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目的探讨重组人促红细胞生成素(rh EPO)对大鼠慢性脑缺血后JAK2/STAT3信号传导通路的影响,以及对Bcl-2和Bax表达的影响,分析rh EPO在慢性脑缺血中可能的作用机制。方法将60只雄性成年SD大鼠,随机分为假手术组、2VO组(双侧颈总动脉结扎模型组)、2VO+rh EPO组、2VO+rh EPO+AG490(JAK2特异性拮抗剂)组、2VO+AG490组,每组12只。各组分别腹腔注射AG490(5 mg/kg)或等量的DMSO(AG490的溶媒),30 min后鼻腔缓慢注入rh EPO(150 U/125μl)或等量生理盐水。造模后3 d给药,每周1次,共8次。给药毕24 h后取脑组织标本,HE染色观察组织形态学变化。免疫组化法检测JAK2、P-JAK2(Tyr1007 1008)、STAT3、P-STAT3(Tyr705),以及Bcl-2、Bax蛋白的表达水平。原位末端标记法(TUNEL)检测凋亡细胞数。q RT-PCR法检测Bcl-2、Bax的m RNA表达水平。结果 (1)与假手术组比较,2VO组中P-JAK2和P-STAT3表达增强,Bcl-2、Bax的表达上调(P均<0.05)。(2)与2VO组比较,2VO+rh EPO组P-JAK2、P-STAT3蛋白表达增强,并上调Bcl-2的表达和下调Bax的表达(P均<0.05)。(3)与2VO+rh EPO组比较,2VO+rh EPO+AG490组可抑制rh EPO诱导的P-JAK2、P-STAT3表达,下调Bcl-2表达和上调Bax表达(P均<0.05)。(4)2VO+AG490组中P-JAK2、P-STAT3、Bcl-2、Bax的表达与2VO组和2VO+rh EPO+AG490组比较差异无统计学意义(P均>0.05)。结论 rh EPO可能通过激活JAK2/STAT3信号转导通路,促进Bcl-2上调和Bax下调,减缓慢性脑缺血大鼠神经细胞凋亡。
Objective To investigate the effect of recombinant human erythropoietin (rhEPO) on JAK2 / STAT3 signal transduction pathway and the expression of Bcl-2 and Bax after chronic cerebral ischemia in rats, and to investigate the possible effect of rh EPO on chronic cerebral ischemia The mechanism of action. Methods Sixty adult male SD rats were randomly divided into sham operation group, 2VO group (bilateral carotid artery ligation model group), 2VO + rh EPO group, 2VO + rh EPO + AG490 (JAK2 specific antagonist group) , 2VO + AG490 group, 12 rats in each group. Each group was intraperitoneally injected with AG490 (5 mg / kg) or an equivalent amount of DMSO (vehicle for AG490), and rhEPO (150 U / 125 μl) or normal saline was slowly infused into the nasal cavity 30 minutes later. 3 d after model administration, once a week, a total of 8 times. Twenty-four hours after administration, brain samples were taken and histological changes were observed by HE staining. The expression of JAK2, P-JAK2 (Tyr1007 1008), STAT3, P-STAT3 (Tyr705) and Bcl-2, Bax protein were detected by immunohistochemistry. Apoptotic cells were detected by TUNEL. q RT-PCR method was used to detect the mRNA expression levels of Bcl-2 and Bax. Results (1) Compared with the sham operation group, the expression of P-JAK2 and P-STAT3 in 2VO group were increased and the expression of Bcl-2 and Bax were up-regulated in both groups (all P <0.05). (2) Compared with 2VO group, the expressions of P-JAK2 and P-STAT3 protein were upregulated in 2VO + rh EPO group, and the expression of Bcl-2 and Bax were up-regulated (all P <0.05). (3) Compared with 2VO + rh EPO group, 2VO + rh EPO + AG490 group inhibited rh EPO-induced P-JAK2 and P-STAT3 expression, down-regulated Bcl-2 expression and up-regulated Bax expression (all P <0.05). (4) The expressions of P-JAK2, P-STAT3, Bcl-2 and Bax in 2VO + AG490 group were not significantly different from those in 2VO group and 2VO + rh EPO + AG490 group (all P> 0.05). Conclusion rhEPO may promote the apoptosis of neurons in chronic cerebral ischemia rats by activating the JAK2 / STAT3 signal transduction pathway and up-regulating the expression of Bcl-2 and Bax.