Diagnosis, Clinical Presentation and Management of Celiac Disease in Children and Adolescents in Poland.

Celiac disease (CD) is a chronic immune-mediated disorder triggered by the ingestion of gluten in genetically predisposed individuals, affecting about 1% of the general population in the developed world. In 2012, the European Society for Pediatric Gastroenterology, Hepatology, and Nutrition (ESPGHAN) recommendations for CD diagnoses in children and adolescents were introduced, allowing the "no-biopsy" approach if certain criteria were met. This approach was also confirmed in the revised guidelines published in 2020. Thus, the aim of this study was to assess-over a one-year period-the clinical presentations and current status of the management of children and adolescents diagnosed with CD in Poland. Medical records of children and adolescents, newly diagnosed with CD in 2022/2023 in three medical centers in Poland, were involved. Gastroenterologists completed the specific anonymous web-based forms developed in the CD SKILLS project, including data routinely assessed at individual visits about the diagnostic approach and clinical presentation of the disease. Our study assessed 100 patients (56% girls) with an age range 1.6-18.0 years. We found that 98% of patients were serologically tested prior to a CD diagnosis and 58% of patients were diagnosed using the "no-biopsy" approach. In the analyzed group, 40% belonged to a known risk group, only 22% had annual screening before the CD diagnosis (the longest for 9 years), and 19% showed no symptoms at the time of the CD diagnosis. Our research confirmed the applicability of the "no-biopsy" approach for the diagnosis of CD in children and adolescents in Poland, and also showed changes in the clinical picture of CD. Moreover, we highlight the need to introduce broad CD serological screening in risk groups of the Polish population.

Celiac-the lone horse? An autoimmune condition without signals of microbiota dysbiosis.

Accumulating evidence supports the role of microbiota in autoimmune processes, but research regarding the role of the gut microbiota in celiac disease (CD) is still emerging, and a consistent CD-associated dysbiosis pattern has not yet been defined. Here, we characterized the microbiota of children newly diagnosed with CD, with their unaffected family members as a healthy control group to reduce confounding factors including genetic background, hygiene, dietary habits, and environment, and followed children with CD over 1 year of dietary intervention (exclusion of gluten) to understand if the microbiota is associated with CD and its mediation. We did not find differences in the microbiota of siblings with and without CD, despite a wealth of evidence in the literature supporting CD-specific microbiota. CD is common among first-degree relatives, so this could suggest that unaffected family members in this study may be living in a pre-CD state, currently below clinical detection. Interestingly, despite the effectiveness of diet in CD control, we did not observe diet-mediated microbiota changes, except for short-term increase in Akkermansia muciniphila. This lack of effect could suggest a very strong CD microbial signature even when controlled or could be a technical shortcoming. Expanded future studies with both related and unrelated controls and diet interventions in both the CD and control arms can provide further context to our findings. IMPORTANCE The microbiota is the community of microbes that live in and on us. These microbes are essential to our health and everyday function. Disruption of the community is associated with diseases ranging from metabolic syndrome to autoimmune diseases to mental disorders. In the case of celiac disease (CD), research remains inconclusive regarding implications of the microbiota in etiology. Here, we compared microbiota of children with CD to those of their unaffected family members and found very few differences in microbiota profiles. We next examined how gluten elimination in CD patients affects the microbiota. Surprisingly, despite diet adherence, microbiota shifts were minimal, with only a short-term increase in Akkermansia muciniphila. Previous studies suggest that family members of CD patients may be living in a pre-CD state, which could explain their microbial similarity. A larger study with unrelated controls and increased microbiota monitoring during diet intervention should give our findings more perspective.

Atherosclerotic Risk Factors in Children with Celiac Disease.

RESULTS: We found significantly lower concentrations of total cholesterol, lipoprotein LDL-C, apolipoproteins A1 and B, as well as hCRP in all children with CD. We showed decreased level (<5 ng/mL) of folic acid among 46% of children treated for >5 years. Moreover, we showed significant decrease of folic acid level already after 1 year of a GFD (12 vs. 5.6 ng/mL; p < 0.001). We also found significant negative correlation of z-score body mass index (BMI) with HDL and APOA1 level (r = -0.33; p = 0.015 and r = -0.28; p = 0.038, respectively) and modest positive correlation of z-score BMI with atherogenic factor of total cholesterol-HDL ratio and LDL-HDL ratio (r = 0.40; p = 0.002 and r = 0.36; p = 0.006, respectively). Analysis of physical activity showed an increase in the insulin levels with inactivity (r = 0.36; p = 0.0025). We also found positive correlation of the sleep duration with the adiponectin level (r = 0.41; p = 0.011). CONCLUSIONS: In children with CD treated with a GFD, decreased level of folic acid together with increased BMI, sedentary behavior, and an improper lipid profile may predispose them to atherosclerosis in the long run. This data suggests the need of further studies to determine the need for metabolic cardiovascular risk screening in children with CD.

Transcriptional and Ultrastructural Analyses Suggest Novel Insights into Epithelial Barrier Impairment in Celiac Disease.

Disruption of epithelial junctional complex (EJC), especially tight junctions (TJ), resulting in increased intestinal permeability, is supposed to activate the enhanced immune response to gluten and to induce the development of celiac disease (CD). This study is aimed to present the role of EJC in CD pathogenesis. To analyze differentially expressed genes the next-generation mRNA sequencing data from CD326+ epithelial cells isolated from non-celiac and celiac patients were involved. Ultrastructural studies with morphometry of EJC were done in potential CD, newly recognized active CD, and non-celiac controls. The transcriptional analysis suggested disturbances of epithelium and the most significant gene ontology enriched terms in epithelial cells from CD patients related to the plasma membrane, extracellular exome, extracellular region, and extracellular space. Ultrastructural analyses showed significantly tighter TJ, anomalies in desmosomes, dilatations of intercellular space, and shorter microvilli in potential and active CD compared to controls. Enterocytes of fetal-like type and significantly wider adherence junctions were observed only in active CD. In conclusion, the results do not support the hypothesis that an increased passage of gluten peptides by unsealing TJ precedes CD development. However, increased intestinal permeability due to abnormality of epithelium might play a role in CD onset.

Vitamin K status in cystic fibrosis patients with liver cirrhosis.

The available data on the influence of liver cirrhosis on vitamin K status in CF patients is scarce. Therefore, the aims of the present study were to assess the prevalence of vitamin K deficiency in cirrhotic CF subjects and to determine whether it correlates with liver cirrhosis. The study group comprised of 27 CF patients with and 63 without liver cirrhosis. Vitamin K status was assessed using prothrombin induced by vitamin K absence (PIVKA-II) and the percentage of undercarboxylated osteocalcin (u-OC). PIVKA-II concentrations were higher in cirrhotic than in non-cirrhotic CF patients (median [1st-3rd quartile]: 3.2ng/ml [1.0-10.0] vs. 1.3ng/ml [0.2-2.6], p=0.0029). However, the differences in u-OC percentages between the studied groups did not reach the level of significance (49.4% [7.0-73.8] vs. 8.0% [2.6-59.1], p=0.0501). Based on multiple linear regression analysis the dose of vitamin K and F508del mutation were potentially defined as determinants of vitamin K deficiency. Liver cirrhosis was not documented to be an independent risk factor. In CF patients with liver cirrhosis vitamin K deficiency is not only more frequent, but also more severe. However, not liver cirrhosis, but the presence of a F508del CFTR mutation constitutes an independent risk factor for vitamin K deficiency.

Clinical utility of quantitative multi-antibody Polycheck immunoassays in the diagnosis of coeliac disease.

AIM: To evaluate the clinical utility of multi-antibody strategies in the diagnosis of coeliac disease (CD), the new quantitative Polycheck immunoassays were analysed. METHODS: Polycheck Celiac Panels (PCPs) are immunoenzyme screening assays for the quantitative measurement of coeliac-specific immunoglobulin class G (IgG) or class A (IgA) in serum. Lines of relevant antigens are coated together with five IgG or IgA standard lines used for the standard curve as positive control. PCP IgA consists of human recombinant human tissue transglutaminase (tTG) and deamidated gliadin peptides (DGP) as targets to detect IgA antibodies. PCP IgG consists of tTG, DGP and IF (intrinsic factor) antigens to detect antibodies in IgG class. PCPs were performed on 50 CD patients, including 6 cases with selective IgA deficiency, and 50 non-coeliac controls. CD diagnosis was performed according to the ESPGHAN recommendations: The presence of specific anti-tTG-IgA or anti-DGP-IgG (in the case of IgA deficiency) antibodies, typical histopathological changes in duodenal mucosa described in Marsh-Oberhüber classification as at least grade 2. The diagnosis of the majority of the control subjects was functional gastrointestinal disorders. The PCP results were compared with reference EliA Celikey. RESULTS: The usage of PCPs led to the correct identification of all CD patients. In our study, PCPs showed 100% agreement with the histopathological results. PCP IgA test showed a 98% concordance and correlated positively (R = 0.651, P = 0.0014) with EliA Celikey test. The highest specificity and positive predictive value (both 100%) were observed for the detection of Polycheck anti-tTG-IgA antibodies. The highest sensitivity and negative predictive value (both 100%) were achieved by Polycheck anti-DGP-IgG antibody detection. The best performance (98% sensitivity and negative predictive value, 100% specificity and positive predictive value, diagnostic accuracy - AU ROC 99%) was observed for the strategy of using both PCP IgA and IgG and determining positive outcomes of the test with two or more coeliac-specific antibodies detected. The majority of coeliac patients had multiple antibodies. All four antibodies were detected in 7 (14%) cases, 19 children (38%) were positive for three antibodies and 23 (46%) were positive for two antibodies. CONCLUSION: The present study showed that detection of coeliac-specific antibodies with multi-antibody PCPs is effective and efficacious in the diagnosis of CD.

Serum Concentrations of Insulin, Ghrelin, Adiponectin, Leptin, Leptin Receptor and Lipocalin-2 in Children with Celiac Disease Who Do and Do Not Adhere to a Gluten-Free Diet.

BACKGROUND/AIMS: The roles of the many bioactive peptides in the pathogenesis of celiac disease remain unclear. To evaluate the serum concentrations of insulin, ghrelin, adiponectin, leptin, leptin receptor, and lipocalin-2 in children with celiac disease who do and do not adhere to a gluten-free diet (GFD, intermittent adherence). METHODS: Prepubertal, pubertal, and adolescent celiac children were included in this study (74 girls and 53 boys on a GFD and 80 girls and 40 boys off of a GFD). RESULTS: Insulin levels in prepubertal (9.01±4.43 µIU/mL), pubertal (10.3±3.62 µIU/mL), and adolescent (10.8±4.73 µIU/mL) girls were higher than those in boys (5.88±2.02, 8.81±2.88, and 8.81±2.26 µIU/mL, respectively) and were neither age-dependent nor influenced by a GFD. Prepubertal children off of a GFD exhibited higher ghrelin levels than prepubertal children on a GFD. Adiponectin levels were not age-, sex- nor GFD-dependent. Adherence to a GFD had no effect on the expression of leptin, leptin receptor, and lipocalin-2. CONCLUSIONS: Adherence to a GFD had no influence on the adiponectin, leptin, leptin receptor, and lipocalin-2 concentrations in celiac children, but a GFD decreased highly elevated ghrelin levels in prepubertal children. Further studies are required to determine whether increased insulin concentrations in girls with celiac disease is suggestive of an increased risk for hyperinsulinemia.

Combination Testing Using a Single MSH5 Variant alongside HLA Haplotypes Improves the Sensitivity of Predicting Coeliac Disease Risk in the Polish Population.

Assessment of non-HLA variants alongside standard HLA testing was previously shown to improve the identification of potential coeliac disease (CD) patients. We intended to identify new genetic variants associated with CD in the Polish population that would improve CD risk prediction when used alongside HLA haplotype analysis. DNA samples of 336 CD and 264 unrelated healthy controls were used to create DNA pools for a genome wide association study (GWAS). GWAS findings were validated with individual HLA tag single nucleotide polymorphism (SNP) typing of 473 patients and 714 healthy controls. Association analysis using four HLA-tagging SNPs showed that, as was found in other populations, positive predicting genotypes (HLA-DQ2.5/DQ2.5, HLA-DQ2.5/DQ2.2, and HLA-DQ2.5/DQ8) were found at higher frequencies in CD patients than in healthy control individuals in the Polish population. Both CD-associated SNPs discovered by GWAS were found in the CD susceptibility region, confirming the previously-determined association of the major histocompatibility (MHC) region with CD pathogenesis. The two most significant SNPs from the GWAS were rs9272346 (HLA-dependent; localized within 1 Kb of DQA1) and rs3130484 (HLA-independent; mapped to MSH5). Specificity of CD prediction using the four HLA-tagging SNPs achieved 92.9%, but sensitivity was only 45.5%. However, when a testing combination of the HLA-tagging SNPs and the MSH5 SNP was used, specificity decreased to 80%, and sensitivity increased to 74%. This study confirmed that improvement of CD risk prediction sensitivity could be achieved by including non-HLA SNPs alongside HLA SNPs in genetic testing.

Evaluation of factors related to bone disease in Polish children and adolescents with cystic fibrosis.

PURPOSE: The aim of the study was to evaluate factors related to bone formation and resorption in Polish children and adolescents with cystic fibrosis and to examine the effect of nutritional status, biochemical parameters and clinical status on bone mineral density. MATERIALS AND METHODS: The study group consisted of 100 children and adolescents with cystic fibrosis with a mean age 13.4 years old. Anthropometric measurements, included body height, body mass and body mass index (BMI); bone mineral densitometry and biochemical testing were performed. Bone mineral density was measured using a dual-energy X-ray absorption densitometer. Biochemical tests included serum calcium, phosphorus, parathyroid hormone and vitamin D concentrations, as well as 24-h urine calcium and phosphorus excretion. Pulmonary function was evaluated using FEV1%, and clinical status was estimated using the Shwachman-Kulczycki score. RESULTS: Standardized body height, body mass and BMI were significantly lower than in the reference population. Mean serum vitamin D concentration was decreased. Pulmonary disease was generally mild, with a mean FEV1% of 81%. Multivariate linear regression revealed that the only factors that had a significant effect on bone marrow density were BMI and FEV1%. There were no significant correlations between bone mineral density and the results of any of the biochemical tests performed. CONCLUSIONS: Nutritional status and bone mineral density were significantly decreased in children and adolescents with cystic fibrosis. In spite of abnormalities in biochemical testing, the factors that were found to have the strongest effect on bone mineral density were standardized BMI and clinical status.

Dietary pattern and its relationship between bone mineral density in girls and boys with cystic fibrosis - preliminary report.

BACKGROUND: Nutrition influence on cystic fibrosis (CF) patients survival is well documented and dietary therapy is one of basic elements of their treatment. Prolonged survival of CF patients might yet emerge comorbidities, which include bone mineral disease. THE AIM: The assessment of the dietary pattern and its relationship between bone density in boys and girls with cystic fibrosis. MATERIAL AND METHODS: 89 patients aged 10-18 years from 3 Polish CF Centres were included into the study. To obtain a knowledge about quality of diet, a 3-day food record was assessed and percent of recommended for CF patients intake of energy, protein, carbohydrates, fat, vitamin D3, calcium, phosphorus was counted. Bone mineral density (BMD) in lumbar spine (L1-L4) was measured and expressed as a Z-score. To assess nutritional status, anthropometric measurements was evaluated (body weight, height and BMI). Descriptive methods, Mann-Whitney test, T-Student test, Spearman correlation and one-way ANOVA were used for statistical analyses. RESULTS: The patients with cystic fibrosis did not meet specific for CF nutritional guidelines. A deficiency in recommended intake was observed in energy (88%), protein (82%), calcium (78%) and vitamin D3 (71%). The intake of phosphorus was higher than recommended (142%). A nutritional status was significantly reduced, as compared with the reference group (p 0.001). Boys characterized significantly lower body weight (p=0.019) and height (p=0.036) than girls as well as worse caloric (p=0.023) and carbohydrates intake (p=0.005). However, girls had reduced vitamin D3 content in their diet (p 0.001). The bone mineral density in the whole group was reduced and Z-score amounted to -0.95 ± 1.17. Tendency to decreasing of BMD with age was observed. BMI showed important correlation with bone mineral density both in girls (p 0.001) and in boys (p=0.020). CONCLUSION: CF patients do not follow specific for them dietary recommendations and essential differences were observed between girls and boys. Nutritional status (BMI) showed correlation with bone mineral density in CF patients. Therefore intensive nutritional therapy according to recommendations is needed. .

Lactose malabsorption is a risk factor for decreased bone mineral density in pancreatic insufficient cystic fibrosis patients.

As decreased bone mineral density (BMD) is a common problem in cystic fibrosis (CF) and milk products may have pivotal dietary role affecting BMD, we aimed to assess the potential influence of adult-type hypolactasia (ATH) and lactose malabsorption (LM) on BMD in adolescent and young adult patients. In 95 CF pancreatic-insufficient patients aged 10-25 years (without liver cirrhosis, steatosis and cholestasis, diabetes mellitus, systemic glucocorticoid therapy), lumbar BMD, the nutritional status, pulmonary function, vitamin D3 concentration, calcium intake and single-nucleotide polymorphism upstream of the lactase gene were assessed. In subjects with the -13910 C/C genotype predisposing to ATH, the presence of LM was determined with the use of a hydrogen-methane breath test (BT). BMD and calcium intake were significantly lower in patients with the C/C genotype (P<0.028 and P<0.043, respectively). The abnormal BMD was stated more frequently in patients with the C/C genotype (P<0.042) and with LM (P<0.007). BMD, daily calcium intake and serum vitamin D concentration were significantly lower in LM subjects than in the other patients (P<0.037, P<0.000004 and P<0.0038, respectively). In logistic regression analysis, the relationship between examined parameters and BMD, was found to be statistically significant (P<0.001). However, only standardized body weight and LM were documented to influence BMD (P<0.025 and P<0.044, respectively). In conclusion, LM seems to be an independent risk factor for decreased BMD in CF patients.

Mild CFTR mutations and genetic predisposition to lactase persistence in cystic fibrosis.

Taking into account the reported incidence of hypolactasia in cystic fibrosis (CF) and the possible impact of milk products on nutritional status we aimed to assess the genetic predisposition to adult-type hypolactasia (ATH) and its incidence in CF. Single nucleotide polymorphism upstream of the lactase gene (LCT) was assessed in 289 CF patients. In subject with -13910C/C genotype (C/C) predisposing to ATH, hydrogen-methane breath test (BT) with lactose loading was conducted and clinical symptoms typical for lactose malabsorption were assessed. The percentage of CF patients with C/C was similar to that observed in healthy subjects (HS) (31.5 vs 32.5% ). Eleven out of 52 (24.5%) CF C/C patients had abnormal BT results. The recalculated frequency of lactose malabsorption was similar for the entire CF and HS populations (6.9 vs 7.2%). Similarly as in the control group, few CF patients have identified and linked to lactose consumption clinical symptoms. The frequency of LCT polymorphic variants in CF patients having and not having severe mutations of CFTR gene showed significant differences. The C allele was more frequent in homozygotes of the severe mutations than in patients carrying at least one mild/unknown mutation (P<0.0028) and in patients with at least one mild mutation (P < 0.0377). In conclusion, CF patients carrying mild CFTR mutations seem to have lower genetic predisposition to ATH. Lactose malabsorption due to ATH in CF is not more frequent than in the general population. Symptomatic assessment of lactose malabsorption in CF is not reliable.

Cystic fibrosis is a risk factor for celiac disease.

BACKGROUND: The coexistence of cystic fibrosis (CF) and celiac disease (CD) has been reported. To our knowledge there is no study directly comparing the incidence of CD in CF patients to that in the general population at the same time. There is no published data on genetic predisposition to CD in CF patients either. Therefore, in the present study we aimed to assess the genetic predisposition to CD and its incidence in CF patients comparing it to data from the general population. PATIENTS AND METHODS: Two hundred eighty-two CF patients were enrolled in the study. In 230 CF patients the genetic predisposition to CD (the presence of HLA-DQ2/ DQ8) was assessed. In all CF patients, serological screening for CD was conducted. In patients with positive antiendomysial antibodies (EMA) gastroduenoscopy was offered. Intestinal histology was classified according to modified Marsh criteria. The results of serological CD screening in 3235 Polish schoolchildren and HLA-DQ typing in 200 healthy subjects (HS) were used for comparison. RESULTS: Positive EMA was found in 2.84% of the studied CF patients. The incidence of proven CD was 2.13%. The incidence of CD as well as positive serological screening were significantly more frequent in the CF group than in the general population. The frequency of CD-related HLA-DQ alleles in CF and HS did not differ. CONCLUSIONS: Genetic predisposition to celiac disease in cystic fibrosis patients is similar to that of the general population. However, our results suggest that cystic fibrosis is a risk factor for celiac disease development.

Analysis of CFTR, SPINK1, PRSS1 and AAT mutations in children with acute or chronic pancreatitis.

OBJECTIVES: Defects of PRSS1, SPINK1, CFTR and AAT are considered causative or predisposing to pancreatitis. The aim of this study was to evaluate the impact of these defects into molecular pathology of chronic pancreatitis (CP) and acute recurrent pancreatitis (ARP). METHODS: Ninety-two children with CP or ARP, 55 family members and 50 controls were investigated. The subjects were screened for PRSS1 mutations: R122H, R122C, A16V, N29I; SPINK1 N34S variant; panel of 14 CFTR defects: INNOLiPA CFTR12, CFTRdele2,3 and IVS8-T variant or panel of 3 CFTR defects-F508del, CFTRdele2,3 and IVS8-T; AAT mutations: E264V, E342K. RESULTS: We identified 1 mutated allele in at least 1 of 4 genes in 31 of 92 patients and 12 of 50 controls (P = 0.157). Mutations in SPINK1 and PRSS1 were most frequent. PRSS1 mutations were identified mainly in CP patients (9.6% of CP vs 2.5% of ARP alleles, P = 0.094), whereas N34S SPINK1 mutation was present with comparable frequency in CP and ARP patients (7.7% vs 10.0%, P = 0.768). The frequency of mutations in CFTR alleles was similar to controls (4.9% vs 5%, P = 0.587). Overall frequency of AAT mutations was lower than in the controls. Family studies showed that defects in the examined genes did not always segregate with disease. CONCLUSIONS: PRSS1 defects seem to be causative for pancreatitis, whereas defects in SPINK1 are suggested to be associated with the disease. No association between CFTR mutations and pancreatitis was observed. The importance of AAT variants remains speculative.