- Email: [email protected]
S30 Th17 cells mediate intestinal fibrosis in mice
30
5th International Meeting on Inflammatory Bowel Diseases, Capri, April 8 10, 2010
S29 Therapeutic effect of adipose-derived mesenchymal stem cells on experimental colitis by inhibiting inflammatory and autoimmune responses E. Lombardo, D. B¨ uscher. R&D Department, Cellerix S.A., Tres Cantos, Spain Background and Aims: Crohn’s disease is a chronic debilitating disease characterized by severe T helper cell (Th)1-driven inflammation of the colon partially caused by a loss of immune tolerance against mucosal antigens. Mesenchymal stem cells (MSCs) of allogeneic origin have been reported to suppress effector T-cell responses in vitro, to have therapeutic effects in some immune disorders and certain capacity to restore immune tolerance. The aim of this work is to investigate the potential anti-inflammatory and therapeutic effect of human adiposederived MSCs (hASCs) in two well-established murine models of inflammatory bowel diseases. Methods: We examined the therapeutic action of hASCs in the colitis-induced by administration of trinitrobenzene sulfonic acid or dextran sodium sulfate, evaluating diverse clinical signs of the disease. We also investigated the mechanisms involved in the potential therapeutic effect of hASCs, such as inflammatory cytokines and chemokines, Th1-type response, and the generation of regulatory T (Tr) cells. Results: Systemic infusion of hASCs significantly ameliorated the clinical and histopathologic severity of colitis, abrogating body weight loss, diarrhea, and inflammation, and increasing survival. The therapeutic effect was associated with downregulation of both inflammatory and Th1-driven autoimmune response, by regulating a wide spectrum of inflammatory mediators directly through activated macrophages, and by generating interleukin-10-secreting Tr cells with suppressive capacity on autoreactive T cells. In addition, hASCs protected from mortality caused by sepsis by downregulating the exacerbated inflammatory response characteristic of this disease. Conclusions: Therefore, these adult MSCs emerge as key regulators of immune tolerance in physiological conditions by inducing the generation/activation of Tr clones in the periphery and as attractive candidates for a cell-based therapy for the treatment of inflammatory and autoimmune disorders. S30 Th17 cells mediate intestinal fibrosis in mice G. Grassl, G. Diehl, E. Ma, Y. Valdez, D.R. Littman, B. Finlay. University of Kiel, Germany Background: Intestinal fibrosis and stricture formation are serious complications of Crohn’s disease, often requiring surgical intervention. We recently developed a model of Salmonella-induced severe and persistent intestinal fibrosis in mice. T cells play a predominant role in mediating intestinal inflammation and Th17 cells are thought to be the primary inducers of inflammation in Crohn’s disease patients. Aim: Th17 cell development is regulated by the transcription factors RORa and RORg. Here we investigate the contribution of Th17 cells to the development of Salmonella-induced intestinal fibrosis using RORa / and RORg / and RORa / g / doubleknockout mice. Material and Methods: Streptomycin-pretreated C57/Bl6, Rag1 / , RORa / , RORg / and RORa / g / mice were orally infected with an attenuated strain of Salmonella enterica serovar Typhimurium. Tissues were analyzed for bacterial colonization and inflammation, while fibrosis was assessed by Masson’s trichrome staining. We assessed induction of fibrogenic and proinflammatory cytokines including TGF-b1,
IGF-1, MCP-1, IFN-g and IL-17 in the ceca of infected mice. Furthermore, cell types present in fibrotic tissues and the presence of type I and type III collagens were assessed by immunohistochemistry. Results: In all mouse strains, infection with Salmonella led to similar levels of chronic intestinal colonization. C57/Bl6, RORa / and RORg / mice displayed excessive transmural inflammation characterized by submucosal edema, destruction of crypt architecture, epithelial ulcerations and hyperplasia of the muscle layers. In contrast, we observed milder histopathology in Rag1 / and RORa / g / mice. Expression of fibrinogenic and pro-inflammatory cytokines was elevated and accompanied by increased numbers of fibroblast and extensive collagen deposition in the submucosa of C57/Bl6, RORa / and RORg / . However, both profibrotic cytokine expression and collagen production were significantly reduced in Rag1 / and RORa / g / mice. Conclusions: These data demonstrate that intestinal inflammation and fibrosis induced by chronic Salmonella infection of the murine gastrointestinal tract is mediated by Th17 cells and requires both RORa and RORg. Session VII
Developments in clinical IBD
S31 Oral presentation Mucosal healing in Crohn’s disease is associated with high infliximab trough levels W. Van Moerkercke2 , G. Compernolle1 , C. Ackaert1 , A. Gils1 , S. Vermeire2 , M. J¨ urgens2 , I. Cleynen2 , G. van Assche2 , P. Rutgeerts2 . 1 Laboratory for Pharmaceutical Biology KULeuven, Leuven, Belgium, 2 University Hospitals Leuven Laboratory of Gastroenterology, Leuven, Belgium Background and Aims: The importance of mucosal healing as a treatment goal in Crohn’s disease has increasingly been recognized. Infliximab (IFX) is a very efficacious treatment and has shown to induce rapid and profound healing. Mucosal healing is associated with deep clinical remission, reduction of hospitalisations and surgical rates. Nevertheless, primary and secondary failure occurs. Secondary loss of response is at least in part explained by immunogenicity leading to lower trough levels of the drug. Little is known about the importance of trough levels in optimizing therapy. We wanted to study whether IFX trough levels are important to achieve mucosal healing and if they are related to the degree of healing. Methods: We studied serial serum samples in 255 patients with Crohn’s disease, all in whom clinical and endoscopic data before and after start of IFX therapy were present. For 232 patients, serum was available at time t0 (before the start of IFX), t1 (2 to 6 weeks after the first infusion) or t2 (at the time of the endoscopy). IFX trough levels were determined by an in-house developed ELISA method, in which 1:150 diluted serum samples were applicated to TNF alpha coated plates. On each plate, an internal standard curve was introduced and a polyclonal HRP-conjugated goat antihuman antibody was used for detection. Endoscopic healing was defined as complete (disappearance of all lesions), partial (clear endoscopic improvement but still ulceration present) or no healing. Data were analysed using non-parametric Kruskal Wallis and Mann Witney test. Results: Complete healing was observed in 39% of patients, partial in 22% and no healing in 39% of patients. Patients who showed (partial or complete) healing had significantly higher IFX trough levels (median 5.00 mg/mL; P25 0.49 P75 9.85) compared to patients without healing (median 0.95 mg/mL; P25 0.35 P75 6.56 respectively; p = 0.006).
5th International Meeting on Inflammatory Bowel Diseases, Capri, April 8 10, 2010
S29 Therapeutic effect of adipose-derived mesenchymal stem cells on experimental colitis by inhibiting inflammatory and autoimmune responses E. Lombardo, D. B¨ uscher. R&D Department, Cellerix S.A., Tres Cantos, Spain Background and Aims: Crohn’s disease is a chronic debilitating disease characterized by severe T helper cell (Th)1-driven inflammation of the colon partially caused by a loss of immune tolerance against mucosal antigens. Mesenchymal stem cells (MSCs) of allogeneic origin have been reported to suppress effector T-cell responses in vitro, to have therapeutic effects in some immune disorders and certain capacity to restore immune tolerance. The aim of this work is to investigate the potential anti-inflammatory and therapeutic effect of human adiposederived MSCs (hASCs) in two well-established murine models of inflammatory bowel diseases. Methods: We examined the therapeutic action of hASCs in the colitis-induced by administration of trinitrobenzene sulfonic acid or dextran sodium sulfate, evaluating diverse clinical signs of the disease. We also investigated the mechanisms involved in the potential therapeutic effect of hASCs, such as inflammatory cytokines and chemokines, Th1-type response, and the generation of regulatory T (Tr) cells. Results: Systemic infusion of hASCs significantly ameliorated the clinical and histopathologic severity of colitis, abrogating body weight loss, diarrhea, and inflammation, and increasing survival. The therapeutic effect was associated with downregulation of both inflammatory and Th1-driven autoimmune response, by regulating a wide spectrum of inflammatory mediators directly through activated macrophages, and by generating interleukin-10-secreting Tr cells with suppressive capacity on autoreactive T cells. In addition, hASCs protected from mortality caused by sepsis by downregulating the exacerbated inflammatory response characteristic of this disease. Conclusions: Therefore, these adult MSCs emerge as key regulators of immune tolerance in physiological conditions by inducing the generation/activation of Tr clones in the periphery and as attractive candidates for a cell-based therapy for the treatment of inflammatory and autoimmune disorders. S30 Th17 cells mediate intestinal fibrosis in mice G. Grassl, G. Diehl, E. Ma, Y. Valdez, D.R. Littman, B. Finlay. University of Kiel, Germany Background: Intestinal fibrosis and stricture formation are serious complications of Crohn’s disease, often requiring surgical intervention. We recently developed a model of Salmonella-induced severe and persistent intestinal fibrosis in mice. T cells play a predominant role in mediating intestinal inflammation and Th17 cells are thought to be the primary inducers of inflammation in Crohn’s disease patients. Aim: Th17 cell development is regulated by the transcription factors RORa and RORg. Here we investigate the contribution of Th17 cells to the development of Salmonella-induced intestinal fibrosis using RORa / and RORg / and RORa / g / doubleknockout mice. Material and Methods: Streptomycin-pretreated C57/Bl6, Rag1 / , RORa / , RORg / and RORa / g / mice were orally infected with an attenuated strain of Salmonella enterica serovar Typhimurium. Tissues were analyzed for bacterial colonization and inflammation, while fibrosis was assessed by Masson’s trichrome staining. We assessed induction of fibrogenic and proinflammatory cytokines including TGF-b1,
IGF-1, MCP-1, IFN-g and IL-17 in the ceca of infected mice. Furthermore, cell types present in fibrotic tissues and the presence of type I and type III collagens were assessed by immunohistochemistry. Results: In all mouse strains, infection with Salmonella led to similar levels of chronic intestinal colonization. C57/Bl6, RORa / and RORg / mice displayed excessive transmural inflammation characterized by submucosal edema, destruction of crypt architecture, epithelial ulcerations and hyperplasia of the muscle layers. In contrast, we observed milder histopathology in Rag1 / and RORa / g / mice. Expression of fibrinogenic and pro-inflammatory cytokines was elevated and accompanied by increased numbers of fibroblast and extensive collagen deposition in the submucosa of C57/Bl6, RORa / and RORg / . However, both profibrotic cytokine expression and collagen production were significantly reduced in Rag1 / and RORa / g / mice. Conclusions: These data demonstrate that intestinal inflammation and fibrosis induced by chronic Salmonella infection of the murine gastrointestinal tract is mediated by Th17 cells and requires both RORa and RORg. Session VII
Developments in clinical IBD
S31 Oral presentation Mucosal healing in Crohn’s disease is associated with high infliximab trough levels W. Van Moerkercke2 , G. Compernolle1 , C. Ackaert1 , A. Gils1 , S. Vermeire2 , M. J¨ urgens2 , I. Cleynen2 , G. van Assche2 , P. Rutgeerts2 . 1 Laboratory for Pharmaceutical Biology KULeuven, Leuven, Belgium, 2 University Hospitals Leuven Laboratory of Gastroenterology, Leuven, Belgium Background and Aims: The importance of mucosal healing as a treatment goal in Crohn’s disease has increasingly been recognized. Infliximab (IFX) is a very efficacious treatment and has shown to induce rapid and profound healing. Mucosal healing is associated with deep clinical remission, reduction of hospitalisations and surgical rates. Nevertheless, primary and secondary failure occurs. Secondary loss of response is at least in part explained by immunogenicity leading to lower trough levels of the drug. Little is known about the importance of trough levels in optimizing therapy. We wanted to study whether IFX trough levels are important to achieve mucosal healing and if they are related to the degree of healing. Methods: We studied serial serum samples in 255 patients with Crohn’s disease, all in whom clinical and endoscopic data before and after start of IFX therapy were present. For 232 patients, serum was available at time t0 (before the start of IFX), t1 (2 to 6 weeks after the first infusion) or t2 (at the time of the endoscopy). IFX trough levels were determined by an in-house developed ELISA method, in which 1:150 diluted serum samples were applicated to TNF alpha coated plates. On each plate, an internal standard curve was introduced and a polyclonal HRP-conjugated goat antihuman antibody was used for detection. Endoscopic healing was defined as complete (disappearance of all lesions), partial (clear endoscopic improvement but still ulceration present) or no healing. Data were analysed using non-parametric Kruskal Wallis and Mann Witney test. Results: Complete healing was observed in 39% of patients, partial in 22% and no healing in 39% of patients. Patients who showed (partial or complete) healing had significantly higher IFX trough levels (median 5.00 mg/mL; P25 0.49 P75 9.85) compared to patients without healing (median 0.95 mg/mL; P25 0.35 P75 6.56 respectively; p = 0.006).