AIM: To investigate whether TL-10-transduced dendritic cells (DCs) could induce tolerogenicity and prolong allograft survival in rat intestinal transplantation.METHODS: Spleen-derived DCs were prepared and genetically...AIM: To investigate whether TL-10-transduced dendritic cells (DCs) could induce tolerogenicity and prolong allograft survival in rat intestinal transplantation.METHODS: Spleen-derived DCs were prepared and genetically modified by hTL-10 gene. The level of IL-10 expression was quantitated by ELTSA. DC function was assessed by MTT in mixed leukocyte reaction. Allogeneic T-cell apoptosis was examined by flow cytometric analysis. Seven days before heterotopic intestinal transplantation, 2x106 donor-derived IL-10-DC were injected intravenously, then transplantation was performed between SD donor and Wistar recipient.RESULTS: Compared with untransduced DC, IL-10-DC could suppress allogeneic mixed leukocyte reaction (MLR). The inhibitory effect was the most striking with the stimulator/effector (S/F) ratio of 1:10. The inhibition rate was 33.25 %,41.19 % (P<0.01) and 22.92 % with the S/E ratio of 1:1,1:10 and 1:50 respectively. At 48 hours and 72 hours by flow cytometry counting, apoptotic T cells responded to IL-10-DC in MLR were 13.8 % and 30.1%, while untransduced group did not undergo significant apoptosis (P<0.05). IL-10-DC pretreated recipients had a moderate survival prolongation with a mean allograft survival of 19.8 days (P<0.01),compared with 7.3±2.4 days in control group and 8.3±2.9days in untransduced DC group. Rejection occurred in the control group within three days. The difference between untreated DC group and control group was not significant.CONCLUSION: IL-10-DC can induce allogenic T-cell hyporesponsiveness in vitro and apoptosis may be involved in it. IL-10-DC pretreatment can prolong intestinal allograft survival in the recipient.展开更多
To establish a cell line for stable expression of human beta-defensin 3 (hBD3). Methods Full length cDNA of hBD3 was isolated from previously constructed pGEM-hBD3 and then inserted into pcDNA3. The recombinant vector...To establish a cell line for stable expression of human beta-defensin 3 (hBD3). Methods Full length cDNA of hBD3 was isolated from previously constructed pGEM-hBD3 and then inserted into pcDNA3. The recombinant vector identified carrying hBD3 with right direction was introduced into COS-7 cells by Lipofe-ctamine. Cell clones survived in G418-rich medium and with stable expression of hBD3 in both mRNA and protein levels were identified by RT-PCR and Western blot respectively. Genomic integration of the hBD3 gene with the COS-7 cells was confirmed by Southern dot blot and primary analysis. The antimicrobial activity of the secreted hBD3 was also evaluated. Results COS-7 cells transfected with pcDNA3-hBD3 expressed hBD3 stably in mRNA and protein level. Southern dot blot analysis showed successful integration of the hBD3 gene into the genome of COS-7 cell and the hBD-3 protein secreted into the culture medium showed antimicrobial activity. Conclusion We successfully established a hBD3-expressing cell line.展开更多
Acute lung injury (ALI) is a leading complication in extensively burnedpatients, especially those with inhalation injury. It can cause hypoxia resulting in injury ofremote organs and dysfunction. p38 mitogen-activated...Acute lung injury (ALI) is a leading complication in extensively burnedpatients, especially those with inhalation injury. It can cause hypoxia resulting in injury ofremote organs and dysfunction. p38 mitogen-activated protein kinase (p38 MARK) is a stress activatedprotein kinase in the MARK family. Most of the previous studies have demonstrated that p38 MARKsignal transduction pathway mediated All in rats with acute severe pancreatitis, sepsis etc.However, there is little information regarding the role of p38 MARK signal transduction pathway inALI after severe burn trauma.展开更多
文摘AIM: To investigate whether TL-10-transduced dendritic cells (DCs) could induce tolerogenicity and prolong allograft survival in rat intestinal transplantation.METHODS: Spleen-derived DCs were prepared and genetically modified by hTL-10 gene. The level of IL-10 expression was quantitated by ELTSA. DC function was assessed by MTT in mixed leukocyte reaction. Allogeneic T-cell apoptosis was examined by flow cytometric analysis. Seven days before heterotopic intestinal transplantation, 2x106 donor-derived IL-10-DC were injected intravenously, then transplantation was performed between SD donor and Wistar recipient.RESULTS: Compared with untransduced DC, IL-10-DC could suppress allogeneic mixed leukocyte reaction (MLR). The inhibitory effect was the most striking with the stimulator/effector (S/F) ratio of 1:10. The inhibition rate was 33.25 %,41.19 % (P<0.01) and 22.92 % with the S/E ratio of 1:1,1:10 and 1:50 respectively. At 48 hours and 72 hours by flow cytometry counting, apoptotic T cells responded to IL-10-DC in MLR were 13.8 % and 30.1%, while untransduced group did not undergo significant apoptosis (P<0.05). IL-10-DC pretreated recipients had a moderate survival prolongation with a mean allograft survival of 19.8 days (P<0.01),compared with 7.3±2.4 days in control group and 8.3±2.9days in untransduced DC group. Rejection occurred in the control group within three days. The difference between untreated DC group and control group was not significant.CONCLUSION: IL-10-DC can induce allogenic T-cell hyporesponsiveness in vitro and apoptosis may be involved in it. IL-10-DC pretreatment can prolong intestinal allograft survival in the recipient.
文摘To establish a cell line for stable expression of human beta-defensin 3 (hBD3). Methods Full length cDNA of hBD3 was isolated from previously constructed pGEM-hBD3 and then inserted into pcDNA3. The recombinant vector identified carrying hBD3 with right direction was introduced into COS-7 cells by Lipofe-ctamine. Cell clones survived in G418-rich medium and with stable expression of hBD3 in both mRNA and protein levels were identified by RT-PCR and Western blot respectively. Genomic integration of the hBD3 gene with the COS-7 cells was confirmed by Southern dot blot and primary analysis. The antimicrobial activity of the secreted hBD3 was also evaluated. Results COS-7 cells transfected with pcDNA3-hBD3 expressed hBD3 stably in mRNA and protein level. Southern dot blot analysis showed successful integration of the hBD3 gene into the genome of COS-7 cell and the hBD-3 protein secreted into the culture medium showed antimicrobial activity. Conclusion We successfully established a hBD3-expressing cell line.
文摘Acute lung injury (ALI) is a leading complication in extensively burnedpatients, especially those with inhalation injury. It can cause hypoxia resulting in injury ofremote organs and dysfunction. p38 mitogen-activated protein kinase (p38 MARK) is a stress activatedprotein kinase in the MARK family. Most of the previous studies have demonstrated that p38 MARKsignal transduction pathway mediated All in rats with acute severe pancreatitis, sepsis etc.However, there is little information regarding the role of p38 MARK signal transduction pathway inALI after severe burn trauma.
