WYE-354调控小鼠同种异体皮肤移植抗排斥反应中的mTOR信号传导通路

WYE-354 regulates signal transduction pathway of mammalian target of rapamycin in anti-rejection of mice after skin allograft

目的探讨mTOR激酶ATP竞争抑制剂WYE-354对小鼠同种异体皮肤移植抗排斥反应的作用及机制。方法以C57BL/6为受者,BALB/c为供者建立同种异体皮肤移植模型,C57BL/6小鼠行同种同体皮肤移植为自体移植组。移植-2d给药,共21d,阳性对照组为同种异体皮肤移植,雷帕霉素组口服给药0.2ml、剂量1mg/(kg.d),WYE-354药物组腹腔注射0.2ml:低剂量1mg/(kg.d)、中剂量1.5mg/(kg.d)、高剂量50mg/(kg.d)。术后3、7、11、21d时,应用免疫印记法留样检测每组脾细胞4E-BP1磷酸化水平,11d行皮肤移植物组织病理染色。结果阳性对照组皮肤移植物存活时间明显少于雷帕霉素组、WYE-354低、中、高剂量组,差异有统计学意义(P<0.01);雷帕霉素组皮肤移植物存活时间明显少于WYE-354中、高剂量组,差异有统计学意义(P<0.01)。雷帕霉素组及WYE-354低剂量组皮肤移植物局部淋巴细胞浸润较阳性对照组减少,WYE-354中、高剂量组皮肤移植物局部淋巴细胞浸润较阳性对照组、雷帕霉素组、WYE-354低剂量组均减少。雷帕霉素组、WYE-354低、中、高剂量组均可下调脾细胞中4E-BP1蛋白的磷酸化水平,以WYE-354中、高剂量组作用最为明显。结论 WYE-354可以抑制mTOR信号转导通路相关蛋白磷酸化水平的调控,并阻滞其细胞周期,减少淋巴细胞向皮肤移植物浸润、显著延长小鼠皮肤移植物存活时间。

Objective To study the role of WYE-354, an ATP-competitive mammalian target of rapamycin（mTOR） kinase inhibitor, in anti-rejection of mice after skin allograft and its mechanism. Methods A skin allograft model was established with BALB/ c mice as the donor and C57BL/6 mice as the recipient. The mice were divided into auto-graft group（control group） and allograft group （WYE-354 group）. Mice in control group were given rapamycin and WYE-354 2 days after operation for 2 days and those in WYE-354 group received oral 0.2ml rapamycin（lmg/kg · d） and injection of WYE-354 at dose of lmg/（kg · d）, 1.Smg/（kg·d） and 50mg/（kg · d）, respectively. On days 3, 7, 11, and 21 after operation, 4E-BP 1 phosphorylation levels in spleen cells were measured by Western blot. On day 11 after operation, skin tissue sample was stained with H＆E. Results The survival time of mice in WYE-354 group was longer than that of mice in control group（P〈0.01 ）. The survival time of mice in control group was longer than that of mice in medium and high dose WYE-354 groups（P〈0.01）. The number of locally-infiltrated lymphocytes was less in low dose WYE-354 group than in control group. The 4E-BP1 phosphorylation level in spleen cells was down-regulated in control group and low, medium and high WYE-354 groups, especially in high dose WYE-354 group. Conclusion WYE-354 can inhibit the 4E-BP1 phosphorylation level related with the mTOR signal transduction pathway, block the cell cycle, reduce the infiltration of lymphocytes to skin grafts, and prolong the survival time of mice after skin graft.