Mitochondria in human reproduction: novel paradigm in the onset of neurodegenerative disorders.

The disease progression of neurodegenerative disorders (NDD), including Alzheimer's, Parkinson's and Huntington's disease, is inextricably tied to mitochondrial dysfunction. However, although the contribution by nuclear gene mutations is recognised for familial onset of NDD, the degree to which cytoplasmic inheritance serves as a predetermining factor for the predisposition and onset of NDD is not yet fully understood. We review the reproductive mechanisms responsible for ensuring a healthy mitochondrial population within each new generation and elucidate how advanced maternal age can constitute an increased risk for the onset of NDD in the offspring, through the increased heteroplasmic burden. On the one hand, this review draws attention to how assisted reproductive technologies (ART) can impair mitochondrial fitness in offspring. On the other hand, we consider qualified ART approaches as a significant tool for the prevention of NDD pathogenesis.

Immunoexpression of type-1 adiponectin receptor in the human intestine.

UNLABELLED: Adiponectin is an important biomarker of metabolic syndrome that was recently identified in human breast milk. We demonstrate the presence of type-1 adiponectin receptor (adipoR1) by immunoperoxidase method in 21 bioptic specimens - duodenum (n = 6), terminal ileum (n = 7) and colon (n = 8) from 14 human subjects (6 females and 8 males aged 9 months-47 years). In all the samples, adipoR1 was detected. The positivity was observed in enterocytes and colonocytes as well as in lymphocytes in the submucosa and in the smooth muscle of the intestinal wall. Thus, adiponectin may influence intestinal physiology through its type-1 receptor. KEYWORDS: adiponectin - adiponectin receptor - intestine - nutritional programming - breast milk.

Role of common canalicular transporter gene variations in aetiology of idiopathic gallstones in childhood.

Variations in genes encoding canalicular transportes, for biliary lipids may affect concentrations of biliary lipids in bile and promote cholesterol crystallization and gallstone formation. In our study we investigated the contribution of heterozygosity for common variations considered either potentially pathogenic or susceptibility alleles for cholesterol cholelithiasis in adults (c.523A>G (p.Thr175Ala) and c.1954A>G (p.Arg652Gly) in ABCB4, c.1331T>C (p.Val444Ala) in ABCB11 and c.55 G>C (p.Asp19His) in ABCG8) to the aetiology of paediatric idiopathic gallstone disease. Genotyping was performed in 35 paediatric subjects with idiopathic gallstones with positive family history for gallstones and 150 population controls. The ABCB4 variant p.Thr175Ala was found only in the controls, not in the patients. The frequency of the remaining three variant alleles and the corresponding genotypes did not differ between patients and controls. We conclude that the studied common variations in genes encoding canalicular transporters known to contribute to genetic predisposition to cholesterol gallstones in adulthood do not contribute specifically to the aetiology of paediatric idiopathic gallstones.

Intestinal microbiota in exclusively breast-fed infants with blood-streaked stools.

Intestinal microbiota in exclusively breast-fed infants with blood-streaked stools and in healthy exclusively breast-fed babies was compared. Total anaerobes, bifidobacteria, lactobacilli, coliform bacteria, enterococci and clostridia were quantified by cultivation methods in feces of 17 full-term exclusively breastfed patients (aged 16.3 +/- 7.4 weeks) with blood-streaked stools and in the control group of 22 healthy fullterm exclusively breast-fed infants (13.7 +/- 6.4 weeks). Specific fluorescence in situ hybridization kits for Bifidobacterium spp. were used for the quantitative detection of bifidobacteria in samples. Control samples had significantly (p < 0.05) higher counts of total anaerobes. Bifidobacteria were not detected in patients' samples in 65 % and in controls in 36 % (p < 0.01). Bifidobacteria counts were also significantly higher in the control group (p < 0.01). Furthermore, clostridia strains were detected only in feces from bifidobacteria-negative infants reaching counts >8 log CFU/g. Lactobacilli were not detected in 65 % patients and in 45 % control samples. However, this difference was not significant as well as the difference in lactobacilli counts. Eosinophilia was observed in 35 % of patients, low IgA concentration in 71 % and also low IgG concentration in 71 %. pANCA positivity was found in 53 % of patients. In conclusion a significant low proportion of bifidobacterial microbiota in patients with blood-streaked stools was shown in comparison with controls.

Infliximab dependency in children with Crohn's disease.

BACKGROUND: Recently, infliximab dependency has been described. AIM: To assess frequency of ID in 82 consecutive Crohn's disease children treated with infliximab 2000-2006 and to describe clinical and genetic predictors of long-term infliximab response. METHODS: A phenotype model of infliximab dependency was used to assess treatment response: 'immediate outcome' (30 days after infliximab start)--complete/partial/no response. 'Long-term outcome': (i) prolonged response: maintenance of complete/partial response; (ii) infliximab dependency: relapse < or = 90 days after intended infliximab cessation requiring repeated infusions to regain complete/partial response or need of infliximab >12 months to sustain response. Polymorphisms TNF-308 A>G, TNF-857 C>T, Casp9 93 C>T, FasL-844 C>T, LTA 252 C>T and CARD15 (R702W, G908R, 1007fs) were analysed. RESULTS: Ninety-four per cent of children obtained complete/partial response. In long-term outcome, 22% maintained prolonged response, 12% had no response, while 66% became infliximab dependent. Perianal disease and no previous surgery were associated with infliximab dependency (OR 5.34, 95% CI: 1.24-22.55; OR 6.7, 95% CI: 1.67-26.61). No association was found with studied polymorphisms. The cumulative probability of surgery 50 months after starting infliximab was 10% in infliximab dependency, 30% in prolonged responders and 70% in nonresponders (P = 0.0002). CONCLUSIONS: Sixty-six per cent of children became infliximab dependent. Perianal disease and no surgery prior to infliximab were associated with infliximab dependency phenotype.

Comparison of intestinal microflora in healthy infants and infants with allergic colitis.

Twenty-eight exclusively breast-fed healthy infants and 16 infants also exclusively breast-fed with allergic colitis (aged 85 +/- 60 and 98 +/- 58 d, respectively) were screened for differences in fecal flora. Bifidobacteria were detected in 23 healthy infants and only in 4 fecal samples of infants with allergic colitis. All bifidobacteria-free infants possessed Gram-positive regular rods as a major group of their fecal flora. These bacteria were identified as clostridia using genus-specific FISH probe. Infants with allergy colitis possessed significantly lower counts of bifidobacteria and total anaerobes and significantly higher counts of clostridia in their feces. In healthy infants, Bifidobacterium longum was the most frequently found species (54.5% of the samples), followed by B. adolescentis (20.0), B. breve (18.2), B. bifidum (16.4), B. dentium (10.9) and B. pseudocatenulatum (1.80). Bifidobacterial isolates from two babies with allergic colitis were identified as B. longum, one child from patients group contained species B. dentium and one baby B. adolescentis. Our results suggest that there are significantly lower counts of bifidobacteria in infants with allergic colitis than in healthy infants.

Variants of CARD15, TNFA and PTPN22 and susceptibility to Crohn's disease in the Czech population: high frequency of the CARD15 1007fs.

Crohn's disease (CD) has been shown to be associated with the variants in the CARD15 gene as well as in other genes involved in the immune response. The frequencies of the variants profoundly differ among populations and so does the associated risk. We examined the associations of variants in the CARD15, TNFA and PTPN22 genes with pediatric-onset and adult-onset CD in the Czech population. Genotype, phenotype and allelic frequencies were compared between 345 patients with CD (136 pediatric-onset and 209 adult-onset patients) and 501 unrelated healthy controls. At least one minor allele of the CARD15 gene was carried by 46% patients and only 21% control subjects (OR = 3.2, 95% CI 2.4-4.4). In a multiple logistic regression model, the strongest association with CD was found for the 1007fs variant (OR = 4.6, 95% CI 3.0-7.0), followed by p.G908R (OR = 2.9, 95% CI 1.5-5.7) and p.R702W (OR = 1.7, 95% CI 1.0-2.9), while no independent association was found for the remaining variants in the CARD15 gene (p.268S, p.955I and p.289S), for the p.R620W variant in the PTPN22 gene or for the g.-308G>A variant in the TNFA gene. The age at CD onset was strongly modified by positivity for the 1007fs allele: it was present in 42% pediatric-onset and only 25% adult-onset patients. In conclusion, we report a high frequency of the minor allele of the CARD15 1007fs polymorphism in the Czech population and a strong effect of this allele on the age at disease onset.

Dynamic changes of orexin A and leptin in obese children during body weight reduction.

In this study, we describe changes of plasma levels of the hypothalamic neuropeptide orexin A in obese children during the reduction of body weight and its relationship to other biochemical and anthropometrical parameters. We measured orexin A fasting plasma levels by the RIA method in 58 obese children--33 girls and 25 boys; mean age 13.1+/-0.38 years (range 7-18.5) before and after 5 weeks of weight-reduction therapy. Leptin, IGF-1, and IGFBP-3 levels were measured in all the subjects and were compared to orexin A levels and anthropometrical data. Average weight in subjects before weight-reduction was 74.2+/-2.79 kg and after weight-loss 67.4+/-2.60 kg (p<0.0001). Orexin A levels before the therapy were 33.3+/-1.97 pg/ml and after the therapy 51.7+/-3.07 pg/ml (p<0.0001). Levels of orexin A were not significantly different between girls and boys (p=0.7842). We found negative correlation between orexin A and age (r = -0.5395; p<0.0001), body height (r = -0.4751; p=0.0002), body weight (r = -0.4030; p=0.0017) and BMI (r = -0.2607; p=0.0481). No correlation was found between orexin A and IGF-1, IGFBP-3 or leptin. Orexin A plasma levels increased during body weight loss, whereas the reverse was true for leptin levels. These findings support the hypothesis that orexin A may be involved in regulation of nutritional status in children.

Detection of infant faecal bifidobacteria by enzymatic methods.

An enzyme-based assay was developed for the detection of bifidobacteria in infant faeces. Ninety-five samples from 51 breast-fed infants in the age between 3 and 276 days were investigated. Bifidobacteria and other bacterial groups were determined by cultivation and fluorescence in situ hybridisation (FISH). Faecal samples were examined for the activity of fructoso-6-phosphate phosphoketolase (F6PPK) and for other enzymatic reactions using the API-ZYM kit. Twenty-nine infants had high numbers of bifidobacteria (usually higher than 9 log CFU/g) in their faeces. Seventeen infants (35%) did not contain detectable amounts of bifidobacteria in their faecal samples. The remaining five individuals had low counts of bifidobacteria (3-6 log CFU/g). Most negative infants possessed major amounts of clostridia in their faecal flora. There were no significant differences among bifidobacterial counts obtained by cultivation and FISH, detection of F6PPK, alpha-galactosidase and alpha-glucosidase activities could routinely be used for the rapid and simple detection of bifidobacteria in infant faecal samples. Bifidobacterial colonies were identified using enzymatic tests and PCR procedure based on 16S rRNA gene sequences species-specific primers. In 14 samples, the identifications of individual isolates were compared with direct analyses of faeces using the nested PCR-denaturing gradient gel electrophoresis (nested DGGE) procedure. The results obtained in several cases are not identical. Bifidobacterium longum and Bifidobacterium breve were most frequently identified. Bifidobacteria-positive samples had high activities of alpha-galactosidase and alpha-glucosidase. On the contrary, negative samples missed either one or both of these enzymatic activities. While all positive samples tested showed distinctive fructose-6-phosphate phosphoketolase activity (F6PPK), none of the negative samples expressed F6PPK activity.

Serum alpha-glutathione S-transferase as a sensitive marker of hepatocellular damage in patients with cystic fibrosis.

The aim of the study was to evaluate serum a-glutathione S-transferase (s-GSTA) levels in patients with cystic fibrosis (CF) and to compare s-GSTA with other liver function tests and with a hepatic ultrasound scan (US). The cytosolic enzyme, alpha-glutathione S-transferase is predominantly found in the liver and is distributed uniformly in the liver tissue. In our study s-GSTA levels were measured in 37 CF patients aged 1 to 28 years (mean age 10.4 years, 24 males). The control group consisted of 27 patients aged 2 to 17 years (mean age 8.5 years, 18 males). The presence of hepatobiliary abnormalities was assessed by clinical examination, ultrasound scan, s-GSTA, and conventional liver enzymes: alanine aminotransferase (ALT), alkaline phosphatase (ALP), aspartate aminotransferase (AST) and gama-glutamyl transferase (GMT). The calculated 5-95 % range of s-GSTA for the control group was 0.098-2.54 microg/l, for the CF group 0.43-9.76 microg/l. Mean s-GSTA level in the control group was 1.55 microg/l (S.D.=1.57), and 2.05 micro/l (S.D.=2.60) in the CF group. In the group of CF patients, the serum levels were significantly higher than in the control group (P<0.01). No significant correlation existed in the CF group between s-GSTA and conventional liver tests (ALT, AST, ALP and GMT). Four patients in the CF group had hepatobiliary abnormalities detectable by conventional liver tests, s-GSTA and US. Four patients had abnormal s-GSTA, while conventional liver tests and US were normal. One other patient had abnormal hepatic US, but normal standard liver tests and s-GSTA. The study has suggested that a raised s-GSTA level might be a marker of possible pathological changes of the hepatobiliar system in CF patients. Serum GSTA seems to be a more sensitive marker than transaminases for the monitoring of hepatocellular integrity and as an early predictor of hepatic damage.

[HLA-DRB1/DQA1/DQB1 alleles and haplotypes in Czech children with celiac sprue]. / HLA-DRB1/DQA1/DQB1 alely a haplotypy ceských deti s celiakální sprue.

BACKGROUND: The celiac disease (CD) is a multifactor disease resulting from a life time abnormal immune response to gluten accompanied by autoimmune characteristics, which can in sensitive individuals evoke small bowel mucosa morphologic changes. The genetically sensitive individual to CD has not been defined yet, it is obvious, however, that this illness is closely linked to the HLA class II genes. The objective of our study was to detect associations of HLA class II alleles and haplotypes DRB1/DQA1/DQB1 in Czech CD children. METHODS AND RESULTS: A group of Czech CD children diagnosed according to ESPGHAN criteria was genotyped HLA for alleles of DRB1/DQA1/DQB1 loci. Genotyping of the HLA-DRB1/DQB1 haplotypes proved statistically significant association CD with haplotypes and alleles of this genetic system. 92.9% of patients have in their HLA phenotype allele DQA1*0501 in either cis or trans configuration with the DQB1 allele *0201/*0202. The extended HLA haplotype DRB*0301/DQA1*0501/DRB1*0201 as well as the haplotype DRB1*0701/DQA1*0201/DQB1*0202, are presented in 63.6% or in 61.0% CD patients respectively. The individual HLA class II alleles DRB*0301, *0701, DQA1*0201, *0501, DQB1*0201, *0202 and the above mentioned HLA haplotypes inclusively provide genotypic frequencies significantly different from healthy Czech individuals (P < or = 0.06 +/- 0.001). These results support the opinion that the HLA molecule expressed on the cellular surface as a alpha beta heterodimer encoded by the DQA1*0501 and DQB1*0201/02 alleles in either cis or trans configuration is responsible for the primary sensitivity to this disease. We were, however, not able to find an association of various clinical forms of the CD with a certain HLA haplotype in the followed group. CONCLUSION: The CD patients have in comparison with healthy population significantly different frequency of HLA class II haplotypes. Though the finding of these alleles is not sufficient for an explicit confirmation of this diagnosis, the proof of this risky haplotype/s may notably contribute to it, namely in case of potential or latent forms of this disease.

Cow's milk-specific cellular and humoral immune responses and atopy skin symptoms in infants from atopic families fed a partially (pHF) or extensively (eHF) hydrolyzed infant formula.

BACKGROUND: Hydrolyzed milk formulas are recommended to feed infants at high risk of atopy if breast-feeding is not possible. We studied the specific cellular and humoral immune response to cow's milk proteins and occurrence of atopic dermatitis under different feeding regimens: two hydrolyzed infant milk formulas (partially [pHF] and extensively hydrolyzed [eHF]) and under exclusive breast-feeding (BF). METHODS: Seventy-two infants from families with atopic symptoms were randomized in the pHF and eHF groups, respectively. At 6 and 12 months of age, peripheral blood mononuclear cell proliferation along with specific IgG and IgE to cow's milk proteins was determined in infants fed pHF or eHF, respectively, and those who had not yet received any formula at 6 months of age (BF). Cases of atopic dermatitis were recorded throughout the first 12 months of life, and their severity was evaluated with SCORAD points. RESULTS: A significantly decreased proliferation to cow's milk caseins was found in the pHF group compared to the exclusively breast-fed group. Medians of stimulation indexes for CAS at 6 months were as follows: pHF 1.18; n=24; BF 1.70; n=24 (P=0.033, Mann-Whitney U-test). Higher levels of plasma IgG antibodies to BCAS were found in infants fed pHF than in those fed eHF at 12 months. Optical density (OD): (25th percentile; median; 75th percentile): pHF: 0.00; 0.14; 0.38; n=30; eHF: 0.00; 0.03; 0.14; n=28; P=0,089, Mann-Whitney U-test. Cow's milk-specific IgE was detected at 6 months as follows: BF: 3 of 24; eHF: 2 of 21; pHF: 0 of 23. The number of cases of atopic dermatitis and their severity did not differ among the groups during the first 12 months. CONCLUSIONS: Feeding pHF appears to suppress cow's milk-specific cellular responses and stimulate specific IgG production. Specific IgE sensitization can occur also with breast-feeding.

Increased levels of circulating ICAM-1, E-selectin, and IL-2 receptors in celiac disease.

Adhesive interactions between endothelium and circulating cells are crucial for the development of inflammatory reactions. We found significantly higher serum levels of soluble intracellular adhesion molecule-1 (sICAM-1, 492.5 +/- 22.1 ng/ml) in patients with active celiac disease (including IgA-deficient patients) than in patients on a gluten-free diet (335.7 +/- 20.0 ng/ml) (P < 0.001) and healthy controls (207.4 +/- 11.2 ng/ml) (P < 0.001). The concentration of soluble E-selectin in sera from celiac patients (37.2 +/- 3.4 ng/ml) was also higher (P < 0.001) than in sera from healthy controls (15.5 +/- 0.7 ng/ml) but, in contrast to sICAM-1, it remained high in the patients after treatment (30.2 +/- 2.7 ng/ml). Interestingly, the concentration of circulating soluble interleukin-2 receptors, molecules indicating lymphocyte activation, was only increased in sera from patients with active celiac disease (2943.0 +/- 214.1 pg/ml), and the level in sera from treated patients and healthy controls was comparable (1936 +/- 349 and 1416 +/- 111.7 pg/ml). The elevated serum level of soluble cell adhesion molecules could be used as a supplementary, noninvasive procedure for monitoring intestinal immune reactions.

[Growth in children with the exudative enteropathy syndrome due to a congenital heart defect--cor triatriatum dextrum]. / Rust u dítete se syndromem exsudativní enteropatie na podklade vrozené srdecní vady--cor triatriatum dextrum.

Retarded growth in a child can be the sign of serious chronic disease. The authors present an account of a six-year-old boy where growth retardation persisted at least from the age of three. During this period his height dropped from the zone between the 25th and 50th percentile into the zone between the 3rd and 10th percentile. From the clinical point of view a large abdomen, loose stools and hypocalcaemia with tetany were striking, as they were moreover refractory to vitamin D2, calcitriol and calcium administration by the oral route. The authors revealed severe hypoproteinaemia, a 150 times increased value of alpha-1-antitrypsin in faeces, and exudative enteropathy syndrome was diagnosed. The cause was venous congestion due to a rare heart disease--cor triatriatum dextrum. The septum in the right atrium was resected. Immediately after surgery the consistency and frequency of stool decreased. Calcaemia and plasma protein levels reached normal levels within two months. A growth spurt of 11 cm/year followed. Fifteen months after operation the patient's height reached almost the 50th percentile.

[Endoscopic sclerotherapy of esophageal varices in children with prehepatic portal hypertension]. / Endoskopická skleroterapie jícnových varixu u detí s prehepatální portální hypertenzí.

BACKGROUND: Endoscopic sclerotization of oesophageal varices in adult patients with prehepatic portal hypertension has become the method of choice in haemorrhage from these varicosities. The objective of the present work was to prove the effectiveness of this treatment in children. METHODS AND RESULTS: Between November 1987 and May 1993 in the authors' departments endoscopic sclerotization was used to treat 20 children (age 2.5-17 years) with bleeding oesophageal varices (o.v.) associated with prehepatic portal hypertension (PPH), caused by thrombosis of the portal vein. Half the children were treated unsuccessfully before sclerotherapy by surgery, some repeatedly. Complete eradication of o.v. was achieved in 19 children (95%). In the course of sclerotherapy before completed obliteration of all varices 5 children (25%) had another spell of haemorrhage. In three instances this early relapse of haemorrhage was controlled by another sclerotization. In one patient continuing haemorrhage from an oesophageal varix was treated by establishment of a splenorenal anastomosis. In another patient the source of haemorrhage were gastric varices which were ligatured and after the operation the authors proceeded with sclerotherapy. A relapse of o.v. during the average 3-year follow up period was recorded in 7 children (35%), while a relapse of haemorrhage from these neovarices occurred only in one child (5%). CONCLUSIONS: Successful eradication of oesophageal varices by sclerotherapy in 95%, and 95% successful prevention of relapsing haemorrhage from neovarices for a period of three years after surgery justify a change of tactics of treatment. The method of first choice should be always sclerotization of bleeding varices.

[Acute reversible liver necrosis after valproate therapy (immunologic hypersensitivity)]. / Akute Reversible Lebernekrose nach Valproat-Behandlung (immunologisch bedingte Idiosynkrasie).

We describe a case of a 9-year-old boy with the severe acute liver injury caused by idiosyncrasy after the administration of valproate. The liver biopsy performed during the first days after the onset of the disease revealed the necrosis of more than 50% of hepatocytes but the injury was almost fully reversible which was well demonstrated by the second biopsy five months later. After five years an evidence for the immunologic idiosyncrasy caused by valproate was still present, which was demonstrated in vitro by blastic transformation of lymphocytes from the patient. The boy is quite healthy and epilepsy is under control of suximide. The family was informed that the boy could be treated with valproate never more.

[Juvenile granulosa cell tumor with subsequent occurrence of gastrointestinal polyposis, subcutaneous lipomatosis and nodular goiter]. / Juvenilní nádor z bunek granulózy s následným výskytem polypózy zazívacího traktu, subkutánní lipomatózy a nodózí strumy.

A juvenile tumour from granulosa cells (M-8622/1), 13 x 8 x 6 cm, in the right ovary in a three-month-old girl produced some symptoms of pseudopubertas praecox isosexualis which disappeared after operation. Microscopic examination of the tumour revealed in addition to typical structures a less common differentiation to Sertoli cells. Despite actinotherapy and chemotherapy one and a half years after the onset of the disease X-ray examination revealed metastases in the lungs which were successfully cured by further doses of the above two types of treatment. Between the age of 6 and 15 years the girl developed successively polyposis of the stomach, small and large intestine (M-7564/0), subcutaneous lipomatosis of the trunk and left lower extremity (M-8881/0) and nodular goitre (M-7164/0), predominantly quiescent. In the literature a connection between gonadal stromal ovarian tumours and mesenchymal tumours, intestinal polyposis and disorders of the thyroid gland is described, but in different patients. The authors' observation is unique by the successive incidence of these changes in a single patient surviving 15 years after operation; and thus genetically conditioned associations could be involved.