Insulin gene promoter methylation status in Greek children and adolescents with Type 1 Diabetes.

The insulin (INS) gene is the one of the most important genes involved in the pathogenesis of Type 1 Diabetes (T1D) after the Major Histocompatibility Complex genes. Studies addressing the issue of hyper- or hypo-methylation status of the INS gene promoter have reported inconsistent results. The majority of studies showed hypomethylation; however a few studies have shown hypermethylation at specific cytosine-guanosine (CpG) sites in the promoter region of the INS gene. The aim of the present study was to analyze the methylation status of the promoter region of the INS gene in Greek children and adolescents with T1D. A total of 20 T1D participants (mean diabetes duration of 6.15±4.12 years) and 20 age- and sex-matched controls were enrolled in the present study. DNA was isolated from whole blood samples, modified using sodium bisulfite and analyzed using PCR and electrophoresis. DNA was then pooled with highly reactive supermagnetic beads at similar molar quantities, submitted for library construction and finally sequenced using next-generation sequencing. The methylation profile at 10 CpG sites around the transcription start site (TSS) of the INS promoter was analysed and expressed as the mean ± standard deviation. The overall mean methylation in patients with T1D did not differ compared with the healthy controls. There was a statistically significant difference between the two groups in hypermethylation at position -345 (P=0.02), while a trend (P=0.06) at position -102 was observed. According to the results of the present study, increased methylation in the INS gene promoter at specific CpG sites around the TSS were already present in childhood T1D. These data may possibly serve as a guide towards the identification of a methylation pattern for detection of development of T1D in genetically predisposed children.

DNA methylation analysis within the IL2RA gene promoter in youth with autoimmune thyroid disease.

BACKGROUND: Alpha-subunit of the interleukin-2 receptor (IL2RA) is involved in the regulation of T-cell function and has been related to autoimmune thyroid disease (AITD). Although the exact mechanisms are not fully understood, promoter methylation might account for differences in gene expression. The aim of this study was to investigate whether there are differences in the percentage of DNA methylation within the IL2RA gene promoter in young patients with AITD. MATERIALS AND METHODS: In a cross-sectional design, the presence of DNA methylation in the IL2RA gene promoter was quantified, by real-time PCR and melting curve analysis, in modified genomic DNA isolated from blood samples of a total of 149 children and adolescents with AITD, including patients with Hashimoto thyroiditis (ΗΤ) (n = 60), Graves' disease (GD) (n = 9), concurrent diagnosis of HT and type 1 diabetes (T1DM + HT) (n = 25), and healthy controls (n = 55). RESULTS: The percentage of DNA methylation in the IL2RA gene promoter was significantly decreased in patients with GD (26.0 ± 4.2%) but not in those with HT (36.3 ± 1.4%) in comparison with controls (41.3 ± 1.5%). CONCLUSIONS: The observed DNA hypomethylation in the IL2RA gene promoter in patients with GD might be related to its increased expression, thus contributing to the etiopathogenesis of GD in childhood and adolescence.

l-selenomethionine supplementation in children and adolescents with autoimmune thyroiditis: A randomized double-blind placebo-controlled clinical trial.

WHAT IS KNOWN AND OBJECTIVE: Although a beneficial effect of selenium (Se) administration has been proposed in adults with autoimmune thyroiditis (AT), there is a paucity of similar data in children and adolescents. The purpose of the study was to investigate whether administration of a high dose of organic Se (200 µg daily as l-selenomethionine) has an effect on antithyroid antibody titres in children and adolescents with AT. METHODS: Seventy-one (71) children and adolescents, with a mean age of 11.3 ± 0.3 years (range 4.5-17.8), diagnosed with AT (antibodies against thyroid peroxidase [anti-TPO] and/or thyroglobulin [anti-Tg] ≥60 IU/mL, euthyroidism or treated hypothyroidism and goitre in thyroid gland ultrasonography) were randomized to receive 200 µg l-selenomethionine or placebo daily for 6 months. Blood samples were drawn for measurement of serum fT4, TSH, anti-TPO and anti-Tg levels, and thyroid gland ultrasonography was performed at the entry to the study and after 6 months of treatment. RESULTS AND DISCUSSION: At the end of the study, a statistically significantly higher reduction in anti-Tg levels was observed in the Se group compared to the placebo group (Δ: -70.9 ± 22.1 vs -6.7 ± 60.6 IU/mL, P = 0.021). Although anti-TPO levels were also decreased in the Se group, this change was not statistically different from that of the control group (Δ: -116.2 ± 68.4 vs +262.8 ± 255.5 IU/mL, P = 0.219). No significant difference in thyroid gland volume was observed between the two study groups (P > 0.05). WHAT IS NEW AND CONCLUSION: In this original study, organic Se supplementation appears to reduce anti-Tg levels in children and adolescents with AT.

Seasonality of month of birth in children and adolescents with autoimmune thyroiditis: a continuing conundrum.

Background The aim of this study was to analyze the seasonal birth month pattern in young patients with autoimmune thyroiditis and compare it with youth controls. Methods Medical records of a total of 298 children and adolescents of Greek origin, with a diagnosis of Hashimoto thyroiditis (HT) before the age of 21 years that were born from 1987 to 2010 were retrospectively reviewed. In addition, 298 consecutive subjects that were born from 1988 to 2012 and evaluated in a tertiary unit for any reason, served as controls, provided that they had no personal or family history of thyroid or any other autoimmune disease. Results Significant differences were found between children and adolescents with HT and healthy controls in the yearly pattern of month of birth distribution (p=0.029). During month-by-month analysis, it was shown that the highest and lowest predispositions to HT were among those born in spring (March) (odds ratio [OR] 2.34, p=0.005), and autumn (November) (OR 0.49, p=0.035), respectively. A binary logistic regression model also revealed that season of birth and sex were the only factors that remained related to HT disease, even after adjustment for confounding factors such as year of birth and age (p<0.001, Nagelkerke r-square 0.151). Conclusions This study suggests that the effect of certain seasonal factors during fetal development, reflected by the seasonal differences in birth pattern, in children and adolescents with autoimmune thyroiditis could contribute to long-term programming of an autoimmune response against the thyroid gland. Further studies are needed to demonstrate a clear cause and effect relationship between month of birth and HT.

Psychological Aspects of Androgen Insensitivity Syndrome: Two Cases Illustrating Therapeutical Challenges.

Androgen Insensitivity Syndrome (AIS) and its heterogeneous phenotypes comprise the pieces of a challenging clinical problem. The lack of standardized guidelines results in controversies regarding the proper diagnostic and therapeutic approach, including the time and type of intervention. Due to its variable phenotype, AIS is not diagnosed at the proper age that would allow optimal psychological and medical support to the patient. Therapeutic approaches are not established, mainly due to the rarity of the disease. In addition, various social and ethical consequences may emerge. The aim of this double case report is to outline the difficulties that may rise during diagnostic, therapeutic, and psychological approach of AIS, especially concerning the handling of the relatives' reaction.

Prevalence of selective immunoglobulin A deficiency in Greek children and adolescents with type 1 diabetes.

BACKGROUND: The association of selective immunoglobulin A (IgA) deficiency with type 1 diabetes (T1D) remains unclear. This study was to evaluate serum IgA concentrations in Greek children and adolescents with T1D. METHODS: In two hundred individuals with T1D, serum IgA concentrations were quantitatively determined using nephelometry. RESULTS: Immunoglobulin A deficiency was detected in 6 (3.0%) of 200 patients who were subjected to immunological evaluation. Recurrent infections were not recorded, but human papilloma virus infection was clinically suspected and confirmed by laboratory examination in a 5-year-old girl. In regard to coincidence of selective IgA deficiency with autoimmune diseases, celiac disease was detected in a girl and juvenile idiopathic arthritis in a boy. Serum IgA concentrations differed significantly when patients were grouped according to age at the beginning of the study (P<0.001), age at diagnosis of T1D (P=0.015) and coincidence of celiac disease (CD) (P=0.038). However, when the age of the patients was adjusted, difference in serum IgA concentrations was not statistically significant despite CD was present or not. Moreover, serum IgA concentrations were positively correlated with serum IgG (P<0.001) and IgE (P=0.001) concentrations and negatively correlated with serum antigliadin antibody IgG (P=0.035) concentrations. There was no association or correlation of serum IgA concentrations with glycemic control. CONCLUSION: The prevalence of selective IgA deficiency in Greek children and adolescents with T1D is high (3.0%). The correlation of serum IgA concentrations with serum IgG, IgE and anti-gliadin antibody IgG concentrations needs further investigation.

Secondary nocturnal enuresis related to central diabetes insipidus as an early manifestation of intracranial germinomatous germ cell tumors in a series of male youngsters.

Nocturnal enuresis is a common symptom in children. It is usually attributed to benign causes and diagnostic evaluation is not carried out. We report three male young patients initially presenting with short stature and nocturnal enuresis, related to diabetes insipidus, caused by intracranial germinomatous germ cell tumors. In all three cases, water deprivation tests confirmed diabetes insipidus. Extensive endocrinological investigation also showed further hormone deficiencies. Magnetic resonance imaging of the brain revealed the presence of a central nervous system lesion and histology confirmed the final diagnosis. Surgery, radiation with or without chemotherapy was conducted and the patients were treated with hormone replacement therapies. The patients after a long follow-up were free of disease. We present these cases to alert clinicians to bear in mind that the presence of an intracranial germinomatous germ cell tumor should at least be considered in a child presenting with bed wetting, especially if additional symptoms and signs, including late onset puberty and growth delay or morning hypernatremia, may coexist.

Suboptimal glycaemic control enhances the risk of impaired prothrombotic state in youths with type 1 diabetes mellitus.

OBJECTIVE: To estimate markers of prothrombotic state and endothelial dysfunction in youths with type 1 diabetes mellitus (T1DM) and investigate possible associations with anthropometric/demographic data, glycaemic control and lipid profile. METHODS: In a cross-sectional design, we recruited 155 youths with T1DM and determined levels of plasminogen activator inhibitor-1-antigen (PAI-1-Ag), von Willebrand factor-antigen (vWF-Ag), fibrinogen (FB), lipids and glycosylated haemoglobin (HbA1c). RESULTS: Of all the participants, 76 (49%) had increased levels of at least one of prothrombotic factors. Suboptimal glycaemic control was associated with a worse lipid profile and an eightfold increased risk of elevated vWF-Ag levels. Higher vWF-Ag concentrations were also correlated with impaired lipid profile and increased HbA1c values, whereas PAI-1-Ag was positively correlated only with triglyceride levels. After adjustment for potential confounders, only HbA1c contributed independently to the variation in vWF-Ag levels. CONCLUSION: Impaired prothrombotic state and consequently endothelial dysfunction are present in youths with T1DM, representing a cumulative risk factor for future cardiovascular disease (CVD). Achievement and maintenance of euglycaemia and normolipidaemia are crucial to decelerate progress of this process.

An unexpected cause of vaginal bleeding: the role of pelvic radiography.

Vaginal bleeding and/or discharge in young girls may result from infection, urological problems, endocrine causes, bleeding disorders, dermatological conditions, trauma, sexual abuse, masses or foreign bodies. We report a case of excessive vaginal bleeding caused by a foreign body in a prepubertal girl with emphasis on the diagnostic challenges and pitfalls regarding imaging techniques. In our patient, although invasive and expensive investigations had been initially made, the foreign body was last detected only when a plain pelvic radiography was performed.

Association between magnesium concentration and HbA1c in children and adolescents with type 1 diabetes mellitus.

BACKGROUND: Magnesium levels may be decreased in patients with type 1 diabetes mellitus (T1DM), influencing disease control. Relevant studies concern mainly adults and there are few data from the pediatric population. The aim of the present study was to evaluate magnesium levels and examine their possible association with glycemic control in youths with T1DM. METHODS: In all, 138 children and adolescents with T1DM aged between 1.9 and 20.3 years were recruited to the study. Using a cross-sectional design, we measured anthropometric parameters, HbA1c, serum magnesium, ionized and total calcium, phosphorus, potassium, sodium, and urinary albumin (UA). Estimated glomerular filtration rate (eGFR), based on serum creatinine concentrations, was also calculated. RESULTS: Lower levels of magnesium were found in subjects with poor versus good glycemic control (0.79 ± 0.09 vs 0.82 ± 0.09 mmol/L, respectively; P = 0.002). Serum magnesium levels were negatively correlated with HbA1c (P < 0.001) and positively correlated with UA, calcium, phosphorus, and potassium levels (P < 0.05). After adjustment for confounding factors, only magnesium levels remained significantly associated with HbA1c (adjusted r(2) = 0.172; P = 0.004). The odds ratio for poor glycemic control, indicated by HbA1c >7.5%, between the highest and lowest magnesium concentration quartiles was 0.190 and amounted to a decrease of 1.7% in the HbA1c level. CONCLUSIONS: The present study shows that low serum magnesium levels in children and adolescents with T1DM are associated with an increased risk of poor glycemic control, potentially contributing to the early development of cardiovascular complications.

Children and adolescents with type 1 diabetes mellitus have a sixfold greater risk for prolonged QTc interval.

BACKGROUND: QT-wave abnormalities have been detected in type 1 diabetes mellitus (T1DM). Prolongation of the heart rate corrected QT interval (QTc) has been associated with cardiovascular mortality. We evaluated how often QT/QTc abnormalities are present in youth with T1DM and if they are associated with disease parameters. METHODS: Sixty-two T1DM youngsters and equal age- and gender-matched controls were studied. Demographic, anthropometric, and laboratory data were determined. QT was measured on a 12-lead resting electrocardiogram. QTc was calculated using Bazett's formula. RESULTS: T1DM patients had significantly longer QT/QTc than controls, but significance disappeared after adjustment for confounders. Abnormally prolonged QTc≥440 ms was observed six times more frequently in those with T1DM. QT was correlated with age, age at disease onset, but not with glycated hemoglobin or diabetes duration; QTc was only correlated with pubertal stage. CONCLUSIONS: T1DM youths have a sixfold increased risk for QT/QTc prolongation and should have regular follow-up for cardiac autonomic dysfunction.

Ghrelin-leptin network influences serum chitinase 3-like protein 1 (YKL-40) levels in obese prepubertal children.

OBJECTIVE: This study aimed to investigate any possible interactions between hormonal regulators of weight gain and markers of subclinical inflammation in childhood obesity. Forty-one obese prepubertal children and 41 age- and gender-matched lean controls were included. Children were classified as obese or non-obese according to international age- and gender-specific body mass index (BMI) cutoff points defined by the International Obesity Task Force to define childhood obesity. Anthropometric measurements, serum insulin, chitinase 3-like protein (YKL-40), ghrelin and leptin levels as well as plasma glucose in the fasting state were determined. RESULTS: Obese children as compared with controls had higher YKL-40 (50.7±15.2 vs 41.0±10.5 ng/ml, p=0.003), higher leptin (33.8±16.0 vs 9.7±7.5 ng/ml, p<0.001) and lower ghrelin serum levels (871.4±368.0 vs 1417.6±387.3 pg/ml, p<0.001). The obese children with ghrelin levels above median (43.8±10.2 ng/ml) as compared to those with ghrelin below median (57.2±16.6 ng/ml) presented lower serum YKL-40 levels (p=0.009), indicating more severe inflammation with lower levels of ghrelin. By contrast, although the obese children with leptin levels above median (49.7±16.3 ng/ml) presented lower serum YKL-40 levels as compared to those with leptin levels below median (51.6±14.6 ng/ml), this difference did not reach the level of statistical significance (p=0.726). Moreover, serum YKL-40 levels were significantly correlated with ghrelin (r=-0.359, p=0.014) but not with leptin levels (r=0.169, p=0.261). A significant negative correlation between ghrelin and leptin levels was also found (r=-0.276, p=0.041). These findings remained unchanged for obese, when analyses were done separately, whereas the significance of correlations was lost for non-obese subjects. CONCLUSIONS: Ghrelin-leptin network had an impact on serum YKL-40 levels in obese prepubertal children; upregulation of YKL-40 secretion seems to be a consequence of reduced ghrelin rather than elevated leptin concentrations.

Elevated circulating levels of the serum acute-phase protein YKL-40 (chitinase 3-like protein 1) are a marker of obesity and insulin resistance in prepubertal children.

YKL-40 (chitinase 3-like protein 1) is a newly recognized protein that is secreted by activated macrophages and neutrophils and expressed in a broad spectrum of inflammatory conditions and cancers. It has also been associated with endothelial dysfunction and diabetes in adults. Its role in childhood obesity has not been evaluated yet. Our aim was to evaluate the associations of serum YKL-40 levels with markers of obesity, inflammation, and insulin resistance in children. Forty-one obese prepubertal children and 41 age- and sex-matched lean controls were included, and serum YKL-40 levels were determined. Body mass index (BMI), blood pressure (BP), body fat percentage, fasting glucose, insulin, homeostasis model assessment for insulin resistance (HOMA-IR) index, whole-body insulin sensitivity index, lipids, white blood cell (WBC) count, C-reactive protein, and fibrinogen levels were also assessed. Obese children had higher YKL-40 levels compared with controls (P = .003). Insulin-resistant individuals showed higher YKL-40 compared with non-insulin-resistant individuals after adjusting for age and BMI (adjusted P = .039). Serum YKL-40 levels were positively correlated with age, BMI, body fat percentage, fasting glucose and insulin, HOMA-IR index, whole-body insulin sensitivity index, systolic BP, mean BP, and WBC count (P < .05). After adjustment for age, sex, BMI, WBC count, and systolic BP, HOMA-IR index remained significantly associated with YKL-40 levels (P < .001). The study suggests that YKL-40 levels are elevated in obese youth and represent a marker of insulin resistance even in childhood. Prospective studies are needed to determine whether children with elevated YKL-40 levels are at higher risk for future cardiovascular disease.

Growth hormone deficiency: an unusual presentation of floating harbor syndrome.

Floating-Harbor Syndrome (FHS) is a very rare condition of unknown etiology characterized by short stature, delayed bone age, characteristic facial features, delayed language skills and usually normal motor development. This syndrome has only once been associated with growth hormone deficiency and precocious puberty in the same patient. We describe a 5 4/12 year-old girl with the typical features of FHS in whom growth hormone deficiency was diagnosed and two years later central precocious puberty was noted. The patient showed a good response to human recombinant growth hormone as well as gonadotropin releasing hormone analogue treatment.

Insulin resistance is associated with at least threefold increased risk for prothrombotic state in severely obese youngsters.

UNLABELLED: Obesity in childhood increases the risk for early adult cardiovascular disease. However, the underlying mechanism is not fully known. The aims of this study were to measure levels of prothrombotic factors and examine their possible association with obesity and insulin resistance in obese children and adolescents. A total of 313 obese children and adolescents were recruited. In a cross-sectional design, we measured anthropometric parameters, plasminogen activator inhibitor-1-antigen (PAI-1-Ag), von Willebrand factor-antigen (vWF-Ag), fibrinogen (FB), lipids, fasting glucose, and insulin (FI) levels. Insulin resistance was estimated using the homeostasis model assessment for insulin resistance (HOMA-IR) index. Boys presented significantly higher PAI-1-Ag levels than girls (82.6 vs. 71.3 ng/ml, p = 0.01). Higher levels of PAI-1-Ag (96.8 vs. 69 ng/ml, p < 0.001), vWF-Ag (123.5 vs. 107.6%, p = 0.004) but not FB (353.1 vs. 337.6 mg/dl, p = 0.137) were found in insulin-resistant (IR) participants after adjusted for age, gender, and pubertal stage. IR patients were at 2.98 (CI: 1.084-8.193) and 4.86 (CI: 1.119-15.606) times greater risk for high PAI-1-Ag and vWF-Ag levels, respectively. All three prothrombotic factors were positively correlated with body mass index (BMI) and FI levels (p < 0.05), but only PAI-1-Ag and vWF-Ag were significantly correlated with HOMA-IR index (p ≤ 0.001). After adjustment for confounding factors, both BMI and HOMA-IR indices remained significantly associated with PAI-1-Ag (r2 = 0.225, p < 0.001) and vWF-Ag levels (r2 =0.077, p = 0.003). CONCLUSION: This study shows that obesity in youngsters, when accompanied with insulin resistance, is associated with at least threefold increased risk for elevated levels of prothrombotic factors, contributing to the early development of atherothrombosis. This impaired prothrombotic state may partially explain the increased risk for developing cardiovascular disease later in adulthood.

Acquired toxoplasmosis accompanied by facial nerve palsy in an immunocompetent 5-year-old child.

Acquired toxoplasmosis, although relatively common in children, is usually asymptomatic but can also be clinically manifested by a benign and self-limited infectious mononucleosis-like syndrome. Neurological complications are very rare in immunocompetent children. The authors report a 5-year-old boy who presented with cervical lymphadenopathy because of acquired toxoplasmosis accompanied with unilateral facial nerve paralysis. Toxoplasma gondii DNA detection in blood by polymerase chain reaction, as well as elevated specific immunoglobulin M antibodies against it, established the diagnosis. Characteristic brain lesions on magnetic resonance imaging were absent and ophthalmologic examination revealed no inflammatory lesions in the retina and choroid. Treatment with pyrimethamine, sulfadiazine, and folic acid resulted in a complete recovery after 2 months of therapy. Although rare, acute facial nerve paralysis of unknown origin can be caused by acquired toxoplasmosis even in the immunocompetent pediatric population. Elevated titers of specific antibodies and the presence of parasite's DNA are key findings for the correct diagnosis.