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1.
Aliment Pharmacol Ther ; 47(7): 951-957, 2018 Apr.
Article in English | MEDLINE | ID: mdl-29368342

ABSTRACT

BACKGROUND: Excessive amounts of bile acids entering the colon due to bile acid malabsorption cause chronic bile acid diarrhoea. Diagnosis is possible by measuring the retention fraction of orally ingested 75 Selenium homotaurocholic acid (SeHCAT). The knowledge of long-term effects of medical treatment is sparse. AIM: To describe diarrhoea, adherence to treatment, treatment effects and quality of life in a large, well-defined cohort of patients with bile acid diarrhoea. METHODS: A retrospective survey was performed among 594 patients with bile acid malabsorption verified by SeHCAT scans at our unit between 2003 and 2016. Questionnaires about medical history, diarrhoea, use of medication, and quality of life scores were mailed to all patients. RESULTS: Among 594 patients 377 (69%) responded. Among respondents, 121 (32%) had bile acid diarrhoea due to ileal disease or resection (type 1), 198 (52%) idiopathic bile acid diarrhoea (type 2) and 58 (16%) bile acid diarrhoea due to other non-ileal disease, mainly cholecystectomy (type 3). At follow-up, half of the patients, 184 (50%), reported improvement of diarrhoea. However, 273 patients (74%) still reported diarrhoea and 234 (62%) regularly used anti-diarrhoeal medication. In spite of treatment, 235 (64%) considered reduced quality of life by diarrhoea and 184 (50%) reported that diarrhoea was unaltered or worse than before established diagnosis. CONCLUSION: Many patients with bile acid diarrhoea continue to have bothersome diarrhoea in spite of correct diagnosis and treatment.


Subject(s)
Antidiarrheals/therapeutic use , Bile Acids and Salts/adverse effects , Diarrhea/etiology , Malabsorption Syndromes/diagnosis , Adult , Aged , Bile Acids and Salts/metabolism , Diarrhea/diagnosis , Diarrhea/drug therapy , Diarrhea/metabolism , Female , Humans , Malabsorption Syndromes/drug therapy , Male , Middle Aged , Quality of Life , Radionuclide Imaging/methods , Retrospective Studies , Taurocholic Acid/analogs & derivatives , Time Factors
2.
Eur J Nutr ; 57(7): 2607-2619, 2018 Oct.
Article in English | MEDLINE | ID: mdl-28913556

ABSTRACT

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-ß (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-ß, 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.


Subject(s)
Cholecalciferol/administration & dosage , Colon/cytology , Dendritic Cells/drug effects , Intestinal Mucosa/drug effects , Dendritic Cells/immunology , Dose-Response Relationship, Drug , Healthy Volunteers , Humans , Intestinal Mucosa/cytology , T-Lymphocytes, Regulatory
3.
Mucosal Immunol ; 11(2): 449-461, 2018 03.
Article in English | MEDLINE | ID: mdl-28766555

ABSTRACT

Toll-like receptor 9 (TLR9) agonists are being developed for treatment of colorectal and other cancers, yet the impact of these drugs on human intestines remains unknown. This, together with the fact that there are additional potential indications for TLR9 agonist therapy (e.g., autoimmune and infectious diseases), led us to investigate the impact of MGN1703 (Lefitolimod) on intestinal homeostasis and viral persistence in HIV-positive individuals. Colonic sigmoid biopsies were collected (baseline and week four) from 11 HIV+ individuals on suppressive antiretroviral therapy, who received MGN1703 (60 mg s.c.) twice weekly for 4 weeks in a single-arm, phase 1b/2a study. Within sigmoid mucosa, global transcriptomic analyses revealed 248 modulated genes (false discovery rate<0.05) including many type I interferon (IFN)-stimulated genes. MGN1703 increased the frequencies of cells exhibiting MX1 (P=0.001) and ISG15 (P=0.014) protein expression. No changes were observed in neutrophil infiltration (myeloperoxidase; P=0.97). No systematic effect on fecal microbiota structure was observed (analysis of similarity Global R=-0.105; P=0.929). TLR9 expression at baseline was inversely proportional to the change in integrated HIV DNA during MGN1703 treatment (P=0.020). In conclusion, MGN1703 induced a potent type I IFN response, without a concomitant general inflammatory response, in the intestines.


Subject(s)
Colon, Sigmoid/physiology , DNA/therapeutic use , Gastrointestinal Microbiome/drug effects , HIV Infections/immunology , HIV-1/physiology , Intestines/immunology , Toll-Like Receptor 9/agonists , Colon, Sigmoid/drug effects , Colon, Sigmoid/virology , Cytokines/genetics , Cytokines/metabolism , DNA, Viral/genetics , Female , Gene Expression Profiling , HIV Infections/drug therapy , Homeostasis , Humans , Immunity, Mucosal/drug effects , Interferon Type I/metabolism , Intestines/drug effects , Intestines/virology , Male , Myxovirus Resistance Proteins/genetics , Myxovirus Resistance Proteins/metabolism , Ubiquitins/genetics , Ubiquitins/metabolism , Viral Load/drug effects
4.
Parasite Immunol ; 39(1)2017 Jan.
Article in English | MEDLINE | ID: mdl-27743501

ABSTRACT

Helminthic therapy of immune-mediated diseases has gained attention in recent years, but we know little of how helminths modulate human immunity. In this study, we investigated how self-infection with Trichuris (T.) trichiura in an adult man without intestinal disease affected mucosal and systemic immunity. Colonic mucosal biopsies were obtained at baseline, during T. trichiura infection, and after its clearance following mebendazole treatment. Unexpectedly, the volunteer experienced a Campylobacter colitis following T. trichiura clearance, and this served as a positive infectious control. Trichuris trichiura colonization induced equally increased expressions of T-helper (h)1-, Th2-, Th17- and Treg-associated cytokines and transcription factors, measured by quantitative polymerase chain reaction. We observed several indicators of modulation of systemic immunity during the T. trichiura infection. Plasma eosinophils and anti-Trichuris antibodies rose markedly during the inoculation phase, and a shift towards a Th2-dominated T cell response at the expense of the Th1-response was observed in circulating T cells. Taken together, our findings corroborate that helminths modulate regional and systemic human immunity.


Subject(s)
Immunity, Mucosal , Trichuriasis/immunology , Trichuris/immunology , Adult , Animals , Campylobacter Infections/complications , Cytokines/metabolism , Humans , Intestinal Mucosa , Male , Mebendazole/therapeutic use , T-Lymphocytes, Regulatory/immunology , Trichuriasis/complications
5.
Neurogastroenterol Motil ; 28(2): 217-24, 2016 Feb.
Article in English | MEDLINE | ID: mdl-26729638

ABSTRACT

BACKGROUND: Gastrointestinal (GI) dysmotility may present secondary to inflammatory bowel disease. The main aim of this study was to investigate GI motility in ulcerative colitis (UC) patients during severe disease activity. METHODS: Twenty patients with severe UC were studied with a novel telemetric capsule system (3D-Transit) designed for minimally invasive, ambulatory assessment of total and regional GI transit times. Ten patients were available for follow-up during remission. Data were compared to those of 20 healthy subjects (HS). KEY RESULTS: Total GI transit time was significantly longer in patients with severe UC (median 44.5 h [range 9.9-102.7 h]) than in HS (median 27.6 h [range 9.6-56.4 h]) (p = 0.032). Additionally, during severe UC, transit time was prolonged through the proximal colon (p = 0.003) and there were strong trends toward longer than normal small intestinal transit time (HS: median 4.9 h [range 3.4-8.3 h] vs severe UC patients: median 5.9 h [range 3.9-11.9 h]; p = 0.053) and colorectal transit times (HS: median 18.2 h [range 1.5-43.7] vs severe UC patients: median 34.9 h [range 0.4-90.9 h]; p = 0.056). Our data further indicate that total GI and colorectal transit times may be prolonged in UC during early remission. CONCLUSIONS & INFERENCES: Total GI transit times are significantly prolonged during severe UC.


Subject(s)
Colitis, Ulcerative , Gastrointestinal Transit , Adult , Aged , Aged, 80 and over , Capsule Endoscopy/methods , Female , Gastroenterology/instrumentation , Gastroenterology/methods , Humans , Male , Middle Aged , Telemetry/instrumentation , Telemetry/methods , Young Adult
6.
Mucosal Immunol ; 9(1): 171-82, 2016 Jan.
Article in English | MEDLINE | ID: mdl-26080709

ABSTRACT

Disruption of the homeostatic balance of intestinal dendritic cells (DCs) and macrophages (MQs) may contribute to inflammatory bowel disease. We characterized DC and MQ populations, including their ability to produce retinoic acid, in clinical material encompassing Crohn's ileitis, Crohn's colitis and ulcerative colitis (UC) as well as mesenteric lymph nodes (MLNs) draining these sites. Increased CD14(+)DR(int) MQs characterized inflamed intestinal mucosa while total CD141(+) or CD1c(+) DCs numbers were unchanged. However, CD103(+) DCs, including CD141(+)CD103(+) and CD1c(+)CD103(+) DCs, were reduced in inflamed intestine. In MLNs, two CD14(-) DC populations were identified: CD11c(int)HLADR(hi) and CD11c(hi)HLADR(int) cells. A marked increase of CD11c(hi)HLADR(int) DC, particularly DR(int)CD1c(+) DCs, characterized MLNs draining inflamed intestine. The fraction of DC and MQ populations expressing aldehyde dehydrogenase (ALDH) activity, reflecting retinoic acid synthesis, in UC colon, both in active disease and remission, were reduced compared to controls and inflamed Crohn's colon. In contrast, no difference in the frequency of ALDH(+) cells among blood precursors was detected between UC patients and non-inflamed controls. This suggests that ALDH activity in myeloid cells in the colon of UC patients, regardless of whether the disease is active or in remission, is influenced by the intestinal environment.


Subject(s)
Aldehyde Dehydrogenase/immunology , Colitis, Ulcerative/immunology , Colon/immunology , Crohn Disease/immunology , Dendritic Cells/immunology , Macrophages/immunology , Adolescent , Adult , Aged , Aged, 80 and over , Aldehyde Dehydrogenase/genetics , Antigens, CD/genetics , Antigens, CD/immunology , Antigens, CD1/genetics , Antigens, CD1/immunology , Antigens, Surface/genetics , Antigens, Surface/immunology , CD11c Antigen/genetics , CD11c Antigen/immunology , Case-Control Studies , Colitis, Ulcerative/genetics , Colitis, Ulcerative/pathology , Colon/pathology , Crohn Disease/genetics , Crohn Disease/pathology , Dendritic Cells/pathology , Female , Gene Expression Regulation , Glycoproteins/genetics , Glycoproteins/immunology , HLA-DR Antigens/genetics , HLA-DR Antigens/immunology , Humans , Integrin alpha Chains/genetics , Integrin alpha Chains/immunology , Intestinal Mucosa/immunology , Intestinal Mucosa/pathology , Lipopolysaccharide Receptors/genetics , Lipopolysaccharide Receptors/immunology , Lymph Nodes/immunology , Lymph Nodes/pathology , Macrophages/pathology , Male , Middle Aged , Severity of Illness Index , Signal Transduction , Thrombomodulin
7.
Clin Exp Immunol ; 181(1): 19-28, 2015 Jul.
Article in English | MEDLINE | ID: mdl-25707738

ABSTRACT

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 = 0027). 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.


Subject(s)
CD4-Positive T-Lymphocytes/immunology , Crohn Disease/drug therapy , Receptors, Calcitriol/biosynthesis , Vitamin D/therapeutic use , 25-Hydroxyvitamin D3 1-alpha-Hydroxylase/biosynthesis , 25-Hydroxyvitamin D3 1-alpha-Hydroxylase/genetics , Adult , Aged , CD28 Antigens/immunology , CD3 Complex/immunology , Cell Line, Tumor , Female , Flow Cytometry , Humans , Interferon-gamma/biosynthesis , Leukocytes, Mononuclear/immunology , Male , Middle Aged , Placebos , RNA, Messenger/biosynthesis , Receptors, Antigen, T-Cell/immunology , Receptors, Calcitriol/genetics , Receptors, Calcitriol/metabolism , Retinoid X Receptor alpha/biosynthesis , Retinoid X Receptor alpha/genetics , Young Adult
8.
Scand J Immunol ; 80(6): 417-23, 2014 Dec.
Article in English | MEDLINE | ID: mdl-25346048

ABSTRACT

Activated macrophages shed the haemoglobin-haptoglobin scavenger receptor CD163 into the circulation as soluble(s)-CD163. We measured sCD163 as an in vivo macrophage activation marker in patients with Crohn's disease (CD) or ulcerative colitis (UC) receiving antitumour necrosis factor (TNF)-α antibody or prednisolone treatment. We also investigated the CD163 expression on circulating monocytes. 58 patients with CD, 40 patients with UC and 90 healthy controls (HC) were included. All patients had active disease at inclusion and were followed for 6 weeks of anti-TNF-α antibody or prednisolone treatment. We measured plasma sCD163 levels at baseline, 1 day, 1 week and 6 weeks after initiating treatment. CD163 expression on circulating CD14(+) monocytes was measured in 21 patients with CD receiving anti-TNF-α antibody treatment. Baseline sCD163 levels were elevated in patients with CD [1.99 (1.80-2.18) mg/l] and in patients with UC [2.07 (1.82-2.32) mg/l] compared with HC [1.51 (1.38-1.63) mg/l] (P < 0.001). Anti-TNF-α antibody treatment induced a rapid decrease in sCD163 levels in patients with CD and in patients with UC 1 day after treatment initiation (P < 0.05). One week of prednisolone treatment did not induce a reduction in sCD163 levels. Anti-TNF-α treatment normalized sCD163 levels in patients with UC, whereas patients with CD exhibited sustained increased sCD163 levels. In patients with CD, CD163 expression on CD14(+) monocytes was increased compared with HC. This study highlights that active CD and UC are associated with increased macrophage activation, as indicated by elevated sCD163 levels and monocytic CD163 expression. Anti-TNF-α antibody treatment induced a rapid decrease in sCD163 levels, suggesting a specific effect on macrophage activation in inflammatory bowel diseases.


Subject(s)
Antibodies, Monoclonal/therapeutic use , Antigens, CD/blood , Antigens, Differentiation, Myelomonocytic/blood , Inflammatory Bowel Diseases/drug therapy , Inflammatory Bowel Diseases/metabolism , Receptors, Cell Surface/blood , Tumor Necrosis Factor-alpha/antagonists & inhibitors , Antibodies, Monoclonal/pharmacology , Antigens, CD/metabolism , Antigens, Differentiation, Myelomonocytic/metabolism , Biomarkers/blood , Case-Control Studies , Colitis, Ulcerative/drug therapy , Colitis, Ulcerative/immunology , Colitis, Ulcerative/metabolism , Crohn Disease/drug therapy , Crohn Disease/immunology , Crohn Disease/metabolism , Humans , Inflammatory Bowel Diseases/immunology , Lipopolysaccharide Receptors/metabolism , Macrophage Activation/immunology , Macrophages/drug effects , Macrophages/immunology , Macrophages/metabolism , Monocytes/immunology , Monocytes/metabolism , Receptors, Cell Surface/metabolism , Steroids/pharmacology , Steroids/therapeutic use
9.
Aliment Pharmacol Ther ; 32(11-12): 1364-72, 2010 Dec.
Article in English | MEDLINE | ID: mdl-21050239

ABSTRACT

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 < 0.02, Wilcoxon rank sum test). Interestingly, vitamin D3 increased the amount of proliferating stimulated CD4+ T cells from median 41% (range: 10-75%) to 56% (range: 26-77%) (P = 0.02, Wilcoxon rank sum test). CONCLUSIONS: Vitamin D3 treatment of Crohn's disease patients increased the IL-6 levels. Interestingly, vitamin D3 treatment enhanced the CD4+ T cell proliferation.


Subject(s)
Cholecalciferol/therapeutic use , Crohn Disease/drug therapy , Interleukin-6/immunology , T-Lymphocytes/immunology , Adult , Aged , Crohn Disease/immunology , Drug Administration Schedule , Female , Humans , Male , Middle Aged , Statistics as Topic , T-Lymphocytes/drug effects , Young Adult
10.
Aliment Pharmacol Ther ; 32(3): 377-83, 2010 Aug.
Article in English | MEDLINE | ID: mdl-20491740

ABSTRACT

BACKGROUND: Vitamin D has immune-regulatory functions in experimental colitis, and low vitamin D levels are present in Crohn's disease. AIM: To assess the effectiveness of vitamin D3 treatment in Crohn's disease with regard to improved disease course. METHODS: We performed a randomized double-blind placebo-controlled trial to assess the benefits of oral vitamin D3 treatment in Crohn's disease. We included 108 patients with Crohn's disease in remission, of which fourteen were excluded later. Patients were randomized to receive either 1200 IU vitamin D3 (n = 46) or placebo (n = 48) once daily during 12 months. The primary endpoint was clinical relapse. RESULTS: Oral vitamin D3 treatment with 1200 IU daily increased serum 25OHD from mean 69 nmol/L [standard deviation (s.d.) 31 nmol/L] to mean 96 nmol/L (s.d. 27 nmol/L) after 3 months (P < 0.001). The relapse rate was lower among patients treated with vitamin D3 (6/46 or 13%) than among patients treated with placebo (14/48 or 29%), (P = 0.06). CONCLUSIONS: Oral supplementation with 1200 IE vitamin D3 significantly increased serum vitamin D levels and insignificantly reduced the risk of relapse from 29% to 13%, (P = 0.06). Given that vitamin D3 treatment might be effective in Crohn's disease, we suggest larger studies to elucidate this matter further. ClinicalTrial.gov(NCT00122184).


Subject(s)
Cholecalciferol/therapeutic use , Crohn Disease/drug therapy , Vitamins/therapeutic use , Double-Blind Method , Humans , Recurrence , Treatment Outcome
11.
J Dairy Sci ; 92(11): 5378-85, 2009 Nov.
Article in English | MEDLINE | ID: mdl-19841198

ABSTRACT

Osteopontin (OPN) is a multifunctional bioactive protein that is implicated in numerous biological processes such as bone remodeling, inhibition of ectopic calcification, and cellular adhesion and migration, as well as several immune functions. Osteopontin has cytokine-like properties and is a key factor in the initiation of T helper 1 immune responses. Osteopontin is present in most tissues and body fluids, with the highest concentrations being found in milk. In the present study, ELISA for human and bovine milk OPN were developed and OPN concentration in human breast milk, bovine milk, and infant formulas was measured and compared. The OPN concentration in human milk was measured to approximately 138 mg/L, which corresponds to 2.1% (wt/wt) of the total protein in human breast milk. This is considerably higher than the corresponding OPN concentrations in bovine milk (approximately 18 mg/L) and infant formulas (approximately 9 mg/L). Moreover, bovine milk OPN is shown to induce the expression of the T helper 1 cytokine IL-12 in cultured human lamina propria mononuclear cells isolated from intestinal biopsies. Finally, the OPN concentration in plasma samples from umbilical cords, 3-mo-old infants, and pregnant and nonpregnant adults was measured. The OPN level in plasma from 3-mo-old infants and umbilical cords was found to be 7 to 10 times higher than in adults. Thus, the high levels of OPN in milk and infant plasma suggest that OPN is important to infants and that ingested milk OPN is likely to induce cytokine production in neonate intestinal immune cells.


Subject(s)
Infant Formula/chemistry , Milk, Human/chemistry , Milk/chemistry , Osteopontin/analysis , Adult , Animals , Cattle , Cells, Cultured , Enzyme-Linked Immunosorbent Assay , Female , Gene Expression Regulation/drug effects , Humans , Infant , Mucous Membrane/cytology , Mucous Membrane/drug effects , Osteopontin/blood , Osteopontin/pharmacology , Pregnancy , Young Adult
12.
Clin Exp Immunol ; 155(3): 487-95, 2009 Mar.
Article in English | MEDLINE | ID: mdl-19094116

ABSTRACT

Pattern recognition receptors (PRRs) are an integral part of the innate immune system and govern the early control of foreign microorganisms. Single nucleotide polymorphisms (SNPs) in the intracellular pattern recognition receptor nucleotide-binding oligomerization domain-containing protein (NOD2, nucleotide oligomerization domain 2) are associated with Crohn's disease (CD). We investigated the impact of NOD2 polymorphisms on cytokine secretion and proliferation of peripheral blood mononuclear cells (PBMCs) in response to Toll-like receptor (TLR) and NOD2 ligands. Based on NOD2 SNP analyses, 41 CD patients and 12 healthy controls were studied. PBMCs were stimulated with NOD2 and TLR ligands. After 18 h culture supernatants were measured using multiplex assays for the presence of human cytokines granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin (IL)-1 beta and tumour necrosis factor (TNF)-alpha. In CD patients, TLR-induced GM-CSF secretion was impaired by both NOD2-dependent and -independent mechanisms. Moreover, TNF-alpha production was induced by a TLR-2 ligand, but a down-regulatory function by the NOD2 ligand, muramyl dipeptide, was impaired significantly in CD patients. Intracellular TLR ligands had minimal effect on GM-CSF, TNF-alpha and IL-1beta secretion. CD patients with NOD2 mutations were able to secrete TNF-alpha, but not GM-CSF, upon stimulation with NOD2 and TLR-7 ligands. CD patients have impaired GM-CSF secretion via NOD2-dependent and -independent pathways and display an impaired NOD2-dependent down-regulation of TNF-alpha secretion. The defect in GM-CSF secretion suggests a hitherto unknown role of NOD2 in the pathogenesis of CD and is consistent with the hypothesis that impaired GM-CSF secretion in part constitutes a NOD2-dependent disease risk factor.


Subject(s)
Crohn Disease/metabolism , Granulocyte-Macrophage Colony-Stimulating Factor/blood , Nod2 Signaling Adaptor Protein/metabolism , Signal Transduction/physiology , Toll-Like Receptors/metabolism , Acetylmuramyl-Alanyl-Isoglutamine/pharmacology , Case-Control Studies , Cell Proliferation , Cells, Cultured , Crohn Disease/pathology , Crohn Disease/physiopathology , Humans , Interleukin-1beta/blood , Ligands , Lipopolysaccharides/pharmacology , Nod2 Signaling Adaptor Protein/genetics , Polymorphism, Single Nucleotide , Statistics, Nonparametric , Tumor Necrosis Factor-alpha/blood
13.
Scand J Immunol ; 65(5): 453-60, 2007 May.
Article in English | MEDLINE | ID: mdl-17444956

ABSTRACT

In Crohn's disease (CD) mucosal T-cells produce increased interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha) levels and TNF-alpha antibody treatment [Infliximab (Ifx)] is effective. Osteopontin (OPN), a glycoprotein stimulating activated T-lymphocytes, may be involved in the disturbed immune-regulation but also in normal immune-homeostasis and mucosal repair, since it is expressed in many tissues and present in human milk. This study investigates plasma-OPN levels in CD patients during Ifx treatment and the in vitro effect of OPN on intestinal T cells. Thirty-seven CD patients received three Ifx doses at week 0, 2 and 6. Blood samples, colonic biopsies and clinical scores were obtained before treatment and at week 8, 26 and 52. In-vivo activated T-cell cultures were established from colonic biopsies in the presence of interleukin (IL)-2 and IL-4. The in vitro effect of OPN stimulation on T-cell IFN-gamma, TNF-alpha, and IL-10 production was measured. Plasma-OPN was increased in active CD (increased CRP-level) compared with quiescent disease (P = 0.02) and declined after three Ifx doses (P = 0.04). It was inversely correlated with in vitro T-cell IL-10 production. OPN increased CD69 and CD25 expression and enhanced T-cell IFN-gamma and TNF-alpha production in a dose-dependent fashion with higher levels in CD than in healthy controls (HC), but induced a concomitant higher IL-10 production in HC than CD. In conclusion, plasma-OPN levels are related to CD inflammation. In vitro, OPN-stimulated IL-10 production increases less in T-cell cultures from CD patients than from HC, indicating that IL-10 deficiency may be involved in the defect immune-regulation in CD, even after OPN stimulation.


Subject(s)
Crohn Disease/metabolism , Crohn Disease/pathology , Cytokines/physiology , Inflammation Mediators/physiology , Osteopontin/physiology , Adolescent , Adult , Aged , Antibodies, Monoclonal/therapeutic use , Cells, Cultured , Crohn Disease/immunology , Crohn Disease/therapy , Cytokines/blood , Female , Humans , Infliximab , Interleukin-10/biosynthesis , Interleukin-10/deficiency , Interleukin-10/metabolism , Male , Middle Aged , Osteopontin/blood , T-Lymphocytes/metabolism
14.
Clin Exp Immunol ; 141(3): 549-57, 2005 Sep.
Article in English | MEDLINE | ID: mdl-16045746

ABSTRACT

Summary CD4(+)CD25(+) regulatory T cells (T(regs)) are involved in the maintenance of peripheral tolerance and ensure a balanced immune response competent of fighting pathogens and at the same time recognizing commensals as harmless. This feature is lost in Crohn's disease (CD). The forkhead/winged helix transcription factor FoxP3 is a master gene for T(reg) function and defects in the FoxP3 gene lead to a clinical picture similar to inflammatory bowel disease (IBD). Murine colitis can be cured by adoptive transfer of T(regs) and ex vivo-generated gut-specific T(regs) represent an attractive option for therapy in CD. Thus, defective T(regs) could contribute to the development of CD. We cultured biopsies of colonic mucosa in the presence of high concentrations of interleukin (IL)-2 and IL-4 to overcome the anergic nature of naturally occurring CD4(+)CD25(+) T(regs) in the mucosa. We investigated the expression of FoxP3 and regulatory potential of gut-derived CD4(+)CD25(+) T cells cultured from patients with CD and healthy individuals. The FoxP3 expression was analysed by reverse transcriptase polymerase chain reaction (RT-PCR), and the suppressive effect of FoxP3(+)CD4(+)CD25(+) T cells on proliferation and cytokine production of autologous CD4(+) T cells was assessed by flow cytometry. Cultured gut-derived T cells with CD4(+)CD25(+) phenotype expressed FoxP3 and were able as the freshly isolated T(regs) from peripheral blood to suppress proliferation and cytokine production of autologous CD4(+) T cells. Thus, we demonstrate that FoxP3(+)CD4(+)CD25(+) T cells with regulatory properties can be propagated in vitro from inflamed mucosa of CD patients, which may be of interest in adoptive immunotherapy.


Subject(s)
CD4-Positive T-Lymphocytes/immunology , Colon , Crohn Disease/immunology , DNA-Binding Proteins/analysis , Intestinal Mucosa/immunology , Receptors, Interleukin-2/analysis , Adult , CD4-Positive T-Lymphocytes/metabolism , Case-Control Studies , Cells, Cultured , DNA-Binding Proteins/genetics , Down-Regulation , Female , Flow Cytometry , Forkhead Transcription Factors , Humans , Interferon-gamma/immunology , Interleukin-2/immunology , Interleukin-4/immunology , Male , RNA, Messenger/analysis , Reverse Transcriptase Polymerase Chain Reaction , Tumor Necrosis Factor-alpha/immunology
16.
Clin Exp Immunol ; 138(1): 66-74, 2004 Oct.
Article in English | MEDLINE | ID: mdl-15373907

ABSTRACT

Integrin alpha4beta 7 is the principal gut-homing receptor, and it is assumed that expression of this specific integrin directs lymphocytes to the gut in vivo. Adoptive cellular immunotherapy against inflammatory bowel disease (IBD) may depend on the expression of integrin alpha4beta 7 to accomplish local delivery of intravenously injected regulatory T cells in inflamed gut mucosa. The present study aimed to investigate whether in vitro expanded human T cells from the colonic mucosa maintain integrin expression, show in vitro adhesion and retain in vivo gut-homing properties during cultivation. Whole colonic biopsies from healthy subjects were cultured in the presence of interleukin-2 (IL-2) and IL-4. The integrin expression of the cultured T cells was determined by flow cytometry and in vitro adhesion was assessed in a mucosal addressin cell adhesion molecule 1 (MAdCAM-1) adhesion assay. We studied the homing pattern after autologous infusion of 3 x 10(8 111)Indium ((111)In)-labelled T cells in five healthy subjects using scintigraphic imaging. The cultured CD4(+)CD45RO(+) gut-derived T cells express higher levels of integrin alpha4beta 7 than peripheral blood lymphocytes (PBLs) and show strong adhesion to MAdCAM-1 in vitro, even after (111)In-labelling. Scintigraphic imaging, however, showed no gut-homing in vivo. After prolonged transit through the lungs, the T cells migrated preferentially to the spleen, liver and bone marrow. In conclusion, it is feasible to infuse autologous T cells cultured from the gut mucosa, which may be of interest in adoptive immunotherapy. Despite high expression of the gut-homing integrin alpha4beta 7 and adhesion to MAdCAM-1 in vitro, evaluation by (111)In-scintigraphy demonstrated no gut-homing in healthy individuals.


Subject(s)
Colon/immunology , Immunoglobulins/immunology , Integrins/analysis , Intestinal Mucosa/immunology , Mucoproteins/immunology , T-Lymphocytes/immunology , Adult , Antigens, CD/immunology , Bone Marrow/immunology , Cell Adhesion/immunology , Cell Adhesion Molecules , Cell Movement/immunology , Cells, Cultured , Female , Humans , Indium Radioisotopes , Integrins/immunology , Interleukin-2/immunology , Interleukin-4/immunology , Liver/immunology , Lung/immunology , Male , Middle Aged , Receptors, Lymphocyte Homing/immunology , Spleen/immunology
17.
Scand J Gastroenterol ; 39(3): 238-45, 2004 Mar.
Article in English | MEDLINE | ID: mdl-15074393

ABSTRACT

BACKGROUND: Characterization of the T-cell receptor variable beta chain (Vbeta) repertoire in inflamed mucosa has been used to identify disease-relevant T-cell populations and antigens in Crohn disease (CD). In vitro expansion of mucosal T cells may reveal changes in Vbeta repertoire not apparent in fresh isolates and we aimed to identify Vbeta subpopulations implicated in Crohn disease. METHODS: In vivo activated mucosal T cells were cultivated using IL-2 and IL-4 from biopsies of whole colonic mucosa without use of Vbeta-modifying exogenous antigen or feeder cells. The Vbeta gene expression in mucosal T-cell cultures was determined in 30 patients with CD and 12 healthy controls using reverse transcriptase polymerase reaction (RT-PCR) covering all 23 functional Vbeta families and the Vbeta receptor prevalence was evaluated by flow cytometry in selected cultures. RESULTS: Early T-cell cultures from both CD patients and healthy controls showed a polyclonal Vbeta gene expression that narrowed during culture, which in CD cultures led to a significant over-expression of the Vbeta5.1 (P = 0.04) and Vbeta8 gene segments (P = 0.03). Together with Vbeta6 and Vbeta18, these Vbeta chains form a pattern of staphylococcal enterotoxin type E (SEE) responsive Vbeta chains, also over-expressed in CD cultures (P = 0.02). Further in vitro stimulation of CD cultures with SEE caused expansion of Vbeta8 receptor positive cells together with a proinflammatory cytokine response. CONCLUSIONS: CD may be associated with (super)antigen-specific Vbeta subpopulations selected during long-term cultivation of mucosal biopsies from inflamed colon.


Subject(s)
Crohn Disease/genetics , Gene Expression , Genes, T-Cell Receptor beta , Receptors, Antigen, T-Cell, alpha-beta/genetics , T-Lymphocytes/physiology , Adolescent , Adult , Case-Control Studies , Cell Culture Techniques , Colon/pathology , Crohn Disease/pathology , Female , Humans , Intestinal Mucosa/pathology , Male , Middle Aged , Reverse Transcriptase Polymerase Chain Reaction
18.
Scand J Gastroenterol ; 38(10): 1050-4, 2003 Oct.
Article in English | MEDLINE | ID: mdl-14621279

ABSTRACT

BACKGROUND: Crohn disease is characterized by up-regulated intestinal inflammation mainly caused by increased tumour necrosis factor alpha (TNF-alpha) levels. However, the complement system (C) may also have a role in maintaining inflammation. METHODS: Plasma from 26 patients with Crohn disease complicated by fistulizing ano-rectal disease was collected before and after three Infliximab infusions (5 mg kg(-1)). RESULTS: Before treatment, the C3-activation capacities (C3-AC) in plasma from patients with Crohn disease were comparable with values obtained from healthy controls. The classical C pathway-mediated C3-AC, mannan-binding lectin C4-AC, leucocyte count, C-reactive protein concentration and Crohn Disease Activity Index decreased significantly 8 weeks after the first infusion of Infliximab (P < 0.04, Wilcoxon test). CONCLUSIONS: Before treatment, all three C pathways were within the normal range in plasma from patients with Crohn disease; the decrease observed in the classical pathway-mediated C3-AC after treatment with Infliximab reflects a general down-regulation in immune activation.


Subject(s)
Antibodies, Monoclonal/therapeutic use , Complement Activation/immunology , Crohn Disease/drug therapy , Crohn Disease/immunology , Gastrointestinal Agents/therapeutic use , Adolescent , Adult , C-Reactive Protein/analysis , Complement C3/immunology , Complement C4/immunology , Complement Pathway, Classical , Down-Regulation , Female , Humans , Infliximab , Leukocyte Count , Male , Middle Aged
19.
Aliment Pharmacol Ther ; 17(5): 703-10, 2003 Mar 01.
Article in English | MEDLINE | ID: mdl-12641520

ABSTRACT

BACKGROUND: Infliximab reduces mucosal inflammation in some, but not all, patients with Crohn's disease. AIM: To monitor clinical data and changes in mucosal cytokine levels after infliximab treatment to identify differences between responders and non-responders. METHODS: Twenty-six patients with fistulating Crohn's disease received three infliximab infusions at weeks 0, 2 and 6. Follow-up was for 1 year and included clinical examination, colonoscopy, ano-rectal ultrasound and magnetic resonance imaging. Biopsies were taken at weeks 0, 8, 26 and 52. Cell cultures were established and analysed for tumour necrosis factor-alpha, interferon-gamma and interleukin-10 levels, and related to clinical status and fistula healing. RESULTS: Eleven of 15 patients (73%) with active disease (Crohn's disease activity index > 150) obtained remission (Crohn's disease activity index < 150) at 8 weeks. In in vitro cell cultures, there was reduced tumour necrosis factor-alpha and interleukin-10 production at week 26, with the latter persistent throughout the study period. When the disease deteriorated or relapsed, there was increased interferon-gamma production in in vitro cell cultures. Fistula healing was associated with reduced production of interferon-gamma, tumour necrosis factor-alpha and interleukin-10. CONCLUSIONS: Infliximab down-regulates mucosal immune activation in Crohn's disease. Monitoring of mucosal cytokine levels after infliximab treatment by whole biopsy cultures may be useful as interleukin-10, tumour necrosis factor-alpha and interferon-gamma production are different in responders and at relapse.


Subject(s)
Antibodies, Monoclonal/therapeutic use , Crohn Disease/drug therapy , Gastrointestinal Agents/therapeutic use , Intestinal Fistula/complications , Rectal Diseases/complications , Adolescent , Adult , Aged , Anus Diseases/complications , Anus Diseases/metabolism , Anus Diseases/pathology , Cells, Cultured , Crohn Disease/metabolism , Crohn Disease/pathology , Cytokines/metabolism , Female , Humans , Infliximab , Intestinal Fistula/metabolism , Intestinal Fistula/pathology , Intestinal Mucosa/immunology , Male , Middle Aged , Rectal Diseases/metabolism , Rectal Diseases/pathology , Recurrence
20.
Scand J Immunol ; 55(1): 105-10, 2002 Jan.
Article in English | MEDLINE | ID: mdl-11841698

ABSTRACT

We have developed a method for quantitating mannan-binding lectin (MBL)-induced activation of the complement system (MBL-C4-AC) in human plasma. This method and an assay for MBL concentration were applied to plasma samples from healthy individuals and patients with systemic lupus erythematosus (SLE), Crohn's disease (CD) and colorectal cancer (CRC). The MBL concentration was measured by an enzyme-linked immunosorbent assay (ELISA) using monoclonal anti-MBL-antibodies and MBL-C4-AC by an ELISA using solid-phase mannan, incubating with plasma samples and quantitating the complement (C) activation by the use of antibodies against the C split-products C4b/C4c. The MBL concentration was nonsignificantly elevated in plasma from SLE-patients, whereas MBL-C4-AC was suppressed (P < 0.04). There was no correlation between MBL concentration and MBL-C4-AC in plasma from SLE-patients. In contrast, a significant correlation was found between the MBL concentration and MBL-C4-AC in plasma from healthy individuals. The C4 concentration was significantly reduced (P < 0.002) in plasma from the SLE patients and showed a significant correlation to MBL-C4-AC. The MBL-C4-AC assay was highly effective in discriminating the SLE patients from the other patient groups and healthy individuals.


Subject(s)
Carrier Proteins/immunology , Colorectal Neoplasms/immunology , Complement Activation , Crohn Disease/immunology , Lupus Erythematosus, Systemic/immunology , Adult , Aged , Carrier Proteins/blood , Case-Control Studies , Collectins , Colorectal Neoplasms/blood , Crohn Disease/blood , Female , Humans , Lupus Erythematosus, Systemic/blood , Male , Middle Aged
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