目的:研究带有蛋白穿膜结构域的小鼠Foxp3融合蛋白(mouse Foxp3 fusin protein combining with transduc-tion domain,PTD-mFoxp3)对小鼠脾淋巴细胞活化增殖能力和皮肤移植排斥反应的影响。方法:取健康C57BL/6小鼠脾淋巴细胞,在体外经刀豆蛋白A(concanavaline A,ConA)活化,并分别给予环孢素A(cyclosporine,CsA)、不同浓度的PTD-mFoxp3,mFoxp3,PTD-GFP和培养基处理,检测脾淋巴细胞增殖指数。在此基础上进行从Balb/c小鼠到C57BL/6小鼠的皮肤移植试验,将40只C57BL/6受体小鼠随机分为4组,每组10只,分别以PTD-mFoxp3,CsA,0.9%氯化钠注射液(NS),PTD-GFP于手术当天开始连续腹腔注射8天为干预手段。术后第9天各组随机取2只移植小鼠,采集皮片标本,进行组织学检查。其余的8只用于观察移植皮瓣的存活时间。结果:PTD-mFoxp3和CsA相似,与其他处理方法相比明显抑制ConA活化的小鼠脾淋巴细胞增殖指数(P〈0.05)。PTD-mFoxp3组,CsA组移植物的存活时间较各对照组显著延长(P〈0.05);病理检查显示PTD-mFoxp3组和CsA组移植物淋巴细胞浸润明显轻于各对照组。结论:PTD-mFoxp3能抑制ConA活化的T淋巴细胞的增殖,且具有抑制小鼠皮肤移植排斥反应的作用。
Objective: To investigate the potential inhibiting ability to active lymphocytes proliferation and skin allograft rejection of the fusion protein of mouse forkhead box P3 combining with protein transduction domain (PTD-mFoxp3) in mice. Methods: The single-cell suspension of lymphocytes was prepared from mouse spleen. Polyclonal activator Con A was added to active the lymphocytes. Then the single-cell suspension of lymphocytes was incubated with different concentrations of PTD-mFoxp3, PTD-GFP, cyclosporine, and blank control for 48h, respectively. The bioactivity of inhibiting lymphocytes proliferation of each group was detected by incorporating of CCK-8 in vitro. Allogeneic full-thickness grafts from Balb/c mice were transplanted onto the back of C57BL/6 mice. Forty C57BL/6 mice were randomly divided into four groups, with each group having 10 mice and receiving a different treatment including PTD-mFoxp3, cyclos-porine, 0.9% physiological saline injection and PTD-GFP, respectively. On the same day of the skin trans-plantation, the treating regents were injected into C57BL/6 mice abdomen and did continuous injection for another 7 days. Two mice of each group were selected out randomly for histological observation till 9th day after skin transplantation. The survival time of other mice were observed. Results: The bioactivity of PTD- mFoxp3 was similar to eyclosporine, they could inhibit cells proliferation of mouse activated lymphocytes efficiently in vitro( P 〈 0.05, comparing with blank control and PTD-GFP control) , they could prolong the mean survival time (MST) of skin allograft significantly( P 〈 0.05, comparing with blank control and PTD- GFP control). The infiltration of the lymphoeytes in the allografts of PTD-mFoxp3 treated groups and cyclosporine were reduced. Conclusion: The PTD-mFoxp3 can inhibit activated mouse lymphocytes proliferation in vitro and skin allograft rejection in vivo.