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Autologous bone marrow-derived mesenchymal stromal cells in the treatment of fistulising Crohn's disease

Objective External fistulas represent a disabling manifestation of Crohns disease with a difficult curability and a high relapse rate despite a large therapeutic armamentarium. Stem cell therapy is a novel and promising approach for treatment of chronic inflammatory conditions. We therefore investigated the feasibility, safety and efficacy of serial intrafistular injections of autologous bone marrow-derived mesenchymal stromal cells (MSCs) in the treatment of fistulising Crohns disease. Patients and methods We enrolled 12 consecutive outpatients (eight males, median age 32 years) refractory to or unsuitable for current available therapies. MSCs were isolated from bone marrow and expanded ex vivo to be used for both therapeutic and experimental purposes. Ten patients (two refused) received intrafistular MSC injections (median 4) scheduled every 4 weeks, and were monitored by surgical, MRI and endoscopic evaluation for 12 months afterwards. The feasibility of obtaining at least 50×106 MSCs from each patient, the appearance of adverse events, and the efficacy in terms of fistula healing and reduction of both Crohns disease and perianal disease activity indexes were evaluated. In addition, the percentage of both mucosal and circulating regulatory T cells expressing FoxP3, and the ability of MSCs to influence mucosal T cell apoptosis were investigated. Results MSC expansion was successful in all cases; sustained complete closure (seven cases) or incomplete closure (three cases) of fistula tracks with a parallel reduction of Crohns disease and perianal disease activity indexes (p<0.01 for both), and rectal mucosal healing were induced by treatment without any adverse effects. The percentage of mucosal and circulating regulatory T cells significantly increased during the treatment and remained stable until the end of follow up (p<0.0001 and p<0.01, respectively). Furthermore, MSCs have been proven to affect mucosal T cell apoptotic rate. Conclusions Locally injected MSCs represent a feasible, safe and beneficial therapy in refractory fistulising Crohns disease.

Helicobacter colonization and histopathological profile of chronic gastritis in patients with or without dyspepsia, mucosal erosion and peptic ulcer: A morphological approach to the study of ulcerogenesis in man

Helicobacter pylori colonization and the incidence, severity, activity and topography of gastritis were investigated systematically in antrum and corpus mucosal biopsies of 1177 subjects undergoing endoscopy in the absence of gastric complaints (asymptomatic, 49) or for non-ulcer dyspepsia (NUD; 631 patients, 72 of whom had gastric and/or duodenal erosions), active gastric ulcer (GU, 76 patients), active duodenal ulcer (DU, 138 patients), and healed gastric (HGU, 39 cases) or duodenal ulcer (HDU, 230 cases). In the antrum,H. pylori colonization and the incidence, severity and activity of gastritis increased progressively in the sequence asymptomatic, erosion-free NUD, erosive NUD, healed ulcer and active ulcer. The same trend was observed in the corpus as regardsH. pylori and gastritis incidence, whereas the severity and activity of gastritis were lower in active DU and erosive NUD and higher in active, proximal GU than in the remaining patients. Active DU and erosive NUD showed the highest incidence of nonatrophic gastritis and lowest type-A or AB atrophic gastritis, while active GU had lowest normal mucosa or type-A gastritis and highest type-B atrophic gastritis. In conclusion,H. pylori colonization and gastritis incidence, severity and, especially, activity of the antrum might all contribute to mucosal erosion and ulceration, whereas the same factors, at least in part and with the exception of proximal GU, seem to have a preventive role when affecting corpus mucosa.

Segmental colitis associated with diverticula : A prospective study

OBJECTIVE: Little is known about the clinical features and natural history of segmental colitis associated with diverticula. Our aim was to evaluate the incidence of segmental colitis associated with diverticula in patients undergoing colonoscopy, its clinical picture, and its outcome. METHODS: This was a multicenter, prospective study. Patients with inflammatory bowel disease (IBD)-like lesions limited to colonic segments with diverticula were enrolled. Patients were treated with oral and topical 5-aminosalicylic (5-ASA) until remission was achieved; clinical and endoscopic follow-up was planned at 6 wk and 12 months. RESULTS: A total of 5457 consecutive colonoscopies were recorded at five participating institutions; 20 patients (0.36%) met the endoscopic criteria for segmental colitis associated with diverticula. All had lesions in the left colon, and one also had lesions in the right colon. In six cases, a specific diagnosis was made thereafter. The remaining 14 patients (0.25% of colonoscopies; eight men; age range, 49–80 yr) were in clinical and endoscopic remission at the first follow-up visit. At onset, 13/14 had hematochezia, seven had diarrhea, and five had abdominal pain; only one had weight loss. No subject had fever. In all but one case, blood chemistries were normal. Five patients had had similar symptoms previously. Thirteen of 14 patients were in clinical and endoscopic remission at 12 months. CONCLUSIONS: This endoscopic picture is not an exceptional finding. Hematochezia was the main clinical feature, and no relation with gender, age, or smoking habit was found. Blood chemistries were generally normal and the rectum was spared. The histological features were not diagnostic and most patients did not complain of any abdominal symptoms 12 months after enrolment.

Segmental colitis associated with diverticula: a prospective study. Gruppo di Studio per le Malattie Infiammatorie Intestinali (GSMII).

OBJECTIVE:Little is known about the clinical features and natural history of segmental colitis associated with diverticula. Our aim was to evaluate the incidence of segmental colitis associated with diverticula in patients undergoing colonoscopy, its clinical picture, and its outcome.METHODS:This was a multicenter, prospective study. Patients with inflammatory bowel disease (IBD)-like lesions limited to colonic segments with diverticula were enrolled. Patients were treated with oral and topical 5-aminosalicylic (5-ASA) until remission was achieved; clinical and endoscopic follow-up was planned at 6 wk and 12 months.RESULTS:A total of 5457 consecutive colonoscopies were recorded at five participating institutions; 20 patients (0.36%) met the endoscopic criteria for segmental colitis associated with diverticula. All had lesions in the left colon, and one also had lesions in the right colon. In six cases, a specific diagnosis was made thereafter. The remaining 14 patients (0.25% of colonoscopies; eight men; age range, 49–80 yr) were in clinical and endoscopic remission at the first follow-up visit. At onset, 13/14 had hematochezia, seven had diarrhea, and five had abdominal pain; only one had weight loss. No subject had fever. In all but one case, blood chemistries were normal. Five patients had had similar symptoms previously. Thirteen of 14 patients were in clinical and endoscopic remission at 12 months.CONCLUSIONS:This endoscopic picture is not an exceptional finding. Hematochezia was the main clinical feature, and no relation with gender, age, or smoking habit was found. Blood chemistries were generally normal and the rectum was spared. The histological features were not diagnostic and most patients did not complain of any abdominal symptoms 12 months after enrolment.

The prevalence and the causes of minimal intestinal lesions in patients complaining of symptoms suggestive of enteropathy. A follow-up study.

Aims: Although they are non-specific, minimal intestinal lesions are at the end of the coeliac histological damage spectrum. To investigate whether minimal intestinal lesions in patients without endomysial antibodies are due to coeliac disease, their prevalence, causes and risk of evolving into frank coeliac disease were studied. Methods: From January 2000 to December 2005, 645 duodenal biopsies were performed. In 209 patients, duodenal biopsies were performed independently of endomysial antibody results. Clinical data and HLA-typing of all the patients negative to endomysial antibodies but with minimal mucosal lesions were re-evaluated. Three years later, they were offered to be seen again, and further investigations were proposed. Results: 14 out of 209 patients had minimal mucosal lesions and negative endomysial antibodies. Two patients were lost to follow-up; in 7/12 patients, symptoms and histological lesions were due to a different condition, not related to coeliac disease. In 11/12 patients, HLA-typing made diagnosis of coeliac disease very unlikely. Only one patient was on a gluten-free diet because of gluten-sensitive symptoms and was DQ2+/DQ8+. Conclusions: Minimal duodenal lesions in patients negative to endomysial antibodies are rare and are likely to be due to conditions unrelated to coeliac disease.

Histologic changes in type A chronic atrophic gastritis indicating increased risk of neuroendocrine tumor development: the predictive role of dysplastic and severely hyperplastic enterochromaffin-like cell lesions.

The role of putative preneoplastic enterochromaffin-like cell lesions, either hyperplastic or dysplastic, in the genesis of type 1 enterochromaffin-like cell neuroendocrine tumors associated with type A chronic atrophic gastritis, their actual neoplastic risk, and their precise histogenetic mechanism deserve further clarification by specific histopathologic studies coupled with patient follow-up. A total of 100 patients with severe type A chronic atrophic gastritis, enterochromaffin-like cell hyperplasia, and antral G-cell hyperplasia were endoscopically and histologically followed up for a median of 90.1 months (total of 9118 person-months). Preneoplastic enterochromaffin-like cell lesions and newly developed neuroendocrine tumors were investigated histologically and histochemically, in parallel with enterochromaffin-like cell lesions found in nontumor mucosa of another 32 well-characterized and previously reported type 1 neuroendocrine tumors. Both neuroendocrine and nonneuroendocrine mucosa changes were analyzed and statistically evaluated. During follow-up, 7 of 100 patients developed neuroendocrine tumors: 5 were in a group of 20 cases with previous enterochromaffin-like cell dysplasia and 2 were among 80 cases showing only enterochromaffin-like cell hyperplasia throughout the study (hazard ratio, 20.7; P < .001). The severity of enterochromaffin-like cell hyperplasia at first biopsy, with special reference to linear hyperplasia with 6 chains or more per linear millimeter, also increased the risk of neuroendocrine tumor development during follow-up (hazard ratio, 13.0; P < .001). Enterochromaffin-like cell microinvasive dysplastic lesions arising at the epithelial renewal zone level, in connection with immature proliferating mucous-neck cells, were found to be linked to early intramucosal neuroendocrine tumor histogenesis. Both enterochromaffin-like cell dysplasia and severe hyperplasia indicate increased risk of neuroendocrine tumor development in type A chronic atrophic gastritis with hypergastrinemia/G-cell hyperplasia.

Ex vivo immunosuppressive effects of mesenchymal stem cells on Crohn’s disease mucosal T cells are largely dependent on indoleamine 2,3-dioxygenase activity and cell-cell contact

IntroductionCrohn’s disease (CD) is a disabling chronic enteropathy sustained by a harmful T-cell response toward antigens of the gut microbiota in genetically susceptible subjects. Growing evidence highlights the safety and possible efficacy of mesenchymal stem cells (MSCs) as a new therapeutic tool for this condition. Therefore, we aimed to investigate the effects of bone marrow-derived MSCs on pathogenic T cells with a view to clinical application.MethodsT-cell lines from both inflamed and non-inflamed colonic mucosal specimens of CD patients and from healthy mucosa of control subjects were grown with the antigen muramyl-dipeptide in the absence or presence of donors’ MSCs. The MSC effects were evaluated in terms of T-cell viability, apoptotic rate, proliferative response, immunophenotype, and cytokine profile. The role of the indoleamine 2,3-dioxygenase (IDO) was established by adding a specific inhibitor, the 1-methyl-DL-tryptophan, and by using MSCs transfected with the small interfering RNA (siRNA) targeting IDO. The relevance of cell-cell contact was evaluated by applying transwell membranes.ResultsA significant reduction in both cell viability and proliferative response to muramyl-dipeptide, with simultaneous increase in the apoptotic rate, was found in T cells from both inflamed and non-inflamed CD mucosa when co-cultured with MSCs and was reverted by inhibiting IDO activity and expression. A reduction of the activated CD4+CD25+ subset and increase of the CD3+CD69+ population were also observed when T-cell lines from CD mucosa were co-cultured with MSCs. In parallel, an inhibitory effect was evident on the expression of the pro-inflammatory cytokines tumor necrosis factor-α, interferon-γ, interleukin-17A and -21, whereas that of the transforming growth factor-β and interleukin-6 were increased, and production of the tolerogenic molecule soluble HLA-G was high. These latter effects were almost completely eliminated by blocking the IDO, whose activity was upregulated in MSCs co-cultured with CD T cells. The use of a semipermeable membrane partially inhibited the MSC immunosuppressive effects. Finally, hardly any effects of MSCs were observed when T cells obtained from control subjects were used.ConclusionMSCs exert potent immunomodulant effects on antigen-specific T cells in CD through a complex paracrine and cell-cell contact-mediated action, which may be exploited for widespread therapeutic use.

Mesenchymal Stromal Cell Infusions as Rescue Therapy for Corticosteroid-Refractory Adult Autoimmune Enteropathy

Adult autoimmune enteropathy (AIE) is a rare cause of malabsorption syndrome unresponsive to dietary restriction. Its diagnostic hallmarks are small-bowel villous atrophy and antienterocyte autoantibodies. Therapy is based mainly on nutritional support and immunosuppression. We treated a 61-year-old woman with corticosteroid-refractory AIE and life-threatening malabsorption syndrome with systemic infusions of autologous, bone marrow-derived, mesenchymal stromal cells (MSCs) as rescue therapy. The MSCs were expanded ex vivo following a previously used Good Manufacturing Practice procedure, and 2 intravenous infusions of 1.8 × 10(6) MSCs/kg body weight were administered 2 weeks apart. Analysis of circulating and mucosal regulatory T-and B-cell numbers, and of serum and secretory immunoglobulin levels, was performed before and after treatment. The MSC infusions were safe and effective, leading to disappearance of disease hallmarks and recovery from the life-threatening condition. Increases in mucosal regulatory T-cell numbers and secretory immunoglobulin levels were also observed. The benefit, however, was transient, and a further MSC infusion resulted in the same short efficacy. This case encourages the use of MSCs to treat patients with life-threatening, corticosteroid-refractory AIE and suggests that MSC infusion can attenuate, albeit transiently, the autoimmune attack.

Increase in neuroendocrine cells in the duodenal mucosa of patients with refractory celiac disease.

OBJECTIVES:Several immune-mediated gastrointestinal disorders, including celiac disease (CD), are associated with neuroendocrine cell hyperplasia. However, neuroendocrine cells have never been explored in refractory CD (RCD).METHODS:Serial duodenal sections from 17 patients with RCD (6 type 1 and 11 type 2), 16 uncomplicated CD patients before and after gluten-free diet, 14 patients with potential CD, 27 patients with non-CD villous atrophy, i.e., common variable immunodeficiency (n=12), Whipples disease (n=10) and giardiasis (n=5), and 16 healthy subjects were processed for the immunohistochemical detection of chromogranin A (CgA), serotonin, and somatostatin. Mucosal tryptophan hydroxylase (TpH)-1 and serotonin-selective reuptake transporter (SERT) transcripts were measured by quantitative reverse transcription-PCR. Serum CgA and 24-h urine 5-hydroxyindoleacetic acid (5-HIAA) were assessed. Biopsies from treated CD patients were cultured with serotonin or peptic tryptic digest of gliadin (PT-gliadin), and interferon (IFN)-γ was detected by ELISA in culture supernatants.RESULTS:Epithelial cells positive for CgA and serotonin, but not somatostatin, were significantly increased in RCD. Raised mucosal transcripts of TpH-1, but not SERT, were found in RCD. On biopsies from treated CD patients, serotonin upregulated IFN-γ production at levels comparable to those induced by PT-gliadin. Serum CgA, but not urine 5-HIAA, was increased in RCD. No significant difference was found between RCD type 1 and type 2 in terms of neuroendocrine cells, mucosal TpH-1 transcripts, and serum CgA.CONCLUSIONS:Serotonin-producing neuroendocrine cells are increased in RCD mucosa. IFN-γ upregulation induced by serotonin suggests that this monoamine may have a role in sustaining the local inflammatory response in CD.

Endoscopic evaluation of gastrointestinal tract in patients with hereditary hemorrhagic telangiectasia and correlation with their genotypes.

Purpose:Hereditary hemorrhagic telangiectasia (HHT) is an autosomal-dominant vascular dysplasia characterized by telangiectases and arteriovenous malformations. Three causative genes are known: ENG (HHT-1), ACVRL1 (HHT-2), and SMAD4 (mutated in HHT in association with juvenile polyposis). Gastrointestinal bleeding is the most common symptom after epistaxis. The stomach and the duodenum are the main gastrointestinal sites of telangiectases. Our aim was to explore gastrointestinal tract of consecutive HHT patients to assess distribution, number, size, and type of telangiectases in relation to genotype.Methods:HHT patients underwent gastroduodenoscopy, video capsule endoscopy, and colonoscopy. Molecular analysis of ENG and ACVRL1 was performed to identify the disease-causing mutation.Results:Twenty-two patients (13 men; mean age: 59 ± 9 years) were analyzed: 7 with HHT-1, 13 with HHT-2, and 2 undefined. Gastrointestinal telangiectases were identified as follows: at gastroduodenoscopy in 86% of HHT-1 patients and in 77% of HHT-2 patients, at video capsule endoscopy in all HHT-1 patients and in 84% of HHT-2 patients, and at colonoscopy in 1 patient for each group. HHT-1 showed multiple telangiectases with a higher prevalence, more relevant in the duodenum.Conclusion:Our data demonstrate extensive involvement of the gastrointestinal tract with a more severe association in HHT-1. Gastroduodenoscopy provides significant information on gastrointestinal involvement, and video capsule endoscopy may be added in selected patients. Colonic polyps/adenomas were identified as occasional findings.Genet Med 16 1, 3–10.