showed mild craniofacial anomalies and retention of some hearing function.
In the case we have described differential diagnosis include several form of acquired hypopigmentation disease enclosing:"e;€¨- pityriasis alba, which is a localized hypopigmented disorder of childhood with many existing clinical variants. Poor cutaneous hydration appears to be a common theme for most risk factors and may help elucidate the pathogenesis of this disorder. Alleviation of the various risk factors, such as atopy, xerosis and mineral deficiencies, via patient education on proper skin care and hygiene, use of lubricants and emollients, topical corticosteroid therapy in the presence of inflammation, and the novel administration of topical anti-inflammatory drugs such as calcineurin inhibitors can play a crucial role in promoting remission or resolution (14). "e;€¨- iatrogenic causes of hypopigmentation disorders, such as those related to imiquimod, an immune response modifier used to treat certain types of skin growths (actinic keratoses) or skin cancer (superficial basal cell carcinoma) (15, 16). "e;€¨- exposure to toxic agents such as hydroquinone, aromatic organic compound, which represent the major component in most black and white photographic developers for film and paper  and which is contained in some cosmetics too (17). "e;€¨- cutaneous infectious disease such as pityriasis versicolor, a superficial fungal infection of the skin caused by Malassezia species that induces a characteristic rash of well demarcated, thin, scaly plaques that can be hypopigmented, hyperpigmented, or erythematous which need topical treatments including ketoconazole, selenium sulphide, or zinc pyrithione shampoo (18).
Conclusions"e;۬Due to the wide spectrum of causes of hypopigmentation disorders, the diagnosis is not always immediate. In our patient positive results of genetic investigations could lead to a resolution of the case. On the contrary, in the case of negativity we could consider, in addition of aforementioned causes of acquired hypopigmentation diseases, the diagnosis of a specific and endemic skin disease such as pityriasis rotunda sardoa, an idiopathic dyschromic-desquamative genodermatoses characterized by hypopigmented lesions which were resolved in the pubertal age, described in the last century in some Mediterrean areas, especially in Sardinia (19).

References
1.    Dessinioti C, Stratigos AJ, Rigopoulos D, Katsambas AD. A review of genetic disorders of hypopigmentation: lessons learned from the biology of melanocytes. Exp Dermatol. 2009 Sep; 18 (9):741-9.
2.    Mollet I, Ongenae K, Naeyaert JM. Origin, clinical presentation and diagnosis of hypomelanotic skin disorders. Dermatol Clin 2007; 25: 363–371.
3.    OtrÄ™ba M, MiliÅ„ski M, Buszman E, WrzeÅ›niok D, Beberok A. Hereditary hypomelanocytoses: the role of PAX3, SOX10, MITF, SNAI2, KIT, EDN3 and EDNRB genes. Postepy Hig Med Dosw (Online). 2013 Nov 26;67:1109-18.
4.    Grob A, Grekin S. Piebaldism in children. Cutis 2016 Feb;97 (2): 90-2.
5.    Saurabh A, Amit O. Piebaldism: a brief report and review of the literature. Indian Dermatol Online J. 2012 May-Aug; 3 (2): 144–147.
6.    Janjua SA, Khachemoune A, Guldbakke KK. Piebaldism: a case report and a concise review of the literature. Cutis. 2007;80:411-414.
7.    Sleiman R, Kurban M, Succaria F, et al. Poliosis circumscripta: overview and underlying causes. J Am Acad Dermatol. 2013;69:625-633.
8.    Mahakrishnan A, Srinivasan MS. Piebaldism with Hirschsprung's disease. Arch Dermatol. 1980;116:1102.
9.    Keena T. Que S, Weston G, Sucheki J, Ricketts J. Pigmentary disorders of the eyes and skin. Clinics in Dermatology. 2015; 33:147-150.
10.    Pingault V, Ente D, Dastot-Le Moal F, et al. Review and update of mutations causing Waardenburg syndrome. Hum Mutat. 2010;31:391-406.
11.    Bansal Y, Jain P, Goyal G, Singh M, Mishra C. Waardenburg syndrome—A case report. Cont Lens Anterior Eye. 2013;36:49-51.
12.    Farrer LA, Grundfast KM, Amos J, et al. Waardenburg syndrome (WS) type I is caused by defects at multiple loci, one of which is near ALPP on chromosome 2: First report of the WS consortium. Am J Hum Genet. 1992; 50:902-913.
13.    Libro di dermatologia
14.    Jadotte YT, Janninger CK. Pityriasis alba revisited: perspectives on an enigmatic disorder of childhood. Cutis. 2011 Feb;87 (2):66-72.
15.    Mori S, Akasaka T. Imiquimod-induced vitiligo-like hypopigmentation after treatment for radiation keratosis. Clin Exp Dermatol. 2016 Dec; 41 (8): 930-932.
16.    Gowda S, Tillman DK, Fitzpatrick JE, Gaspari AA, Goldenberg G. Imiquimod-induced vitiligo after treatment of nodular basal cell carcinoma. J Cutan Pathol. 2009 Aug; 36 (8):878-81.
17.    Matsumoto M, Todo H, Akiyama T, Hirata-Koizumi M, Sugibayashi K, Ikarashi Y, Ono A, Hirose A, Yokoyama K. Risk assessment of skin lightening cosmetics containing hydroquinone. Requl Toxicol Pharmacol. 2016 Aug 10; 81:128-135.
18.    Renati S, Cukras A, Bigby M. Pityriasis versicolor. BMJ. 2015 Apr 7; 350:h1394.
19.    Ena P, Chiarolini F, Gallus D, Spano G, Masotti A, Leigheb G. Pityriasis rotonda sardoa. Studio clinico, epidemiologico e follow-up su 47 casi osservati nel nord Sardegna dal 1981 al 1995. Giornale Italiano di Dermatologia e Veneorologia. 1998 Apr:133 (2):93-9.

 

geneticapediatrica.it trimestrale di divulgazione scientifica dell'Euromediterranean Paediatric Foundation
Legge 7 marzo 2001, n. 62 - Registro della Stampa Tribunale di Messina n. 3/09 - 11 maggio 2009

Introduction
We describe a 9 year-old Caucasian girl with a complex background of premature birth, cerebral palsy, spastic quadriplegia, generalised dystonia and visual impairment who developed a gastro-colonic fistula following Vygon Mic-Key button change.
She first presented to our Emergency Department (ED) with a 3 weeks-history of watery diarrhoea occurring after every feed and weight loss. She had several admissions to her local hospital over the previous weeks and had been treated for possible gastroenteritis without any improvement.
On admission she was severely dehydrated and was having recurrent seizures. She required a number of fluid boluses. Her gastrostomy site was bleeding and her perianal area was quite inflamed.
Blood tests showed leukocytosis with neutrophilia, hypernatraemia, hypokalaemia, deranged creatinine and urea and prolonged APTT. Blood gas revealed metabolic acidosis. Inflammatory markers, urine and stools cultures (including testing for Clostridium Difficile) were within normal limits. There were dilated bowel loops on abdominal X-ray. After her resuscitation, decision made was to stop her enteral feeds and start her on parenteral nutrition and intravenous metronidazole, ceftriaxone and gentamicin. A further contrast study done through her gastrostomy showed the Mic-Key button extension lying in the transverse colon.
She underwent a laparotomy which showed a gastro-colonic fistula with the wall of the transverse colon attached to the internal abdominal wall. There were no signs of peritoneal contamination. Gastro-colonic fistula was closed and a new gastrostomy was performed. Five days after the operation, the patient tolerated her feeds, diarrhea disappeared, parenteral nutrition was stopped and she reached full enteral feeds.
On further questioning it became clear that her symptoms started at the time of her last Vygon Mic-Key button replacement that took place in the Community where she was receiving her usual care. It is possible that during the initial percutaneous endoscopic gastrostomy (PEG) insertion, the tube passed through the transverse colon before entering the posterior wall of the stomach therefore forming a gastro-colonic fistula. On changing the Vygon Mic-key button, it was placed into the transverse colon rather than going all the way to the stomach. As a result, enteral feeds and medications bypassed the usual way and were given directly into the transverse colon. This explained the diarrhoea and the rest of her symptoms such as the weight loss and the seizures (poor absorption of nutrients and medicines).

Discussion
A gastrostomy tube is an artificial device placed into an opening made through the abdominal wall and into the stomach lumen (Figure A).
Until 1980 all permanent intragastric feeding tubes were inserted surgically by a laparotomy. Following 1980, it has become widely accepted that endoscopic insertion is less invasive, more rapid, creates less incisional pain, and reduces length of stay and procedure costs, as well as being at least as safe as the surgical approach (1).
As all surgical procedures there are a number of complications. One of these complications is malposition of tube which is placed in an organ other than the stomach such as small bowel, large bowel, peritoneal cavity or abdominal wall (2). During the procedure, if a loop of small or large bowel is between the stomach and the abdominal wall, it may be caught by the needle and as result, the PEG tube will be inserted through the bowel wall into the stomach; as the time passes, the tract around the gastrostomy epithelializes and forms a fistula, for example a gastro-colonic (Figures B and C). If the fistula does not have time to mature (about six weeks), the removal or replacement of the tube may dislodge the tube from the stomach to another organ such as the colon (as in our case) with a high risk of peritonitis (2) (Figure C).
Gastro-colonic fistula should be considered in any patients with PEG who develops diarrhoea after replacement of the gastrostomy tube (3). Gastro-colonic fistula associated with diarrhea is often seen after the replacement of the original PEG with a Mic-Key button because it is shorter than a PEG tube and can easily be found directly into the colon (4). Investigations needed after a suspicion of a PEG malposition are: 1) CT scan with oral/gastrostomy gastrograffin to show the position of the hub and the tube. 2) Contrast study of the gastrostomy tube by injecting gastrograffin in the tube and performing a plane X-ray of the abdomen or doing a fluoroscopy to show the position of the hub and any extravasation of the contrast into the peritoneal cavity. 3)Endoscopy can confirm the migration of the PEG from the stomach (1, 5). 4) Gastrograffin enema identifying a filling defect in the colon can verify the position of the hub in the colon (1, 5).
Gastro-colonic fistula is a known complication of PEG insertion and if it is not recognized, can lead to life-threating symptoms as hypovolemic shock and death. Clinicians should be aware of signs and symptoms of PEG malposition, that are obviously non-specific, and they should ask for the proper diagnostic investigations.

Fig.A, B, C. (A) Normal insertion of a gastrostomy tube. (B) Formation of a gastro-colonic fistula. (C) Mic-key button in the colon after the formation of a gastro-colonic fistula. Re-designed by T. Alterio from: Okutani D et al. “A case of gastrocolocutaneous fistula as a complication of percutaneous endoscopic gastrostomy”. Acta Medica Okayama. Vol. 62, 2008 Issue 2, Art. 9

References
1.    Heuschkel RB, Gottrand F, Devarajan K, Poole H, Callan J, Dias JA, Karkelis S, Papadopoulou A, Husby S, Ruemmele F, Schäppi MG, Wilschanski M, Lionetti P, Orel R, Tovar J, Thapar N, Vandenplas Y, European Society for Pediatric Gastroenterology, Hepatology, and Nutrition. ESPGHAN position paper on management of percutaneous endoscopic gastrostomy in children and adolescents. J Pediatr Gastroenterol Nutr. 2015 Jan;60 (1):131-41. doi: 10.1097/MPG.
2.    Milanchi S, Wilson MT. Malposition of percutaneous endoscopic-guided gastrostomy: Guideline and management”. “J Minim Access Surg 2008. 4 (1): 1-4.
3.    Hudziak H, Loudu P, Bronowicki JP, Clavière C, Chone L, Bigard MA. Diarrhea following the replacement of percutaneous endoscopic gastrostomy tube: To think of gastrocolic fistula. Gastroenterol Clin Biol. 1996;20:1139–40.
4.    Jensen SW, Eriksen J, Kristensen K. Severe diarrhea after the original well-functioning percutaneous endoscopic gastrostomy tube was replaced by a MicKey button. Ugeskr Laeger. 2006;168:1038–9.
5.    Roozrokh HC, Ripepi A, Stahlfeld K. Gastrocolocutaneous Fistula as a complication of peg tube placement. Surg Endosc. 2002;16:538–9

 

geneticapediatrica.it trimestrale di divulgazione scientifica dell'Euromediterranean Paediatric Foundation
Legge 7 marzo 2001, n. 62 - Registro della Stampa Tribunale di Messina n. 3/09 - 11 maggio 2009

Introduction

Microduplications are a type of genetic mutation, that result from the duplication of a small part of a chromosome. They are typically one to three megabases long, they involve several contiguous genes and represent the reciprocal product of duplicon mediated non-allelic homologous recombinations, a mechanism which often lead to microdeletions.Their location is extremely variable, even if  a specific "critical region" may be consistently involved.
Lots of phenotypic effects of  microduplications are caused by  changes in some critical dose-sensitive genes or in a single gene, but it’s essential the intervention of a duplication that disrupts its integrity. Microduplication syndromes often present a non- clear phenotype and another peculiarity is that they are inherited from apparently normal parents. Since many microduplications are not associated with physical or developmental anomalies, the functional role of genes included with in the copy number variations of speciï¬�c chromosome regions requires careful evaluation. Selected dosage-sensitive genes may affect the phenotype by both haploinsufï¬�ciency and duplication. We report the case of a male pediatric patient with multiple congenital anomalies and  mild mental retardation whose genotype was characterized by two microduplication phenomena: the first one involving 16p13.3  region and the second one involving 16q11.2 region, both interesting a non codifying sequence.

Case report
The probando, a 6-year-old male, is the �rst child of healthy, nonconsanguineous parents. The mother has had two pregnancies: by the first one our patient was born, while the second one was prematurely interrupted because of fetal heart disease (complete atrioventricular canal).
He was born at the 34th + 6 days of gestation by abortion threats complicated pregnancy. Birth weight was 1, 800 g (under 5th centile), birthlength 42 cm (10th centile) and head circumference 31 cm (between 10th and 25th centile).
At birth, congenital hypotiroidism was diagnosed, so hormone replacement therapy was initiated. A month later, the child underwent a surgical intervention to correct  isthmian coarctation of the aorta and interventricular defect), a congenital heart defect formerly diagnosed with fetal echocardiography.
Since the first months of life he was fed with artificial nutrition, mainly through gavage.  Psychomotor development was delayed and a severe impairment of both ponderal and statural growth was observed.
Another surgical intervetion was carried out at the age of three because of bilateral inguinal hernia
At the time of our ï¬�rst observation, at the age of five years, his speech was limited to few short sentences and impaired because of vocal cord paralysis. He wasn’t autonomous in everyday activities. On neurological exam he had moderate mental retardation and no behavioral problems were observed. He never had had seizures.
Transfontanellar ultrasonography, carried out in the first months of life, didn’t highlight anomalies: the supratentorial ventricular system appears on axis if compared to the midline;  morphology and volume limits were also normal like audiological evaluation. No alteration at the eye exam with fundus oculi , except for bilateral strabismus.  Urogenital system was free from malformation events.
On laboratory findings hypercalcemia was detected, but other bioumoral parameters weren’t modified.
On physical examination, weight was 11.5 kg (< 3th centile), height 97 cm (- 2 SD) and head circumference 44 cm (< 3th centile). He had a peculiar face appearance characterized by microcephaly, epicanthus , hypertelorism, strabismus, flat nasal root, big low-set ears with left folded lobe. Hands were characterized by bilateral clinodattilia.

Methods and results
Genetic investigations
Fluorescence in situ hybridization (FISH) for Williams syndrome was negative. Analysis for Kabuki syndrome and chromosome 11 methylation test are both negative.
ArrayCGH documented 2 microduplications, the first one situated in the 16p13.3 region, extended about 80 Kb, and the second one  involving 16q11.2 region, externded about 152 Kb. Both microduplication were inherited from patient’s father, who was healthy.
NGS was also performed, but we are still attend the result.  

Discussion

The described patient presents a double microduplication interesting two different regions of chromosome 16.
According to literature case-report and studies, most of duplications phenomena interesting 16p13.3 region, are associated with a peculiar phenotype characterized by mental retardation, delay in weight and height developement, small head circumference, facial anormalities and fingers anomalies, e.g. clinodattilia observed in our probando. How is demonstrate by Digilio et al., phenotypic severity, such as facial dysmorphism and intellectual disability in patients with 16p duplication , could be correlated with the size of the duplicated segment.
Other 16p13.3 duplication cases are also associated with a more severe phenotype, characterized by major malformations, such as cleft palate, congenital heart defects and genitourinary abnormalities, severe psychomotor retardation, growth delay with microcephaly, seizures, and a speci�c facial appearance (round head, upslanting and narrow palpebral �ssures, sparse eyebrows, broad nasal bridge, rounded nasal tip, , long philtrum, thin upper lip, prominent glabella and micrognathia). 16q11.2 duplication is less frequent, in fact there ore only few studies about it. One of them describes the association with q13 in a 15 week foetus, who presented facial anomalies, mild pyelectasia, minor dysgenesis of the ovaries and an atypical outflow of the right arteria thyroidea ima.
All these duplications are much larger than those detected in our patient and they interest the codifying sequence, differently from our case.
 

Conclusions
Both the microduplication described in our patient can’t be considered as only responsible of the phenotype. They are benign, short, interesting a non codifying sequence and they are also present in his father, who doesn’t present phenotypical alteration or health problems. Although errors in development and phenotypical alteration will not be curable by such strategies, the observed problems in mental function might beneï¬�t from therapeutic intervention.   



References

1. Marangi G. , Leuzzi V., Orteschi D., Grimaldi M.E. , Lecce R., Neri G., Zollino M., Duplication of the Rubinstein–Taybi Region on 16p13.3 Is Associated With a Distinctive Phenotype, Am J Med Gen Part A 146A:2313–2317 (2008)
2. Chen J-L, Yang Y-F, Huang C, Wang J, Yang J-F, Tan Z-P. 2012. Clinical and molecular delineation of 16p13.3 duplication in a patient with congenital heart defect and multiple congenital anomalies. Am J Med Genet Part A 158A:685–688
3. Digilio MC, Bernardini L, Capalbo A, Capolino R, Gagliardi MG, Marino B, Novelli A, Dallapiccola B. 2009. 16p subtelomeric duplication: A clinically recognizable syndrome. Eur J Hum Genet 17:1135– 1140.
4. Kokalj-Vokac N., Medica I., Zagorac A., Zagradisnik B., Erjavec A., Gregoric A. 2000. A case of insertional translocation resulting in partial trisomy 16p. Ann Genet 43:131–135.
5. Martin CL, Waggoner DJ, Wong A, Uhrig S., Roseberry JA. , Hedrick JF., Pack SD., Russell K., Zackai E., Dobyns WB., Ledbetter DH. 2002. ‘‘Molecular rulers’’ for calibrating phenotypic effects of telomere imbalance. J Med Genet 39:734–740.
6. Trimborn M., Wegner R.D., T¨onnies H., Sarioglu N., Albig M., Neitzel H., Prenatal diagnosis and molecular cytogenetic characterisation of a small de novo interstitial duplication 16q11.2-q13 , Prenat Diagn 2006; 26: 273–276.
7. Thienpont B., Béna F., Breckpot F., Devriendt N.P., Bottani A., Vermeesch J.R., Sau Wai Cheung and Koen Jean-Pierre Fryns, Geert Mortier, Jan M Friedman, Laurent Villard, Gorski, Farah Zahir, Siu Li Yong, Michael M Morris, Stefania Gimelli, Chin-To Fong, Jennifer L Kussmann, Dorothy K Grange, Jerome L Björn Menten, Hilde Van Esch, Emmanuel Scalais, Jessica M Salamone, Duplications of the critical RubinsteineTaybi deletion region on chromosome 16p13.3 cause a novel recognisable syndrome, J Med Gen  2010 47: 155-161 originally published online October 14.

geneticapediatrica.it trimestrale di divulgazione scientifica dell'Euromediterranean Paediatric Foundation
Legge 7 marzo 2001, n. 62 - Registro della Stampa Tribunale di Messina n. 3/09 - 11 maggio 2009

Abstract
The term “vascular ring” is generally used to describe a variety of congenital vascular abnormalities of the aortic arch system that can cause tracheal or oesophageal compression. Its clinical presentation is extremely heterogeneous, representing a clinical challenge for paediatricians especially when atypical and/or aspecific clinical symptoms are occurring.
Here, we report three cases of children with three different clinical presentations of a double aortic arch, one of them showed both middle lobe syndrome and double aortic arch, firstly reported in literature.

Key words: children; double aortic arch; middle lobe syndrome; vascular ring.


Introduction
The term “vascular ring” is generally used to describe a variety of congenital vascular abnormalities of the aortic arch system that can cause tracheal or oesophageal compression. The multiple different varieties of vascular rings are due to a defective persistence, involution, or regression of embryonic aortic arches. The clinical manifestations vary primarily with the severity of encroachment by the abnormal vessel on the oesophagus, trachea or bronchus and therefore include stridor, wheezing, cough and dysphagia1.
The double  aortic arch is the most common vascular ring and comprise 1-2% of all cardiac abnormalities2-3. It results from the failure of the fourth embryonic branchial arch to regress, leading to an ascending aorta that divides into a left and right arch that fuse together to completely encircle the oesophagus and the trachea4. The three main types of double aortic arch are the dominant right arch with a small left arch (80%), dominant left arch (10%), and balanced aortic arches (10%)5.
Here, we report three cases of children with three different clinical presentations of a double aortic arch, one of them showed both middle lobe syndrome and double aortic arch, firstly reported in literature.

Case reports
Case 1
We report the case of a one-month-old baby who came to our attention because affected by episodes of asphyxia during the feeding. After delivery she was hospitalized elsewhere due to tremor and dyspnoea. She was discharged and diagnosed with convulsions and gastroesophageal reflux. Afterwards, she suffered from 3 chocking episodes during the feeding. Due to the persistence of the symptoms, she was admitted to our attention. At the entrance her general conditions were poor: mental dullness, stridor was found after chest auscultation, jugular-epigastric inspiratory dyspnoea and rhonchi and rales spread all over the lungs.  For this reason, nourishment was suspended; Infusion therapy and constant monitoring have been carried out. Blood test results were normal. Chest X-ray examination, electroencephalogram and electrocardiogram were negative. Laryngeal-fibroscopy highlighted a posterior hyperaemia of the larynx due to gastroesophageal reflux. Then a contrast-enhanced oesophagogram has been carried out. It showed an anterior tracheal deviation and an oesophageal notch (Fig.1). The tracheoscopy showed a narrowing due to extrinsic compression at about 1cm from the carina. For this reason, we advanced the hypothesis of vascular ring, then confirmed following a computed tomography (CT)-angiography. The newborn girl underwent surgery in order to get her vascular defect repaired. The long-term follow-up showed a clear improvement and resolution of symptoms.

Fig. 1. Esophagogram showing an anterior tracheal deviation and an esophageal notch.

Case 2
We report the case of a 8-month-old baby girl born at 35 weeks gestation by emergency caesarean section. The baby girl came to our attention due to persistent cough since the first months of her life. When she entered the ward her general conditions were satisfactory. The general examination showed hyperaemic pharynx; the chest auscultation revealed sour breath with inconstant rhonchi and breathing noises. Blood tests only showed leucocytosis. Then a chest x-ray examination and ultrasound have been carried out. Laryngoscopy revealed morphological aspects that were consistent with laryngomalacia and signs of gastro-oesophageal reflux disease. The echocardiography highlighted a dominant right double aortic arch with anterior trachea. Furthermore, the oesophagogram showed a minor anomaly of the oesophageal profile in the middle tract, as no dilation in the upper cervical-thoracic duct was revealed. Vascular ring was diagnosed and confirmed by the CT-angiography. For this reason, the baby girl has undergone repair surgery, which was followed by improvement of the symptoms.

Case 3
We report the case of a 3-year-old baby girl born at 41 weeks gestation with an history of acute respiratory distress after birth. She had persistent cough since the first months of her life. For this reason, the baby girl came to our attention due to persistent barking cough and recurrent pneumonia. When she entered the ward her general conditions were poor. Physical examination revealed a febrile, baby-girl with cyanotic skin and oxygen saturation of 88% on room air. She was grunting with severe jugular and epigastric retractions, and the chest auscultation revealed rhonchi and rales all over the lungs. Blood tests showed a neutrophil leucocytosis. Then a chest x-ray examination showed a right lower and middle lobe consolidation with a wedge-shaped area of increased density with apex at the hilum and the base towards the pleura (Fig. 2). The CT-angiography highlighted an apparently right-sided aortic arch (Fig. 3, 4) with an area of increased density of pervious bronchi on the middle lobe, which refers to as subtotal atelectasis as middle lobe syndrome (MLS). Vascular ring was so diagnosed and for this reason the patient has later undergone repair surgery, which was followed by improvement of the symptoms.

Fig. 2. Antero-posterior chest radiograph showing a right lower and middle lobe consolidation.

Fig. 3. Sagittal view of CT showing the vascular ring.

Fig. 4. Coronal view of CT showing the double aortic arch.

Discussion
We reported three clinical cases of double aortic arch with three different dominant clinical manifestations: dysphagia with reflux and choking episodes in the first one; stridor, cough and episodes of regurgitation in the second one; recurrent respiratory infections and middle lobe syndrome in the third one; confirming the heterogeneity of clinical manifestations of double aortic arch.
In general, patients with a double aortic arch tend to have symptoms at an earlier age than do patients with other types of vascular ring. The compression of the trachea and/or the oesophagus causes the onset of respiratory and/or gastrointestinal symptoms respectively 6-7.
Our third clinical case is important because we describe for the first time in literature the simultaneous presence of the MLS and double aortic arch.
The MLS generally refers to atelectasis in the right middle lobe of the lung. Pathophysiologically, there are two forms of MLS, namely obstructive and non-obstructive. Obstructive MLS can be caused by extrinsic compression of the middle lobe bronchus or by endobronchial lesions. Aspirated foreign bodies and recurrent aspiration represent the main causes of intrabronchial obstruction in childhood8.
The middle lobe syndrome and double aortic arch are rare and usually isolated congenital malformations. However, the third described patient showed a MLS associated to the presence of double aortic arch. To date, the coexistence of these malformations it has been ever described, providing to paediatricians the proof that these entities can also coexist in the same patient. However, actually, we can not clarify whether this association is “casual” or “causal”.
In our case, we hypothesized that the MLS was originally caused by obstruction from the vascular ring but it has been perpetuated over time due to recurrent respiratory infections and aspiration. A long-term follow-up, during which the child will pursue a routine airway clearance regimen with aerosol therapy and chest physiotherapy, will give us the confirmation of this association.
The diagnosis of a child with vascular ring should proceed in a stepwise fashion without obtaining excessive studies. Since most patients with recurrent or persistent respiratory problems have already had chest radiographs, a careful review of these films may be helpful. A barium esophagram historically is the study used to diagnose vascular ring and still today a double aortic arch may be suggested by bilateral incidentations, usually at slightly different levels, in addition to a large posterior incidentation6. Cross-sectional imaging with CT-angiography or magnetic resonance angiography are then crucial for the definitive diagnosis and surgical management9. Echocardiography sometimes provides accurate diagnostic imaging of anomalies such as double aortic arch by visualization of both aortic arches but it is not the gold standard of the diagnosis of vascular ring. Finally, bronchoscopic examination would show extrinsic pulsatile compression of the trachea 3.
Surgery to correct a double aortic arch is the only treatment in symptomatic patients and is reportedly well tolerated 10.
The presentation of our cases confirms the clinical heterogeneity of this vascular malformation and, to date, in the best of our knowledge, it has been ever described the coexistence of middle lobe syndrome and double aortic arch in the same patient. In conclusion, although it remains to still elucidate whether this association is “casual” or “causal”, we suggest to also investigate this vascular malformation in patients with MLS.



References
1.    Backer CL, Mongé MC, Popescu AR, et al. Vascular rings. Seminars in Pediatric Surgery. 2016; 25:165-75
2.    Seo HK, JE HG, Kang IS, et al.  Prenatal double aortic arch presenting with a right aortich arch and an anomalous artery arising from the ascending aorta. International Journal of Cardiovascular Imaging. 2010;26:165-8
3.    Turner A, Gavel G, Coutts J. Vascular rings: presentation, investigation and outcome. European Journal of Pediatrics. 2005;164:266-70
4.    Becit N, Erkut B, Karaca Y. Vascular ring: tracheoesophageal compression associated with symmetrical double aortic arch. Texas Heart Institute Journal. 2008;35:209-10
5.    Backer CL, Mavroudis C, Rigsby CK, et al. Trends in vascular ring surgery. Journal of Thoracic Cardiovascular Surgery. 2005;129:1339-47
6.    Licari A, Manca E, Rispoli A, et al. Congenital vascular rings: a clinical challenge for the pediatrician. Pediatr Pulmonology. 2015;5:511-24
7.    Krishnasarma R, Golan Mackintosh LG, Bynum F. ALTE and feeding intolerance as a presentation of double aortic arch. Case reports in Pediatrics. 2016;2016:8475917
8.    Romagnoli V, Priftis KN, De Benedictis FM. Middle lobe syndrome in children today. Pediatric Respiratory Reviews. 2014;14:188-93
9.    Etesami M, Ashwath R, Kanne J, et al. Computed tomography in the evaluation of vascular rings and slings. Insights into Imaging. 2014;5:507-21
10.    Bonnard A, Auber F, Fourcade L. Vascular ring abnormalities: a retrospective study of 62 cases. Journal od Pediatric Surgery. 2003;38:539-43
 

geneticapediatrica.it trimestrale di divulgazione scientifica dell'Euromediterranean Paediatric Foundation
Legge 7 marzo 2001, n. 62 - Registro della Stampa Tribunale di Messina n. 3/09 - 11 maggio 2009

geneticapediatrica.it trimestrale di divulgazione scientifica dell'Euromediterranean Paediatric Foundation
Legge 7 marzo 2001, n. 62 - Registro della Stampa Tribunale di Messina n. 3/09 - 11 maggio 2009