摘要
AIM:To explore the anti-inflammatory potential of adeno-associated virus-mediated delivery of Tregitope 167 in an experimental colitis model.METHODS:The trinitrobenzene sulfonate(TNBS) model of induced colitis was used in Balb/c mice.Subsequently after intravenous adeno-associated virusmediated regulatory T-cell epitopes(Tregitope) delivery,acute colitis was initiated by intra-rectal administration of 1.5 mg TNBS in 40% ethanol followed by a second treatment with TNBS(0.75 mg in 20% ethanol) 8 d later.Control groups included mice not treated with TNBS(healthy control group) and mice treated by TNBS only(diseased group).At the time of sacrifice colon weight,the disease activity index and histology damage score were determined.Immunohistochemical staining of the colonic tissues was performed to asses the cellular infiltrate and the presence of transcription factor forkhead Box-P3(Foxp3).Thymus,mesenteric lymph nodes,liver and spleen tissue were collected and the corresponding lymphocyte populations were further assessed by flow cytometry analysis for the expression of CD4+ T cell and regulatory T cell associated markers.RESULTS:The Tregitope 167 treated mice gained an average of 4% over their initial body weight at the time of sacrifice.In contrast,the mice treated with TNBS alone(no Tregitope) developed colitis,and lost 4% of their initial body weight at the time of sacrifice(P < 0.01).The body weight increase that had been observed in the mice pre-treated with Tregitope 167 was substantiated by a lower disease activity index and a decreased colon weight as compared to the diseased control group(P < 0.01 and P < 0.001,respectively).Immunohistochemical staining of the colonic tissues for CD4+ showed that inflammatory cell infiltrates were present in TNBS treated mice with or without administration with tregitope 167 and that these cellular infiltrates consisted mainly of CD4+ cells.For both TNBS treated groups CD4+ T cell infiltrates were observed in the sub-epithelial layer and the lamina propria.CD4+ T cell infiltrates were also present in the muscularis mucosa layer of the diseased control mice,but were absent in the Tregitope 167 treated group.Numerous Foxp3 positive cells were detected in the lamina propria and sub-epithelium of the colon sections from mice treated with Tregitope 167.Furthermore,the Foxp3 and glycoprotein A repetitions predominant markers were significantly increased in the CD4+ T lymphocyte population in the thymus of the mice pre-treated with adeno-associated virus serotype 5(cytomegalovirus promoter-Tregitope 167),as cytomegalovirus promoter compared to lymphocyte populations in the thymus of diseased and the healthy control mice(P < 0.05 and P < 0.001,respectively).CONCLUSION:This study identifies adeno-associated virus-mediated delivery of regulatory T-cell epitope 167 as a novel anti-inflammatory approach with the capacity to decrease intestinal inflammation and induce longterm remission in inflammatory bowel disease.
AIM：To explore the anti-inflammatory potential of adeno-associated virus-mediated delivery of Tregitope 167 in an experimental colitis model.METHODS：The trinitrobenzene sulfonate（TNBS） model of induced colitis was used in Balb/c mice.Subsequently after intravenous adeno-associated virusmediated regulatory T-cell epitopes（Tregitope） delivery,acute colitis was initiated by intra-rectal administration of 1.5 mg TNBS in 40% ethanol followed by a second treatment with TNBS（0.75 mg in 20% ethanol） 8 d later.Control groups included mice not treated with TNBS（healthy control group） and mice treated by TNBS only（diseased group）.At the time of sacrifice colon weight,the disease activity index and histology damage score were determined.Immunohistochemical staining of the colonic tissues was performed to asses the cellular infiltrate and the presence of transcription factor forkhead Box-P3（Foxp3）.Thymus,mesenteric lymph nodes,liver and spleen tissue were collected and the corresponding lymphocyte populations were further assessed by flow cytometry analysis for the expression of CD4＋ T cell and regulatory T cell associated markers.RESULTS：The Tregitope 167 treated mice gained an average of 4% over their initial body weight at the time of sacrifice.In contrast,the mice treated with TNBS alone（no Tregitope） developed colitis,and lost 4% of their initial body weight at the time of sacrifice（P 〈 0.01）.The body weight increase that had been observed in the mice pre-treated with Tregitope 167 was substantiated by a lower disease activity index and a decreased colon weight as compared to the diseased control group（P 〈 0.01 and P 〈 0.001,respectively）.Immunohistochemical staining of the colonic tissues for CD4＋ showed that inflammatory cell infiltrates were present in TNBS treated mice with or without administration with tregitope 167 and that these cellular infiltrates consisted mainly of CD4＋ cells.For both TNBS treated groups CD4＋ T cell infiltrates were observed in the sub-epithelial layer and the lamina propria.CD4＋ T cell infiltrates were also present in the muscularis mucosa layer of the diseased control mice,but were absent in the Tregitope 167 treated group.Numerous Foxp3 positive cells were detected in the lamina propria and sub-epithelium of the colon sections from mice treated with Tregitope 167.Furthermore,the Foxp3 and glycoprotein A repetitions predominant markers were significantly increased in the CD4＋ T lymphocyte population in the thymus of the mice pre-treated with adeno-associated virus serotype 5（cytomegalovirus promoter-Tregitope 167）,as cytomegalovirus promoter compared to lymphocyte populations in the thymus of diseased and the healthy control mice（P 〈 0.05 and P 〈 0.001,respectively）.CONCLUSION：This study identifies adeno-associated virus-mediated delivery of regulatory T-cell epitope 167 as a novel anti-inflammatory approach with the capacity to decrease intestinal inflammation and induce longterm remission in inflammatory bowel disease.
基金
Supported by Grant from the Broad Medical Research Program of The Broad Foundation,No. IBD-029 5R
