8-Br-cAMP和槲皮素单独和联合应用对Eca-109细胞凋亡和Bcl-2、Bax、XRCC1等凋亡相关蛋白的作用

目的探讨8-Br-cAMP和槲皮素联合应用对Eca-109细胞的凋亡和Bcl-2、Bax、XRCC1等凋亡相关蛋白的作用.方法将Eca-109细胞随机分为4组①8-Br-cAMP组(Br组); ②槲皮素组(Q组); ③(Br+Q)组; ④不加药物的对照(C)组.同时培养48 h,对以上4组细胞制备细胞滴片及硝酸纤维素膜(NCM)滴膜2种标本,分别进行Bcl-2、Bax、XRCC1的免疫组化和免疫斑点印迹实验,并以TUNEL法检测各组细胞凋亡率.结果细胞凋亡率,C组为4%,(Br+Q)组为70%,Br组为42%,Q组为35%,(Br+Q)组>Br组或Q组>C组,P<0.001.Br、Q及(Br+Q)组与对照组相比,Bcl-2-IR及XRCC1-IR皆低于C组,P<0.001,而Bax-IR则高于C组,P<0.001;Br组Q组或Br组(Br+Q)组,P>0.05.Bcl-2-IR与XRCC1-IR呈显著正相关,与Bax-IR和Bax-IR与XRCC1皆呈显著负相关.结论Br与Q联合用药与单独用药皆可下调Bcl-2及XRCC1表达,上调Bax表达;联合用药的细胞凋亡率显著高于单用Br或Q.提示,中西医联合用药可能更具有临床治疗意义.

Effects of 8-Br-cAMP alone or combined with quercetin on Bcl-2, Bax, XRCC1 in Eca-109 cells

Aim:To study the effects of 8 Br cAMP combined with quercetin on apoptosis and apoptotic related proteins, including Bcl 2, Bax, and XRCC1 in Eca 109 cells. Methods: The cultured Eca 109 cells randomized into 4 groups: 8 Br cAMP(Br) group, quercetin (Q) group, (Br+Q) group,and Control (C) group treated with no any drugs, all cultured 48 h.Two kinds of specimens including cell slide and NCM specimens were prepared and Bcl 2 IR, Bax IR,and XRCC1 IR were detected by immunohistochemistry and the immunodot blot technique. Besides, the cell apoptosis was detected with TUNEL method in each group. Results: The cell apoptotic percentage was 70% in (Br+Q) group,42% in Br group, 35% in Q group, and 4% in C group. (Br+Q)>Br or Q>C, P <0.001. When compared with C group, Bcl 2 IR and XRCC1 IR were decreased and Bax IR increased in the other 3 groups, P <0.001, Br vs Q or Br vs Q+Br, P >0.05. There was positive correlation between Bcl 2 IR and XRCC1 IR, but negative correlated with Bax IR, P <0.01. Conclusion: The Br combined with Q or single Br or Q alone could down regulate the expression of Bcl 2 and XRCC1, and up regulate that of Bax. (Br+Q) with highest cell apoptosis suggest that the traditional medicine, quercetin, combined with 8 Br cAMP treatment may be more efficient in clinic.