IgA-Type Enterovirus Antibodies Are Increased among Adults and Children with Recently Diagnosed Type 1 Diabetes.

Enteroviruses (EV) are among the leading environmental triggers of childhood-onset type 1 diabetes (T1D). Our aim was to determine the prevalence of antibodies against EV and their association with T1D in different age groups (n = 62), including young adults, and to compare these data with results from HLA-matched control participants (n = 62). IgA, IgG, and IgM antibodies against EV were detected. IgA EV antibodies were present in 46.8% of participants with T1D (median level 10.9 EIU) and in 11.3% of controls (median level 3.4 EIU). IgA EV positivity and higher level of IgA EV antibodies were both significant risk factors for T1D (odds ratio (OR) 8.33; 95% confidence interval (CI) 2.52-27.6; p = 0.0005 and OR 1.04; 95% CI 1.01-1.06; p = 0.0105, respectively). Importantly, the prevalence of IgA EV antibodies in the subgroups of both children and young adults was also significantly different between participants with T1D and their matched controls (p = 0.0089 and p = 0.0055, respectively). Such differences were not seen for IgG and IgM EV antibodies. However, IgG EV antibodies were associated with 65 kDa glutamic acid decarboxylase antibodies, but not with zinc transporter 8 and protein tyrosine phosphatase IA2 antibodies. The genotype frequency of PTPN22 (rs2476601) and IFIH1 (rs1990760) was not associated with EV positivity. This study showed that EV infections may be an important disease-promoting factor of T1D not only in childhood-onset but also in adult-onset T1D. However, to further confirm this association, direct virological studies are needed in the latter T1D group.

FABP4 and I-FABP Levels in Pregnant Women Are Associated with Body Mass Index but Not Gestational Diabetes.

Objectives: Gestational diabetes mellitus (GDM) is glucose intolerance detected initially during pregnancy. GDM poses an increased risk for the development of diabetes later in life. Fatty acid-binding protein 4 (FABP4) is a regulator of lipid metabolism and is associated with obesity, insulin resistance, and type 2 diabetes. Increased level of intestinal fatty acid-binding protein (I-FABP) may indicate impaired intestinal permeability, which may be an important contributor to the pathogenesis of type 1 diabetes and GDM. We aimed to compare FABP4 and I-FABP levels in pregnant women with GDM and in healthy pregnant controls, taking into consideration their prepregnancy body mass index (BMI), past exposures to enteroviruses (EV), and adipokine and cytokine levels, which have been shown to decrease insulin sensitivity. Material and Methods. Forty patients with GDM (median age 30.5) and 40 pregnant healthy controls (median age 31.1) were divided on the basis of their prepregnancy BMI into two groups: normal weight (BMI < 25, n = 20) and overweight (BMI ≥ 25, n = 20). FABP4 and I-FABP were measured from serum samples using commercial ELISA kits. Results: FABP4 and I-FABP levels did not differ between women with GDM and healthy pregnant controls (p > 0.05 for both comparisons). However, both levels were associated with BMI (p < 0.001 for both comparisons). Median I-FABP level was the highest in healthy controls with lower BMI (<25) (p = 0.0009). FABP4 levels correlated with BMI and C-peptide values in both groups (p < 0.001). Anti-EV antibody levels did not correlate with FABP4 or I-FABP levels. FABP4 and adiponectin levels were negatively correlated in controls (r = -0.61, p = 0.0009), while I-FABP correlated positively with adiponectin (r = 0.58, p = 0.04) and resistin (r = 0.67, p = 0.04) levels in the GDM group. Conclusion: FABP4 and I-FABP levels were not dependent on the diagnosis of GDM, but rather on BMI. The correlation of I-FABP with adiponectin and resistin levels in women with GDM may suggests the importance of lipid metabolism in GDM-associated changes in intestinal permeability.

Celiac Disease in Children, Particularly with Accompanying Type 1 Diabetes, Is Characterized by Substantial Changes in the Blood Cytokine Balance, Which May Reflect Inflammatory Processes in the Small Intestinal Mucosa.

Cytokines play a pivotal role in the maintenance of intestinal homeostasis inducing pro- or anti-inflammatory response and mucosal barrier function in celiac disease (CD) and type 1 diabetes (T1D). We aimed to compare the levels of pro- and anti-inflammatory cytokines in CD patients without and with coexisting T1D, as well as to evaluate its association with the presence of enteroviruses (EV), regulatory T cells (Tregs), and dendritic cells (DCs) in small bowel mucosa. Altogether, 72 patients (median age 10.1 years) who had undergone small bowel biopsy were studied. The study group consisted of 24 patients with CD (median age 6.5 years), 9 patients with CD and concomitant T1D (median age 7.0 years), two patients with T1D (median age 8.5 years), and 37 patients (median age 14.0 years) with functional gastrointestinal disorders (FGD) and a normal small bowel mucosa as controls. The levels of 33 cytokines in serum were measured by multiple analysis using the Milliplex® MAP Magnetic Bead assay. The densities of FOXP3+ Tregs, CD11c+ DC, indoleamine 2,3-dioxygenase+ (IDO+) DC, langerin+ (CD207+) DCs, and EV were evaluated by immunohistochemistry as described in our previous studies. Circulating anti-EV IgA and IgG were evaluated using ELISA. The most important finding of the study is the significant increase of the serum levels of IL-5, IL-8, IL-13, IL-15, IL-17F, IL-22, IL-27, IP-10, MIP-1ß, sIL-2Rα, sTNFRII, and TNFα in CD patients compared to controls and its correlation with the degree of small bowel mucosa damage graded according to the Marsh classification. The leptin level was higher in females in all study groups. The levels of IL-2, IL-6, IL-12 (P70), IL-15, IP-10, and IFNγ correlated significantly with the density of FOXP3+ Tregs in lamina propria of the small bowel mucosa, which supports the evidence about the signaling role of these cytokines in the peripheral maintenance of FOXP3+ Tregs. At the same time, a significant negative correlation occurred between the level of IL-4 and density of FOXP3+ Tregs in controls. Another important finding of our study was the correlation of IL-17F, IP-10, sTNFRII, MCP-1, and GM-CSF with the density of EV-positive cells in the lamina propria of the small bowel mucosa. Correlation of MIP-1 (CCL-4) with CD103+ DC and langerin+ DC densities may point to their significance in the recruitment of immune cells into the lamina propria and in driving the inflammatory response in CD patients. Our results suggest the predominance of Th1 and Th17 immune responses over EV VP1 protein in CD and T1D patients. The significant elevation of Th2 cytokines, like IL-5 and IL-13, but not IL-4, in CD patients and its correlation with the degree of small bowel mucosa damage could reflect the role of these cytokines in gut defense and inflammation.

Circulating Zonulin Correlates with Density of Enteroviruses and Tolerogenic Dendritic Cells in the Small Bowel Mucosa of Celiac Disease Patients.

BACKGROUND: Impaired intestinal integrity, including increased permeability of the small bowel mucosa, has been shown in patients with celiac disease (CD) as well as with type 1 diabetes (T1D). Zonulin (ZO, pre-haptoglobin), a tight junction regulator, plays a particular role in the regulation of intestinal barrier function and in the pathogenesis of the above-mentioned diseases. AIM: To investigate whether enteroviruses (EVs) and immunoregulatory cells are associated with intestinal permeability in patients with CD alone and with coexistent T1D. MATERIALS AND METHODS: Altogether 80 patients (mean age 10.68 ± 6.69 years) who had undergone small bowel biopsy were studied. Forty patients with functional dyspepsia and normal small bowel mucosa formed the control group. The circulating ZO level in sera was evaluated using ELISA. The densities of EV, FOXP3+ regulatory T cells (Tregs), indoleamine 2,3-dioxygenase (IDO+) dendritic cells (DCs) and glutamic acid dexarboxylase (GAD)65+ cells in small bowel mucosa were investigated by immunohistochemistry. The expression analysis of FOXP3, tight junction protein 1 (TJP1), gap junction (GJA1), IDO and CD103 genes was evaluated by real-time PCR. RESULTS: The ZO level was higher in CD patients compared to subjects with a normal small bowel mucosa, particularly in those with Marsh IIIc atrophy (p = 0.01), and correlated with the density of EV (r = 0.63; p = 0.0003) and IDO+ DCs (r = 0.58; p = 0.01) in the small bowel mucosa. The density of GAD65+ epithelial cells was correlated with the density of EV (r = 0.59; p = 0.03) and IDO+ DCs (r = 0.78; p = 0.004) in CD patients. The relative expression of FOXP3 mRNA in the small bowel mucosa tissue was significantly higher in patients with CD, compared to subjects with a normal mucosa, and correlated with the density of EV (r = 0.62; p = 0.017) as well as with the relative expression of IDO mRNA (r = 0.54; p = 0.019). CONCLUSIONS: The CD is associated with elevation of the circulating ZO level, the value of which correlates with the density of EV in CD patients with severe atrophic changes in the small bowel mucosa, particularly in cases of concomitant T1D. The CD is also characterized by the close relationship of the density of GAD65+ epithelial cells with the EV, ZO level and IDO+ DCs.

Antibodies to Lactobacilli and Bifidobacteria in young children with different propensity to develop islet autoimmunity.

The intestinal microbiota is essential to the maturation and homeostasis of the immune system. Immunoblot assays were used to establish the prevalence of serum IgG, IgM, and IgA antibodies specific for Bifidobacterium adolescentis, Bifidobacterium longum, and Lactobacillus rhamnosus GG proteins in young children presenting with or without type 1 diabetes (T1D). We demonstrated that children between the ages of 6 and 12 months had a substantial increase in the frequency of IgG antibodies specific for L. rhamnosus GG proteins. We measured IgG, IgM, and IgA class antibody reactivity against B. adolescentis DSM 20083, B. adolescentis DSM 20086, and B. longum DSM 20088 proteins demonstrating significantly higher IgA responses against B. adolescentis DSM 20083 strain proteins in children who developed islet autoimmunity and T1D later in life. B. adolescentis strains showed more IgM type antibodies in children who developed T1D later in life, but the difference was not statistically significant. B. longum proteins were recognized by IgG and IgA antibodies to a higher extent compared to other bacteria studied. These results confirm that differences in immune reactivity against some commensal strains in young children may represent a different risk factor for developing T1D.

Celiac disease in patients with type 1 diabetes: a condition with distinct changes in intestinal immunity?

Two common chronic childhood diseases-celiac disease (CD) and type 1 diabetes (T1D)-result from complex pathological mechanisms where genetic susceptibility, environmental exposure, alterations in intestinal permeability and immune responses play central roles. In this study, we investigated whether these characteristics were universal for CD independently of T1D association. For this purpose, we studied 36 children with normal small-bowel mucosa and 26 children with active CD, including 12 patients with T1D. In samples from the small-bowel mucosa, we detected the lowest expression of tight junction protein 1 (TJP1) mRNA in CD patients with T1D, indicating an increase in intestinal permeability. Furthermore, these samples displayed the highest expression of forkhead box P3 (FoxP3) mRNA, a marker for regulatory T cells, as compared with other patient groups. At the same time, serum levels of IgA antibodies specific for the CD-related antigens deamidated gliadin and tissue transglutaminase (tTG) were the highest in CD patients with T1D. In contrast, no significant differences were found in IgA or IgG antibodies specific for bovine beta-lactoglobulin or Bifidobacterium adolescentis DSM 20083-derived proteins. There were also no differences in the transamidating activity of serum autoantibodies between patients and control individuals. Our results show that patients with T1D and newly detected CD exhibit severely altered intestinal permeability, strong local immune activation and increased immunoregulatory mechanisms in the small bowel. Further study is required to determine whether these extreme changes in this CD subgroup are due to some specific environmental factors (virus infections), unknown genetic effects or autoimmune reactions to antigenic targets in intracellular tight junctions.

Thyroid autoimmunity and treatment response to escitalopram in major depression.

BACKGROUND: There is evidence that immune alterations play an important part in the pathogenesis of major depression. Thyroid autoimmunity has been found in association with major depression in several studies. AIM: 1) to examine whether the prevalence of anti-thyroid peroxidase autoantibodies (anti-TPO) in depressive patients differs from that in healthy controls; 2) to investigate the possible relationship between thyroid autoimmunity, total T3, free T3, free T4, thyroid-stimulating hormone (TSH), clinical status and treatment outcome in depression. METHOD: The study group consisted of 129 outpatients (69.8% female; mean age 31.7+/-12.0 years) with major depressive disorder with a Montgomery-Azsberg Depression Rating Scale total score of 22 or higher and 72 healthy controls (62.5% female; mean age 31.7+/-13.1 years). The patients were treated with escitalopram 10-20 mg/day for 12 weeks using open-label placebo non-controlled design. Anti-TPO, total T3, free T3, free T4 and TSH were measured before the treatment. RESULTS: The anti-TPO was found in eight (8.9%) depressive and two (4.8%) healthy females without statistical difference between these groups. Since anti-TPO was not seen in males, all further statistical analyses were carried out in females. At the end of week 12 of the treatment, 60 female patients (66.7%) were defined as responders and 30 depressive females (33.3%) showed insufficient response to treatment. Although there were no significant differences in the measurements between responders and non-responders, the last group showed a trend for a higher prevalence of anti-TPO compared with responders. CONCLUSION: Thyroid autoimmunity might be a factor predicting treatment response to antidepressants in depressive patients.

Antigenic proteins of Lactobacillus acidophilus that are recognised by serum IgG antibodies in children with type 1 diabetes and coeliac disease.

Immune responses to lactobacilli have been so far insufficiently investigated in patients with autoimmune diseases. We used whole-cell lysate of an indigenous Lactobacillus acidophilus strain isolated from an Estonian child to study serum IgG antibodies in children groups with type 1 diabetes [insulin dependent diabetes mellitus (IDDM)] (n = 21, age 4-18 yr) and with acute coeliac disease (CD) (n = 20, age 0.6-15 yr) and to compare the results with the controls (n = 24, age 2-17 yr). We found that our developed 1-D immunoblot assay readily enables to reveal antibodies against 28 L. acidophilus antigenic proteins in patients' and controls' sera. As verified by immunoproteomics analysis with 2-D and LC ESI-MS/MS the antigens of L. acidophilus were mainly common cytoplasmic proteins GroEL (HSP60), enolase, transcription factor EF-Ts and EF-Tu. However, in addition we identified formyl-CoA transferase being target for antibodies in every tested IDDM patients' serum. We have characterized for the first time the antigenic profile of L. acidophilus whole-cell lysate using sera from children with IDDM, CD, and controls. The different prevalence of reactions against tested antigens in patients and controls sera may indicate significant differences in immune system and commensal bacteria cross-talk in these groups.

Antibodies to neurofilaments.

Immune responses to neuronal proteins are a frequent occurrence in neurodegenerative diseases. This study determines the occurrence of autoantibodies to the three neurofilament subunits in phosphorylated and dephosphorylated forms and relates these measures to age, human leukocyte antigen (HLA), and severity of disease in Down syndrome (DS). IgG-type antibodies to three neurofilament (NF) subunits, NF-L, NF-M, and NF-H, in both phosphorylated and dephosphorylated forms were tested by immunoblot in 128 patients with DS and compared to antibody levels in 94 normal controls. DS was revealed by karyotype analysis. Antibodies to dephospho-NF-M, NF-M, and NF-L were more common in DS samples than in controls. Total pools of DS and control samples had similar frequency of antibodies to NF-H, but there was a higher prevalence of antibodies against NF-H in DS patients with moderate or mild disability (43.0%) compared with those having serious disability (14.3%). No NF-H antibody was seen among young children (age 14 years or younger) of the latter group. The HLA-DR15 allele was negatively correlated with antibodies against NF-H. The high prevalence of antibodies to neurofilament proteins and the differences between DS and control samples may reflect the cross-talk between neural and immune systems that could have an important role in defending neural structures from neurodegenerative processes. In children with DS the presence of antibodies to NF-H might reflect a more favorable prognosis.

Putative predictors of antibodies against follicle-stimulating hormone in female infertility: a study based on in vitro fertilization patients.

PROBLEM: We have previously demonstrated the presence of naturally occurring antibodies against follicle-stimulating hormone (FSH) in patients with endometriosis and polycystic ovary syndrome (PCOS). Here, we investigated the parameters associated with anti-FSH antibodies in in vitro fertilization (IVF) patients. METHODS OF STUDY: The following parameters were studied in 135 patients: peripheral FSH levels, FSH beta-subunit gene (FSHB) haplotypes, history of previous IVF, and susceptibility to autoimmune reactions in general [seven common autoantibodies (against nuclear antigens on human and rodent substrates, smooth muscle, gastric parietal cells, beta2-glycoprotein I, cardiolipin, and thyroid peroxidase) and HLA-DQB1 alleles]. RESULTS: Although the anti-FSH levels were higher in patients when compared with controls, those higher levels were not associated with FSHB haplotypes. The anti-FSH IgM associated with (i) the levels of FSH in women with male and tubal factor infertility; (ii) the history of IVF in patients with PCOS, endometriosis, and unexplained infertility; and (iii) the production of common autoantibodies among all IVF patients. The anti-FSH IgA associated with HLA-DQB1*03. The anti-FSH IgG correlated with the values of anti-FSH IgA and IgM. CONCLUSION: Anti-FSH may be naturally occurring antibodies associated with peripheral FSH concentrations, but increased in infertile women with dysregulation of immune reactions and repeatedly performed IVF.

Controlled ovarian hyperstimulation changes the prevalence of serum autoantibodies in in vitro fertilization patients.

PROBLEM: Autoimmune mechanisms are involved in etiology of female infertility, the medical problem frequently treated by in vitro fertilization (IVF). Controlled ovarian hyperstimulation (COH) with supraphysiological levels of sex hormones is achieved by IVF. METHODS OF STUDY: Anti-human-ovary and eight common autoantibodies [nuclear (ANA-H, ANA-R on human HEp-2 cell line and rodent antigen, respectively), smooth muscle (SMA), parietal cell, thyroid microsomal, mitochondrial, beta2-glycoprotein-I, cardiolipin antibodies] found in IVF patients (n = 129) were analyzed with regard to the number of previous IVF procedures and the age of the patient. The changes in autoimmune reactions caused by the COH were determined. RESULTS: Endometriosis and polycystic ovary syndrome were associated with a higher number of common serum autoantibodies compared with the tubal factor infertility (Proportion test, P < 0.05). ANA-R was associated with unexplained infertility [adjusted odds ratio (aOR) 8.79, P = 0.038]. SMA correlated with endometriosis (aOR 37.29, P = 0.008), male factor infertility (aOR 20.45, P = 0.018) and with the previous IVF procedures (aOR 2.87, P = 0.013). There was an overall decrease in the number of detectible autoantibodies after COH (Proportion test, P < 0.05). CONCLUSION: COH may have a suppressive effect on the humoral immunity by the time of embryo transfer but more conclusive studies are needed.