Circulating Dendritic Cells, Farm Exposure and Asthma at Early Age.

Farm environment has been shown to protect from childhood asthma. Underlying immunological mechanisms are not clear yet, including the role of dendritic cells (DCs). The aim was to explore whether asthma and farm exposures are associated with the proportions and functional properties of DCs from 4.5-year-old children in a subgroup of the Finnish PASTURE birth cohort study. Myeloid DCs (mDCs), plasmacytoid DCs (pDCs) and CD86 expression on mDCs ex vivo (n = 100) identified from peripheral blood mononuclear cells (PBMCs) were analysed using flow cytometry. MDCs and production of interleukin (IL)-6 and tumour necrosis factor alpha (TNF-α) by mDCs were analysed after 5 h in vitro stimulation with lipopolysaccharide (LPS) (n = 88). Prenatal and current farm exposures (farming, stables, hay barn and farm milk) were assessed from questionnaires. Asthma at age 6 years was defined as a doctor's diagnosis and symptoms; atopic sensitization was defined by antigen-specific IgE measurements. Asthma was positively associated with CD86 expression on mDCs ex vivo [adjusted odds ratio (aOR) 4.83, 95% confidence interval (CI) 1.51-15.4] and inversely with IL-6 production in mDCs after in vitro stimulation with LPS (aOR 0.19, 95% CI 0.04-0.82). In vitro stimulation with LPS resulted in lower percentage of mDCs in the farm PBMC cultures as compared to non-farm PBMC cultures. Our results suggest an association between childhood asthma and functional properties of DCs. Farm exposure may have immunomodulatory effects by decreasing mDC proportions.

Moisture damage in home associates with systemic inflammation in children.

This study investigated the association between confirmed moisture damage in homes and systemic subclinical inflammation in children. Home inspections were performed in homes of 291 children at the age of 6 years. Subclinical inflammation at the age of 6 years was assessed by measuring the circulating levels of C-reactive protein (CRP) and leukocytes in peripheral blood and fractional exhaled nitric oxide (FeNO). Proinflammatory cytokines interleukin (IL)-1ß and IL-6 and tumor necrosis factor (TNF)-α were measured in unstimulated, and in phorbol 12-myristate 13-acetate and ionomycin (PI), lipopolysaccharide (LPS), or peptidoglycan (PPG)-stimulated whole blood. Major moisture damage in the child's main living areas (living room, kitchen, or child's bedroom) and moisture damage with mold in the bathroom were associated with increased levels of CRP and stimulated production of several proinflammatory cytokines. There were no significant associations between moisture damage/visible mold and leukocyte or FeNO values. The results suggest that moisture damage or mold in home may be associated with increased systemic subclinical inflammation and proinflammatory cytokine responsiveness.

High level of fecal calprotectin at age 2 months as a marker of intestinal inflammation predicts atopic dermatitis and asthma by age 6.

BACKGROUND: Gut microbiota and intestinal inflammation regulate the development of immune-mediated diseases, such as allergies. Fecal calprotectin is a biomarker of intestinal inflammation. OBJECTIVE: We evaluated the association of early-age fecal calprotectin levels to the later development of allergic diseases in children from farming and non-farming environments and further studied the effect of gut microbiota on the fecal calprotectin levels. METHODS: Fecal calprotectin was measured from 758 infants participating in the PASTURE study at the age of 2 months using the ELISA method. Serum-specific IgE levels were measured at 6 years of age. Data of environmental factors, doctor-diagnosed atopic dermatitis (AD) and asthma were collected by questionnaire. Multivariate logistic regression models were used for analysis. The composition of fecal microbiota was analysed in a subgroup of 120 infants with 16S rRNA pyrosequencing. The effect of Escherichia coli lipopolysaccharide (LPS) on in vitro monocyte IL-10 secretion was studied by flow cytometry. RESULTS: The infants with high fecal calprotectin levels at 2 months, that is above the 90th percentile, had an increased risk of developing AD and asthma/asthmatic bronchitis by the age of 6 years (aOR 2.02 (1.06-3.85) and 2.41 (1.25-4.64), respectively). High fecal calprotectin levels correlated negatively with fecal Escherichia. LPS from E. coli stimulated production of IL-10 in monocytes. CONCLUSION AND CLINICAL RELEVANCE: High degree intestinal inflammation at 2 months of age, detected as high fecal calprotectin, predicted asthma and AD by the age of 6 years and was linked to low abundance of fecal Escherichia. Impaired IL-10 activation due to the lack of colonization with E. coli could explain the intestinal inflammation associated high fecal calprotectin and later risk of asthma and AD. Our results have implications for the design of probiotic treatments and suggest that early intestinal colonization has long-term health effects.

Monocyte-derived dendritic cells from Crohn's disease patients exhibit decreased ability to activate T helper type 17 responses in memory cells.

An increased activation of interleukin (IL)-17A-producing immune cells is a well-established feature of Crohn's disease (CD). Mechanisms that contribute to this aberrant immune activation are, however, less clear. Given that an enhanced induction of innate-immunity associated cytokines IL-6 and IL-23, which promote IL-17 immunity, is also clearly implicated in CD, we hypothesized that monocyte-derived dendritic cells (moDCs) of CD patient origin would mount exaggerated IL-17A responses in T cells. However, we found a significantly attenuated up-regulation of the IL-17A response in allogeneic T helper memory cells in the presence of culture media from lipopolysaccharide (LPS)-stimulated moDCs of CD patients when compared with moDCs of control subjects (median fold-increase in IL-17A mRNA expression 1·09 versus 1·44, P = 0·038). This was accompanied by a lower expression of IL-1ß and IL-6 transcripts in the LPS-treated moDCs (median 9·55 versus 13·9 relative units, P = 0·042, and 2·66 versus 9·06 relative units, P = 0·049, respectively). In addition, the up-regulation of autophagy-related LC3 transcripts was decreased in moDCs of CD patients (median fold-increase in mRNA expression 1·22 versus 1·52, P = 0·029). Our findings reveal similar immunological aberrancies in CD in the general population as reported in CD patients with mutated intracellular bacterial sensor NOD2, namely attenuated activation of innate cytokines and impaired autophagy, combined with a reduced capacity to up-regulate the T helper type 17 (Th17) response. The results presented here emphasize a defective anti-microbial response in the pathogenesis of CD. The increased mucosal Th1 and Th17 responses, which may contribute to the pathogenesis, could be the consequences of primary defects in the innate immunity.

Soluble immunoglobulin A in breast milk is inversely associated with atopic dermatitis at early age: the PASTURE cohort study.

BACKGROUND: The role of breastfeeding for the development of atopic diseases in childhood is contradictory. This might be due to differences in the composition of breast milk and levels of antimicrobial and anti-inflammatory components. OBJECTIVE: The objective of this study was to examine whether levels of total immunoglobulin A (IgA) or transforming growth factor-ß1 (TGF-ß1) in breast milk were associated with the risk of developing atopic dermatitis (AD), atopic sensitization or asthma at early age taking breastfeeding duration into account. METHODS: The birth cohort study PASTURE conducted in Finland, France, Germany and Switzerland provided 610 breast milk samples collected 2 months after delivery in which soluble IgA (sIgA) and TGF-ß1 levels were measured by ELISA. Duration of breastfeeding was assessed using weekly food frequency diaries from month 3 to month 12. Data on environmental factors, AD and asthma were collected by questionnaires from pregnancy up to age 6. Atopic status was defined by specific IgE levels in blood collected at the ages of 4 and 6 years. Multivariate logistic regression models were used for statistical analysis. RESULTS: Soluble IgA and TGF-ß1 levels in breast milk differed between countries, and sIgA levels were associated with environmental factors related to microbial load, for example, contact to farm animals or cats during pregnancy, but not with raw milk consumption. sIgA levels were inversely associated with AD up to the of age 2 years (P-value for adjusted linear trend: 0.005), independent of breastfeeding duration. The dose of sIgA ingested in the first year of life was associated with reduced risk of AD up to the age of 2 (aOR, 95% CI: 0.74; 0.55-0.99) and 4 years (0.73; 0.55-0.96). No clear associations between sIgA and atopy or asthma up to age 6 were observed. TGF-ß1 showed no consistent association with any investigated health outcome. CONCLUSION AND CLINICAL RELEVANCE: IgA in breast milk might protect against the development of AD.

No association between vitamin D and ß-cell autoimmunity in Finnish and Estonian children.

BACKGROUND: Vitamin D has immunomodulatory properties, such as regulation of FOXP3 expression and regulatory T-cell activity. Our aim was to investigate whether plasma 25-hydroxyvitamin D [25(OH)D] concentrations associate with the development of ß-cell autoimmunity and the transcriptional activity of FOXP3 or vitamin D3 convertase gene (CYP27B1) in CD4+ memory T cells. METHODS: We studied 83 Finnish and 32 Estonian children participating in the DIABIMMUNE and DIPP studies. Twenty-nine Finnish and six Estonian children tested positive for at least one diabetes-associated autoantibody. The plasma concentrations of 25(OH)D and 1,25(OH)2D were analysed with an enzyme immunoassay. Gene expression of FOXP3 and CYP27B1 in the isolated CD4+ memory T cells was studied with reverse transcription quantitative polymerase chain reaction. RESULTS: Vitamin D status did not differ between subjects positive and negative for ß-cell autoantibodies. Finnish children had higher vitamin D status than Estonian children (p < 0.001). FOXP3 expression was higher in Estonian CD4+ memory T-cell samples than in Finnish samples (p < 0.01) even when including in both groups only children with serum 25(OH)D concentrations in the range of 50-80 nmol/L (p < 0.001). CONCLUSIONS: These findings do not support a crucial role of circulating 25(OH)D as a regulator of ß-cell autoimmunity or FOXP3 expression.

Increased peripheral blood CD4+ T cell responses to deamidated but not to native gliadin in children with coeliac disease.

T cell recognition of gliadin from dietary gluten is essential for the pathogenesis of coeliac disease (CD). The aim of the present study was to analyse whether gliadin-specific T cells are detectable in the circulation of children with newly diagnosed coeliac disease by using a sensitive carboxfluorescein diacetate succinimidyl ester (CFSE) dilution method. Peripheral blood CD4(+) T cell responses were analysed in 20 children at diagnosis of CD and compared to those in 64 healthy control children carrying the CD-associated human leucocyte antigen (HLA)-DQ2 or -DQ8 alleles. Deamidated gliadin (gTG)-specific T cells were detectable in the peripheral blood of more than half the children with CD (11 of 20, 55%) compared to 15 of 64 (23.4%) of the control children (P = 0.008). Proliferative responses to gTG were also significantly stronger in children with CD than in controls (P = 0.01). In contrast, T cells specific to native gliadin were detectable at comparable frequencies in children with CD (two of 19, 10.5%) and controls (13 of 64, 20.3%). gTG-specific T cells had a memory phenotype more often than those specific to native gliadin in children with CD (P = 0.02), whereas controls had similar percentages of memory cells in both stimulations. Finally, gTG-specific CD4(+) T cells had a higher expression of the gut-homing molecule ß7 integrin than those specific to the control antigen tetanus toxoid. Collectively, our current results demonstrate that the frequency of circulating memory CD4(+) T cells specific to gTG but not native gliadin is increased in children with newly diagnosed CD.

Altered expression of intestinal human leucocyte antigen D-related and immune signalling molecules in juvenile idiopathic arthritis.

We aimed to study intestinal immune activation status in juvenile idiopathic arthritis (JIA) by assessing intestinal human leucocyte antigen (HLA) class II expression and the mRNA expression levels of the pro- and anti-inflammatory mediators and pattern recognition receptors. HLA-D-related (HLA-DR) expression was assessed using immunohistochemical staining of frozen sections in 11 children with JIA and 17 controls. The gene expression levels of the anti- and proinflammatory cytokines, lymphocyte recognition receptors and pattern recognition receptors were studied with reverse transcription-polymerase chain reaction (RT-PCR) in 14 children with JIA and 12 controls. All subjects had various gastrointestinal (GI) symptoms indicating endoscopic examinations, but eventually were not diagnosed with GI disease. In JIA patients, the expression of HLA-DR was increased in the crypt epithelial cells and in the epithelial basement membrane of the ileum when compared with the controls. Positive HLA-DR staining in the ileal mucosa was associated with the presence of high clinical disease activity of JIA and low mRNA expression of anti-inflammatory mediators, such as forkhead box protein P3 (FoxP3), glucocorticoid-induced tumour necrosis factor receptor-related protein (GITR) and transforming growth factor (TGF)-beta. Low ileal expression of interleukin (IL)-10, TGF-ß, FoxP3, Toll-like receptor 2 (TLR-2) and TLR-4 transcripts correlated significantly with a high clinical disease activity in the JIA patients. The increased HLA-DR expression suggests enhanced intestinal antigen presentation in JIA. A correlation between clinical disease activity and low gene expression of tolerogenic mediators in the ileum supports the hypothesis that a link exists between the gut immune system and JIA.

Up-regulation of small intestinal interleukin-17 immunity in untreated coeliac disease but not in potential coeliac disease or in type 1 diabetes.

Up-regulation of interleukin (IL)-17 in small intestinal mucosa has been reported in coeliac disease (CD) and in peripheral blood in type 1 diabetes (T1D). We explored mucosal IL-17 immunity in different stages of CD, including transglutaminase antibody (TGA)-positive children with potential CD, children with untreated and gluten-free diet-treated CD and in children with T1D. Immunohistochemistry was used for identification of IL-17 and forkhead box protein 3 (FoxP3)-positive cells and quantitative polymerase chain reaction (qPCR) for IL-17, FoxP3, retinoic acid-related orphan receptor (ROR)c and interferon (IFN)-γ transcripts. IL-1ß, IL-6 and IL-17 were studied in supernatants from biopsy cultures. Expression of the apoptotic markers BAX and bcl-2 was evaluated in IL-17-stimulated CaCo-2 cells. The mucosal expression of IL-17 and FoxP3 transcripts were elevated in individuals with untreated CD when compared with the TGA-negative reference children, children with potential CD or gluten-free diet-treated children with CD (P < 0·005 for all IL-17 comparisons and P < 0·01 for all FoxP3 comparisons). The numbers of IL-17-positive cells were higher in lamina propria in children with CD than in children with T1D (P < 0·05). In biopsy specimens from patients with untreated CD, enhanced spontaneous secretion of IL-1ß, IL-6 and IL-17 was seen. Activation of anti-apoptotic bcl-2 in IL-17-treated CaCo-2 epithelial cells suggests that IL-17 might be involved in mucosal protection. Up-regulation of IL-17 could, however, serve as a biomarker for the development of villous atrophy and active CD.

Interaction of enterovirus infection and cow's milk-based formula nutrition in type 1 diabetes-associated autoimmunity.

BACKGROUND: Enteral virus infections and early introduction of cow's milk (CM)-based formula are among the suggested triggers of type 1 diabetes (T1D)-associated autoimmunity, although studies on their role have remained contradictory. Here, we aimed to analyse whether interactions between these factors might clarify the controversies. MATERIALS: The study population comprised 107 subjects developing positivity for at least two T1D-associated autoantibodies and 446 control subjects from the Finnish diabetes prediction and prevention cohort. Enterovirus, rotavirus, adenovirus, respiratory syncytial virus and bovine insulin-binding antibodies were analysed from prospective serum samples at 3-24 months of age. Data on infant cow's milk exposure were available for 472 subjects: 251 subjects were exposed to cow's milk before 3 months of age and 221 subjects later in infancy. RESULTS: Signs of an enterovirus infection by 12 months of age were associated with the appearance of autoimmunity among children who were exposed to cow's milk before 3 months of age. Cox regression analysis revealed a combined effect of enterovirus infection and early cow's milk exposure for the development of ICA and any of the biochemically defined autoantibodies (p = 0.001), of IAA (p = 0.002), GADA (p = 0.001) and IA-2A (p = 0.013). CONCLUSIONS: The effect of enterovirus infection on the appearance of T1D-associated autoimmunity seems to be modified by exposure to cow's milk in early infancy suggesting an interaction between these factors. Moreover, these results provide an explanation for the controversial findings obtained when analysing the effect of any single one of these factors on the appearance of T1D-associated autoimmunity.

Intra-articular corticoid injection induces circulating glucocorticoid bioactivity and systemic immune activation in juvenile idiopathic arthritis.

OBJECTIVE: To study the systemic effects of intra-articular (IA) glucocorticoid (GC) injections in juvenile idiopathic arthritis (JIA). METHODS: The study group comprised 21 JIA patients being treated with IA methylprednisolone [MP (n = 15) or MP plus triamcinolone hexacetonide (THA) (n = 6)] prescribed on clinical indications. The systemic effect of MP was assessed by measuring circulating glucocorticoid bioactivity (GBA) with a recombinant cell transactivation assay 7 and 24 h after the IA injections, and after 2 months. The systemic immunological responses were studied with a novel assay for testing patient serum-induced changes in the secretion of interferon (IFN)-γ and interleukin (IL)-5 from target cells. RESULTS: Administration of IA GC induced serum GBA (p = 0.001) and suppressed circulating cortisol levels (p = 0.002) 7 h after the injection. Serum withdrawn 24 h after the IA injection induced less IL-5 secretion from mitogen-activated target cells when compared with pre-treatment sera (p = 0.036). This decrease in target cell T helper (Th)2 response (IL-5) was MP dose related (r = -0.550, p = 0.018). High IL-5 secretion from target cells prior to the IA injections was associated with good clinical outcome at 2 months, seen as a low number of active (p = 0.044) and restricted joints (p = 0.049). CONCLUSION: IA GC injections have systemic effects that are reflected in the serum as an immediate elevation of GBA, a decrease of endogenous cortisol as well as a suppressive effect of patient serum on target cell IL-5 secretion. These systemic effects may play a role in the attenuation of disease activity.

Enhanced influenza A H1N1 T cell epitope recognition and cross-reactivity to protein-O-mannosyltransferase 1 in Pandemrix-associated narcolepsy type 1.

Narcolepsy type 1 (NT1) is a chronic neurological disorder having a strong association with HLA-DQB1*0602, thereby suggesting an immunological origin. Increased risk of NT1 has been reported among children or adolescents vaccinated with AS03 adjuvant-supplemented pandemic H1N1 influenza A vaccine, Pandemrix. Here we show that pediatric Pandemrix-associated NT1 patients have enhanced T-cell immunity against the viral epitopes, neuraminidase 175-189 (NA175-189) and nucleoprotein 214-228 (NP214-228), but also respond to a NA175-189-mimic, brain self-epitope, protein-O-mannosyltransferase 1 (POMT1675-689). A pathogenic role of influenza virus-specific T-cells and T-cell cross-reactivity in NT1 are supported by the up-regulation of IFN-γ, perforin 1 and granzyme B, and by the converging selection of T-cell receptor TRAV10/TRAJ17 and TRAV10/TRAJ24 clonotypes, in response to stimulation either with peptide NA175-189 or POMT1675-689. Moreover, anti-POMT1 serum autoantibodies are increased in Pandemrix-vaccinated children or adolescents. These results thus identify POMT1 as a potential autoantigen recognized by T- and B-cells in NT1.

Scientific rationale for the Finnish Allergy Programme 2008-2018: emphasis on prevention and endorsing tolerance.

In similarity to many other western countries, the burden of allergic diseases in Finland is high. Studies worldwide have shown that an environment rich in microbes in early life reduces the subsequent risk of developing allergic diseases. Along with urbanization, such exposure has dramatically reduced, both in terms of diversity and quantity. Continuous stimulation of the immune system by environmental saprophytes via the skin, respiratory tract and gut appears to be necessary for activation of the regulatory network including regulatory T-cells and dendritic cells. Substantial evidence now shows that the balance between allergy and tolerance is dependent on regulatory T-cells. Tolerance induced by allergen-specific regulatory T-cells appears to be the normal immunological response to allergens in non atopic healthy individuals. Healthy subjects have an intact functional allergen-specific regulatory T-cell response, which in allergic subjects is impaired. Evidence on this exists with respect to atopic dermatitis, contact dermatitis, allergic rhinitis and asthma. Restoration of impaired allergen-specific regulatory T-cell response and tolerance induction has furthermore been demonstrated during allergen-specific subcutaneous and sublingual immunotherapy and is crucial for good therapeutic outcome. However, tolerance can also be strengthened unspecifically by simple means, e.g. by consuming farm milk and spending time in nature. Results so far obtained from animal models indicate that it is possible to restore tolerance by administering the allergen in certain circumstances both locally and systemically. It has become increasingly clear that continuous exposure to microbial antigens as well as allergens in foodstuffs and the environment is decisive, and excessive antigen avoidance can be harmful and weaken or even prevent the development of regulatory mechanisms. Success in the Finnish Asthma Programme was an encouraging example of how it is possible to reduce both the costs and morbidity of asthma. The time, in the wake of the Asthma Programme, is now opportune for a national allergy programme, particularly as in the past few years, fundamentally more essential data on tolerance and its mechanisms have been published. In this review, the scientific rationale for the Finnish Allergy Programme 2008-2018 is outlined. The focus is on tolerance and how to endorse tolerance at the population level.

Infiltration of forkhead box P3-expressing cells in small intestinal mucosa in coeliac disease but not in type 1 diabetes.

Because the role of regulatory T cells in the intestinal inflammation is unknown in coeliac disease (CD) and type 1 diabetes (T1D), the expression of forkhead box P3 (FoxP3), CD25, transforming growth factor-beta, interferon (IFN)-gamma, interleukin (IL)-4, IL-8, IL-10, IL-15 and IL-18 was measured by quantitative reverse transcription-polymerase chain reaction in the small intestinal biopsies from paediatric patients with active or potential CD, T1D and control patients. The numbers of FoxP3- and CD25-expressing cells were studied with immunohistochemistry. Enhanced intestinal expressions of FoxP3, IL-10 and IFN-gamma mRNAs were found in active CD when compared with controls (P-values < 0.001, 0.004, <0.001). In potential CD, only the expression of IFN-gamma mRNA was increased. The numbers of FoxP3-expressing cells were higher in active and potential CD (P < 0.001, P = 0.05), and the ratio of FoxP3 mRNA to the number of FoxP3-positive cells was decreased in potential CD when compared with controls (P = 0.007). The ratio of IFN-gamma to FoxP3-specific mRNA was increased in active and potential CD (P = 0.001 and P = 0.002). Patients with T1D had no changes in regulatory T cell markers, but showed increased expression of IL-18 mRNA. The impaired up-regulation of FoxP3 transcripts despite the infiltration of FoxP3-positive cells in potential CD may contribute to the persistence of inflammation. The increased ratio of IFN-gamma to FoxP3 mRNA in active and potential CD suggests an imbalance between regulatory and effector mechanisms. The increased intestinal expression of IL-18 mRNA in patients with T1D adds evidence in favour of the hypothesis that T1D is associated with derangements in the gut immune system.

Probiotics in infancy induce protective immune profiles that are characteristic for chronic low-grade inflammation.

BACKGROUND: Probiotics are widely studied both in the treatment and prevention of allergic diseases, but their mode of action is poorly known. OBJECTIVE: Our aim was to examine the effect of probiotic bacteria on in vivo cytokine, antibody, and inflammatory responses in allergy-prone infants. METHODS: In a randomized double-blind study, probiotic bacteria or placebo were given for 1 month before delivery to mothers and for 6 months to infants with a family history of allergy. Plasma samples were analysed for C-reactive protein (CRP), total IgA and IgE, food-specific IgA, IgG, and IgE, IL-2, IL-4, IL-6, IL-10, TNF-alpha, and IFN-gamma. We analysed the associations of immunological and inflammatory parameters at age 6 months with probiotic treatment and allergic phenotype at 2 years. RESULTS: Infants receiving probiotic bacteria had higher plasma levels of CRP (P=0.008), total IgA (P=0.016), total IgE (P=0.047), and IL-10 (P=0.002) than infants in the placebo group. Increased plasma CRP level at age 6 months was associated with a decreased risk of eczema [odds ratio (OR) 0.41 [95% confidence interval (CI) 0.17-0.99], P=0.046], and with a decreased risk of allergic disease [OR 0.38 (95% CI 0.16-0.87), P=0.023] at age 2 years, when adjusted with probiotic use. CONCLUSION: The association of CRP with a decreased risk of eczema at 2 years of age in allergy-prone children supports the view that chronic, low-grade inflammation protects from eczema. Probiotic-induced low-grade inflammation was characterized by elevation of IgE, IgA, and IL-10, the changes typically observed in helminth infection-associated induction of regulatory mechanisms. The findings emphasize the role of chronic microbial exposure as an immune modulator protecting from allergy.

Relation among mannose-binding lectin 2 genotype, beta-cell autoantibodies, and risk for type 1 diabetes in Finnish children.

Mannose-binding lectin (MBL) is a key mediator of innate immunity, the insufficiency of which is caused by point mutations in the MBL2 gene. MBL insufficiency is associated with increased susceptibility to infections and certain autoimmune diseases, but its impact in the pathogenesis and risk of type 1 diabetes (T1D) is controversial. We investigated the significance of the MBL2 genotype on the risk of T1D in a Finnish study population comprising 470 diabetic children and 501 controls. Furthermore, the effect of MBL2 gene polymorphism on the emergence of beta-cell autoantibodies in 289 unaffected children with human leukocyte antigen-conferred susceptibility to T1D was assessed. MBL genotype had no significant effect on the risk or onset age of T1D. However, children with the biallelic variant genotype reflecting total MBL deficiency tested positive more frequently for > or =3 autoantibodies compared with children with another genotype (odds ratio = 6.0, 95% confidence interval 1.3-28; p = 0.013). In conclusion, the MBL2 genotype did not affect susceptibility to T1D in children, and this finding does not support previous reports implicating a role of the MBL2 genotype as a factor predisposing to T1D. The association of the biallelic variant genotype with positivity for multiple autoantibodies suggests that intermolecular epitope spreading may be linked with impaired clearance of autoantigens as a result of MBL deficiency.

Effect of HLA genotype or CTLA-4 polymorphism on cytokine response in healthy children.

Type 1 diabetes (T1D) is considered to be a T-cell-mediated autoimmune disease in which genetic predisposition is affected by HLA class II alleles and polymorphisms in cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) gene. We tested the hypothesis whether these T1D-related gene polymorphisms modulate cytokine response and thus contribute to the development of autoimmunity. The study includes 67 non-diabetic children, typed for HLA class II alleles and CTLA-4 polymorphisms (+49A/G, CT60A/G, CTBC217_1C/T). We measured cytokine secretion of peripheral blood mononuclear cells after stimulation with tetanus toxoid (TT), polio virus, coxsackie virus B4, pertussis toxin (PT) and phytohemagglutinin (PHA). We saw higher IL-13 response to TT in individuals with DR3-DQ2 haplotype (P = 0.002). HLA class II protective haplotype, DR2-DQ6, showed association with increased production of IFN-gamma (P < 0.001) and IL-2 (P = 0.005) in response to polio virus. In children with the autoimmunity-related homozygous genotypes CTLA-4 +49G/G, CT60G/G and CTBC217_1T/T, we found enhanced PT- and PHA-induced IFN-gamma production (P < 0.05). The cytokine responses to studied antigens were weakly modified by HLA class II risk haplotypes, and children with T1D-associated HLA risk haplotypes are not specifically inclined to develop an immune response in general. Higher IFN-gamma and IL-2 response to enterovirus in children with HLA class II protective haplotype DR2-DQ6 could be of importance in the protection from T1D-associated enterovirus infections. All autoimmunity related CTLA-4 polymorphisms were associated with enhanced IFN-gamma. This suggests impaired downregulation of cellular immunity by these CTLA-4 polymorphisms.

Maternal diet during pregnancy and lactation and cow's milk allergy in offspring.

BACKGROUND/OBJECTIVES: Diet during pregnancy and lactation may have a role in the development of allergic diseases. There are few human studies on the topic, especially focusing on food allergies. We sought to study the associations between maternal diet during pregnancy and lactation and cow's milk allergy (CMA) in offspring. SUBJECTS/METHODS: A population-based birth cohort with human leukocyte antigen-conferred susceptibility to type 1 diabetes was recruited in Finland between 1997 and 2004 (n=6288). Maternal diet during pregnancy and lactation was assessed by a validated, 181-item semi-quantitative food frequency questionnaire. Register-based information on diagnosed CMA was obtained from the Social Insurance Institution and completed with parental reports. The associations between maternal food consumption and CMA were assessed using logistic regression, comparing the highest and the lowest quarters to the middle half of consumption. RESULTS: Consumption of milk products in the highest quarter during pregnancy was associated with a lower risk of CMA in offspring (odds ratio (OR) 0.56, 95% confidence interval (CI) 0.37-0.86; P<0.01). When stratified by maternal allergic rhinitis and asthma, there was evidence of an inverse association between high use of milk products and CMA in offspring of non-allergic mothers (OR 0.30, 95% CI 0.13-0.69, P<0.001). Cord blood IgA correlated positively with the consumption of milk products during pregnancy, indicating exposure to CMA and activation of antigen-specific immunity in the infant during pregnancy. CONCLUSIONS: High maternal consumption of milk products during pregnancy may protect children from developing CMA, especially in offspring of non-allergic mothers.

Glutamate decarboxylase-reactive peripheral blood lymphocytes from patients with IDDM express gut-specific homing receptor alpha4beta7-integrin.

Migration of lymphocytes to the pancreas is a prerequisite for insulitis in IDDM. Mucosal vascular addressin (MAdCAM-1), involved in the recirculation of lymphocytes to the gut, has been found in the inflamed islets in NOD mice. In humans, triggers of the gut immune system (e.g., early exposure to cow's milk proteins in infancy, exposure to enteroviral infections) have been associated with IDDM. To study the possible link between the gut immune system and IDDM, we tested the expression of the alpha4beta7-integrin, a homing receptor for MAdCAM-1, on GAD65-reactive lymphocytes. Using immunomagnetic cell sorting, we depleted the lymphocytes with high expression of alpha4beta7-integrin in the peripheral blood mononuclear cell population from IDDM patients and patients with autoimmune polyendocrine disease type 1 (APD-I). The depletion led to a marked decrease (mean 70%) in the cellular response against GAD65 in three of six IDDM patients and in one subject at high risk for IDDM. A decrease of 37% in the GAD response was observed after depletion in the case of one APD-I patient who also had IDDM. Cellular response to tetanus toxoid increased in the majority of patients as well as in three control subjects studied. We demonstrated that a remarkable population of islet cell antigen-reactive lymphocytes express the gut-specific homing receptor, which emphasizes the role of gut immunity in IDDM. The manipulation of the gut immune system is therefore proposed as a tool for modulation of the autoimmunity against pancreatic beta-cells in IDDM.

Effect of cow's milk exposure and maternal type 1 diabetes on cellular and humoral immunization to dietary insulin in infants at genetic risk for type 1 diabetes. Finnish Trial to Reduce IDDM in the Genetically at Risk Study Group.

Type 1 diabetes is considered to be a T-cell-mediated autoimmune disease in which insulin-producing beta-cells are destroyed. Immunity to insulin has been suggested to be one of the primary autoimmune mechanisms leading to islet cell destruction. We have previously shown that the first immunization to insulin occurs by exposure to bovine insulin (BI) in cow's milk (CM) formula. In this study, we analyzed the development of insulin-specific T-cell responses by proliferation test, emergence of insulin-binding antibodies by enzyme immunoassay, and insulin autoantibodies by radioimmunoassay in relation to CM exposure and family history of type 1 diabetes in infants with a first-degree relative with type 1 diabetes and increased genetic risk for the disease. The infants were randomized to receive either an adapted CM-based formula or a hydrolyzed casein (HC)-based formula after breast-feeding for the first 6-8 months of life. At the age of 3 months, both cellular and humoral responses to BI were higher in infants exposed to CM formula than in infants fully breast-fed (P = 0.015 and P = 0.007). IgG antibodies to BI were higher in infants who received CM formula than in infants who received HC formula at 3 months of age (P = 0.01), but no difference in T-cell responses was seen between the groups. T-cell responses to BI at 9 months of age (P = 0.05) and to human insulin at 12 (P = 0.014) and 24 months of age (P = 0.009) as well as IgG antibodies to BI at 24 months of age (P = 0.05) were lower in children with a diabetic mother than in children with a diabetic father or a sibling, suggesting possible tolerization to insulin by maternal insulin therapy. The priming of insulin-specific humoral and T-cell immunity occurs in early infancy by dietary insulin, and this phenomenon is influenced by maternal type 1 diabetes.