Search

Background. Recent epidemiological studies in inflammatory bowel disease (IBD) indicate that paediatric onset of IBD (pIBD) more often requires biological therapy compared to adult onset of IBD (aIBD). Whether this is due to a more aggressive disease phenotype or lower threshold of prescribing biologicals is unknown. In order to expand these findings in a clinical setting, we compared the inflammatory burden in pIBD and aIBD patients requiring biological therapy. Methods. We retrospectively included 70 pIBD and 83 aIBD patients initiating biological therapy. Symptoms and biomarker levels were recorded prior to and 6, 14, 22, and 52 weeks after initiation of biological therapy. Results. In Crohn’s disease (CD), the baseline levels of faecal calprotectin and C-reactive protein (CRP) were increased in paediatric CD patients compared to adult CD patients ( p < 0.0001 and p = 0.01 , respectively). No significant differences were seen in ulcerative colitis (UC). In CD, baseline vitamin D levels ≥ 75 nmol/L and baseline CRP levels < 5 mg/L were associated with higher remission rate ( p = 0.02 ) at the end of follow-up. Moreover, aIBD patients had a higher risk of loss of response to biological therapy and treatment discontinuation compared to pIBD patients ( HR = 4.7 [1.6-13.4], p = 0.004 ). Conclusions. pCD patients had increased inflammation markers compared to aCD patients prior to biological treatment. In addition to this, vitamin D < 75 nmol/L and high CRP levels predicted poor response to treatment in IBD patients.

Background: Vitamin D treatment may reduce Crohn&rsquo, s disease (CD) activity by modulating the mucosal immune function. We investigated if high-dose vitamin D +/&minus, infliximab modulated the mucosal cytokine expression in active CD. Methods: Forty CD patients were randomized into: infliximab + vitamin D, infliximab + placebo-vitamin D, placebo-infliximab + vitamin D or placebo-infliximab + placebo-vitamin D. Infliximab (5 mg/kg) and placebo-infliximab were administered at weeks 0, 2 and 6. Oral vitamin D was administered as bolus 200,000 international units (IU) per week 0 followed by 20,000 IU/day for 7 weeks or placebo. Endoscopy with biopsies was performed at weeks 0 and 7 where endoscopic activity was measured and mucosal mRNA cytokine expression was examined. C-reactive protein (CRP), fecal calprotectin and Harvey-Bradshaw Index (HBI) were measured at weeks 0, 2 and 6. Results: High-dose vitamin D treatment alone and combined with infliximab decreased the IL17A, IFN&gamma, and IL10 expression. High-dose vitamin D alone did not significantly decrease the disease activity, CRP or calprotectin. Combined infliximab and vitamin D treatment was not clinically significantly superior to monotherapy with infliximab. Conclusions: High-dose vitamin D as monotherapy and combined with infliximab decreases IL17A, IFN&gamma, and IL-10 expression in mucosa within treatment groups. This did not induce a statistically significant decreased disease activity. EudraCT no.2013-000971-34.

Inflammatory bowel diseases (IBDs) may result from dysregulated mucosal immune responses directed toward the resident intestinal microbiota. This review describes the hallmark immunobiology of Crohn's disease and ulcerative colitis as well as therapeutic targets and mechanisms of action for current, experimental, and future treatments in IBD. Conventional therapies include 5-aminosalicylic acid, glucocorticosteroids, thiopurines, and methotrexate. Since 1997, monoclonal antibodies have gained widespread use. These consist of antibodies directed against pro-inflammatory cytokines such as tumor necrosis factor α, interleukin (IL)-12, and IL-23, or anti-homing antibodies directed against α4β7 integrin. Emerging oral therapies include modulators of intracellular signal transduction such as Janus kinase inhibitors. Vitamin D may help to regulate innate and adaptive immune responses. Modulation of the intestinal microbiota, using live microorganisms (probiotics), substrates for the colonic microbiota (prebiotics), or fecal microbiota transplantation (FMT), is in development. Dietary supplements are in widespread use, but providing evidence for their benefit is challenging. Stem cell treatment and nervous stimulation are promising future treatments.

PURPOSE: Vitamin D may induce tolerance in the intestinal immune system and has been shown to regulate the phenotype of tolerogenic intestinal dendritic cells (DCs) in vitro. It is unknown whether vitamin D supplementation affects human intestinal DCs in vivo, and we aimed to investigate the tolerability and effect on intestinal CD103+DCs of high-dose vitamin D3 treatment in healthy subjects.METHODS: Ten healthy subjects received a total of 480,000 IU oral vitamin D3 over 15 days and colonic biopsies were obtained before and after intervention by endoscopy. Lamina propria mononuclear cells (LPMCs) were isolated from the biopsies, stained with DC surface markers and analysed with flow cytometry. Snap-frozen biopsies were analysed with qPCR for DC and regulatory T cell-related genes.RESULTS: No hypercalcemia or other adverse events occurred in the test subjects. Vitamin D decreased the number of CD103+ DCs among LPMCs (p = 0.006). Furthermore, vitamin D induced mRNA expression of TGF-β (p = 0.048), TNF-α (p = 0.006) and PD-L1 (p = 0.02) and tended to induce IL-10 expression (p = 0.06). Multivariate factor analysis discriminated between pre- and post-vitamin D supplementation with a combined increased qPCR expression of PD1, PD-L1, TGF-β, IL-10, CD80, CD86, FOXP3, NFATc2 and cathelicidin.CONCLUSION: High-dose vitamin D supplementation is well tolerated by healthy subjects and has a direct effect on the CD103+ DCs, local cytokine and surface marker mRNA expression in the colonic mucosa, suggestive of a shift towards a more tolerogenic milieu.

// Mia Bendix 1 , Stinne Greisen 2,3 , Anders Dige 1 , Christian L. Hvas 1 , Nina Bak 1 , Soren P. Jorgensen 1 , Jens F. Dahlerup 1 , Bent Deleuran 2,3 and Jorgen Agnholt 1 1 Department of Hepatology and Gastroenterology, Aarhus University Hospital, Aarhus, Denmark 2 Department of Immunology, Institute of Biomedicine, Aarhus University, Aarhus, Denmark 3 Department of Rheumatology, Aarhus University Hospital, Aarhus, Denmark Correspondence to: Mia Bendix, email: // Keywords : PD-1, Crohn’s disease, vitamin D, T cells, Immunology and Microbiology Section, Immune response, Immunity Received : November 18, 2016 Accepted : February 07, 2017 Published : February 18, 2017 Abstract Background: Vitamin D modulates inflammation in Crohn’s disease (CD). Programmed death (PD)-1 receptor contributes to the maintenance of immune tolerance. Vitamin D might modulate PD-1 signalling in CD. Aim: To investigate PD-1 expression on T cell subsets in CD patients treated with vitamin D or placebo. Methods: We included 40 CD patients who received 1200 IU vitamin D3 for 26 weeks or placebo and eight healthy controls. Peripheral blood mononuclear cells (PBMCs) and plasma were isolated at baseline and week 26. The expressions of PD-1, PD-L1, and surface activation markers were analysed by flow cytometry. Soluble PD-1 plasma levels were measured by ELISA. Results: PD-1 expression upon T cell stimulation was increased in CD4 + CD25 +int T cells in vitamin D treated CD patients from 19% (range 10 – 39%) to 29% (11 – 79%)( p = 0.03) compared with placebo-treated patients. Vitamin D treatment, but not placebo, decreased the expression of the T cell activation marker CD69 from 42% (31 – 62%) to 33% (19 - 54%)( p = 0.01). Soluble PD-1 levels were not influenced by vitamin D treatment. Conclusions: Vitamin D treatment increases CD4 + CD25 +int T cells ability to up-regulate PD-1 in response to activation and reduces the CD69 expression in CD patients.

BACKGROUND: In ulcerative colitis (UC), dietary supplements may have anti-inflammatory properties and improve disease course. We investigated the effects of casein glycomacropeptide (CGMP), a fraction of bovine whey protein, in active UC.MATERIALS AND METHODS: In a randomized open-label intervention study, 24 patients with active UC involving 10-40 cm of the distal colon were randomized in a 2 : 1 ratio into two groups. The first group was administered their usual treatment plus a daily supplement of CGMP 30 g, and the second group was administered a dose escalation to 4800 mg oral mesalamine daily (standard treatment) for 4 weeks. Clinical, endoscopic, mucosal and circulating disease activity markers were monitored. Acceptance of and adherence to CGMP up to 8 weeks were documented.RESULTS: After 4 weeks of treatment, 10 of 16 (63%) patients who received CGMP had an unchanged or decreased Simple Clinical Colitis Activity Index (SCCAI), which was similar to the four of eight (50%) (P = 0·67) patients on the standard treatment. The number of patients in which SCCAI decreased by three or more did not differ between the two groups: nine of 16 (56%) in the CGMP group vs. four of eight (50%) in the standard treatment group (P = 0·77). Changes in disease extent and severity were similar between the two groups. CGMP was well tolerated and accepted by the patients.CONCLUSIONS: The addition of CGMP as a nutritional therapy to standard treatment was safe and accepted by patients with active distal UC. The disease-modifying effect of CGMP was similar to that of the mesalamine dose escalation.

Summary Crohn's disease (CD) is a chronic inflammatory disease associated with a dysregulated T cell response towards intestinal microflora. Vitamin D has immune modulatory effects on T cells through the nuclear vitamin D receptor (VDR) in vitro. It is unclear how oral vitamin D treatment affects VDR expression. The aim of this study was to establish a flow cytometry protocol, including nuclear and cytoplasmic VDR expression, and to investigate the effects of vitamin D treatment on T cell VDR expression in CD patients. The flow cytometry protocol for VDR staining was developed using the human acute monocytic leukaemia cell line (THP-1). The protocol was evaluated in anti-CD3/CD28-stimulated peripheral blood mononuclear cells (PBMCs) from vitamin D3- (n = 9) and placebo-treated (n = 9) CD patients. Anti-VDR-stained PBMCs were examined by flow cytometry, and their cytokine production was determined by cytokine bead array. VDR, CYP27B1 and RXRα mRNA expression levels in CD4+ T cells were measured by quantitative reverse transcriptase polymerase chain reaction. The flow cytometry protocol enabled detection of cytoplasmic and nuclear VDR expression. The results were confirmed by confocal microscopy and supported by correlation with VDR mRNA expression. VDR expression in CD4+ T cells increased following stimulation. This VDR up-regulation was inhibited with 30% by vitamin D treatment compared to placebo in CD patients (P = 0·027). VDR expression was correlated with in-vitro interferon-γ production in stimulated PBMCs (P = 0·01). Flow cytometry is a useful method with which to measure intracellular VDR expression. Vitamin D treatment in CD patients reduces T cell receptor-mediated VDR up-regulation.

Intestinal CD4+T cell depletion is rapid and profound during early HIV-1 infection. This leads to a compromised mucosal barrier that prompts chronic systemic inflammation. The preferential loss of intestinal T helper 17 (Th17) cells in HIV-1 disease is a driver of the damage within the mucosal barrier and of disease progression. Thus, understanding the effects of new therapeutic strategies in the intestines has high priority. Histone deacetylase (HDAC) inhibitors (e.g., panobinostat) are actively under investigation as potential latency reversing agents in HIV eradication studies. These drugs have broad effects that go beyond reactivating virus, including modulation of immune pathways. We examined colonic biopsies from ART suppressed HIV-1 infected individuals (clinicaltrials.gov:NCT01680094) for the effects of panobinostat on intestinal T cell activation and on inflammatory cytokine production. We compared biopsy samples that were collected before and during oral panobinostat treatment and observed that panobinostat had a clear biological impact in this anatomical compartment. Specifically, we observed a decrease in CD69+intestinal lamina propria T cell frequency and increased IL-17A mRNA expression in the intestinal epithelium. These results suggest that panobinostat therapy may influence the restoration of mucosal barrier function in these patients.

Aliment Pharmacol Ther 2010; 32: 1364–1372 Summary Background Vitamin D3 has shown immune-modulating effects in CD4+ T cells from Crohn’s disease patients in vitro. Aim To investigate the effects of in vivo vitamin D3 treatment on T cells in Crohn’s disease patients. Methods Peripheral blood mononuclear cells (PBMC) were isolated at week 0 and at week 26 from 10 vitamin D3- and 10 placebo-treated Crohn’s disease patients participating in a randomized, placebo-controlled, clinical trial study. Monocyte-depleted PBMC were stimulated with anti-CD3 and anti-CD28, and cultured for 7, days, to investigate CD4+ T-cell proliferation and T-cell cytokine production. Results In vitamin D3-treated patients, the median 25-hydroxyvitamin D3 levels increased 70 nmol/L compared with −5 nmol/L in the placebo group. Vitamin D3 treatment increased interleukin-6 production (delta = 188 pg/mL, range: −444 to 4071) compared with a decrease in the placebo group (delta = −896 pg/mL, range: −3841 to 1323) (P

Low serum vitamin D levels may provoke or aggravate Crohn's disease (CrD). Vitamin D3 is a well-known immune modulator that affects immune functions in vitro and may prevent CrD flares. Dendritic cells (DC) are key mediators of vitamin D3 effects. In this study, we describe changes in monocyte-derived DC (mo-DC) maturation marker expression and cytokine production following 26 weeks of oral vitamin D3 supplementation in CrD patients.Ten CrD patients who had increased serum 25-hydroxy vitamin D levels after oral vitamin D3 and calcium treatment and ten seasonally matched placebo-treated patients were selected for this study. Mo-DC were generated before and after the 26 weeks and induced to mature upon lipopolysaccharide (LPS) stimulation. Maturation marker expression and cytokine production were analysed. Mo-DC function was analysed in a mixed leucocyte reaction (MLR).Compared with baseline values, LPS-matured mo-DC exhibited reduced expression of CD80 and reduced production of the cytokines IL-10, IL-1β, and IL-6 following 26 weeks of oral vitamin D3 supplementation. Mo-DC performance in an allogeneic MLR was unchanged after vitamin D3 supplementation. Treatment with the placebo did not affect maturation markers, cytokine production, or the MLR.Vitamin D3 treatment in CrD patients led to hypo-responsive LPS-stimulated mo-DC. This finding indicates that vitamin D3 levels have an impact on the monocytic precursors of mo-DC in vivo and may explain the positive effects of vitamin D3 supplementation on CrD patients. Alternatively, CrD patients with high serum vitamin D3 levels may represent a subgroup with low disease activity.

In Crohn’s disease (CrD), vitamin D may help to balance an exaggerated immune response and thereby improve the disease course. The immunomodulating effects depend on the activation of 25-hydroxy vitamin D3 (25-D3), into 1,25-dihydroxy vitamin D3 (1,25-D3). This activation has previously been shown to take place in dendritic cells (DC) from healthy individuals. We hypothesised that DC from CrD patients are able to regulate and control inflammatory responses through 25-D3 activation. During differentiation, monocyte-derived DC from 20 CrD patients were cultured with either 25-D3 or 1,25-D3 and matured with lipopolysaccharide (LPS). We examined DC surface marker expression, cytokine production, and the ability to induce cell proliferation in an allogeneic mixed leukocyte reaction. Following stimulation with LPS, DC exposed to either 25-D3 or 1,25-D3 exhibited lower expression levels of CD80, CD83, CD86, and HLA-DR and diminished TNF-α production compared with DC cultured with LPS alone. In contrast, CD14 expression and IL-6 production were higher following 25-D3 or 1,25-D3 treatment. Compared with LPS alone, both forms of vitamin D3 reduced the ability of DC to activate lymphocytes. Following stimulation with 25-D3, DC from CrD patients displayed a reduced response to LPS with a diminished capability to activate T cells compared with DC stimulated with LPS alone. These data indicate that intrinsic activation of 25-D3 occurs in DC from CrD patients and show that 25-D3 can modulate DC function in CrD. Our data suggest that vitamin D deficiency may contribute to the uncontrolled inflammatory process seen in CrD.

Anti-TNF-α antibodies has been suggested to modulate regulatory T cell (Treg) percentages in rheumatoid arthritis, but results from studies of Crohn's disease (CD) are conflicting. We investigated dynamic changes of circulating Tregs in CD during treatment with the anti-TNF-α-antibody adalimumab (Humira®, Abbott Laboratories A/S, Emdrupvej 28C, DK-2100 Copenhagen).Blood samples from 26 CD patients were analysed using flow cytometry before and 1 and 26 weeks after initiation of adalimumab treatment to determine the percentage of Tregs among CD4+ T cells.In spite of a significant decline in disease activity scores and biochemical markers of inflammation, during the first week of treatment, we did not observe early modulating effects of adalimumab on Treg percentages. However, we found a long-term increase in Treg percentages in responders who had low Treg percentages (5%) at baseline (p = 0.04). Treg percentage was inversely associated with disease activity (CD activity index or CDAI) (Spearman's rank correlation, ρ = -0.47, p = 0.02). High Treg percentages among CD4+ T cells at baseline predicted clinical response to adalimumab.Adalimumab treatment did not induce early modulatory effects on Treg percentage, even in responders. This finding suggests that adalimumab does not have a direct or selective effect on Tregs. However, Treg percentage was associated with disease activity and high Treg percentage predicted response to adalimumab.

Udgivelsesdato: 2009-Feb-6 We assessed the inter-observer variation for a clinical staging score for tuberculosis (TB). 87 TB patients were examined by 2 independent observers within 2 h, and kappa was calculated. The kappa value of the total Bandim TBscore being at or above 8 points was 0.637, representing a good inter-observer agreement. No systematic bias was found.

To characterize complex coacervates/flocculates of lysozyme and heparin in terms of binding stoichiometry and to determine the effect of complexation on protein structure and stability.Insoluble lysozyme-heparin complexes were formed at pH 7.2 and the binding stoichiometry determined using a solution depletion method. Protein structure was determined by infrared spectroscopy and intrinsic fluorescence. Protein stability was evaluated using differential scanning calorimetry and followed in a 12-weeks storage stability study at 37 degrees C.Binding stoichiometry between heparin and lysozyme was found to be dependent on ionic strength of the solution. At low ionic strength (I approximately 0.01) about 11 lysozyme molecules could bind to a 17 kDa heparin chain, 3 to a 6 kDa chain, and less than 2 to a 3 kDa chain. At higher ionic strength (I approximately 0.1), only 7 lysozyme molecules could bind to a 18 kDa heparin chain.. Above ionic strengths of approximately 0.32 M, no insoluble complexes were observed. Infrared spectroscopy and intrinsic fluorescence did not show any major changes in protein structure upon complexation to heparin. In contrast, differential scanning calorimetry showed a large decrease in the melting temperature of the protein, from 77 degrees C to 61 degrees C. Moreover, after 12-weeks storage at 37 degrees C, only 60% protein recovery was observed for the complexes, with no loss of protein for the uncomplexed protein.Heparin has multiple binding sites for lysozyme, amounting to at most one lysozyme molecule per 3 disaccharide units of heparin. Complexation decreased lysozyme stability, suggesting that heparin has a higher affinity for the unfolded state than the native state. Similar destabilization may occur for other proteins upon interaction with highly charged polymeric compounds or surfaces.