过表达脑红蛋白对大鼠脊髓损伤的神经保护作用及其机制  

Neuroprotective effect of overexpression of neuroglobin on spinal cord injury in rats and relevant mechanism

作　　者：王浩[1] 张雄[2] 唐宁曼 王晓林[1] WANG Hao;ZHANG Xiong;TANG Ning-man;WANG Xiao-lin(Dazhou Central Hospital,Dazhou 635000,Sichuan Province,China;不详)

机构地区：[1]四川省达州市中心医院,四川达州635000 [2]重庆医科大学基础医学院,重庆400016

出　　处：《中国生物制品学杂志》2021年第3期276-281,共6页Chinese Journal of Biologicals

基　　金：国家自然青年科学基金(81100948)。

摘　　要：目的观察过表达脑红蛋白(neuroglobin,NGB)对大鼠脊髓损伤(spinal cord injury,SCI)后神经功能的影响,并探讨其可能的机制。方法将90只SD大鼠随机分为5组:假手术组、SCI模型组(SCI)、阴性对照组(NC)、AAV-Ngb组(Ngb)和AAV-Ngb+LY249002组(Ngb+LY),每组18只。采用改良Allen′s法制备SCI模型,假手术组大鼠仅暴露T10脊髓背侧面,其余各组依次用微量注射器将生理盐水、AAV-Con、AAV-Ngb溶液和AAV-Ngb+LY294002液(5μmol/L)注入损伤脊髓T10所在节段的中点以及头尾两侧,于术后1、3、7、14、21、28 d,采用BBB(Basso Beattie Bresnaha)评分法评估各组大鼠的后肢运动能力;Tunel染色观察各组大鼠脊髓组织细胞的凋亡;caspase-3和caspase-9酶活性检测试剂盒检测各组大鼠脊髓组织中caspase-3和caspase-9的酶活性;Western blot检测各组大鼠脊髓组织中caspase-3、caspase-9、T(p)-Akt蛋白的表达。结果与假手术组比较,SCI和NC组大鼠的BBB后肢运动功能评分均显著降低(P <0.01),细胞凋亡数明显增多(P <0.01),而与凋亡密切相关的caspase-3和caspase-9酶活性以及蛋白表达水平也显著升高(P <0.01),而p-Akt的蛋白表达水平明显降低(P <0.01);注射Ngb后,BBB评分在术后7、14、21、28 d明显升高(P <0.01),凋亡细胞数和caspase-3、caspase-9的酶活性、蛋白表达水平均明显下降(P <0.01),p-Akt的表达也显著升高(P <0.01),且呈时间依赖性(P <0.01);而注射Akt抑制剂LY249002后,Ngb的作用部分被显著抑制(P <0.05)。结论过表达Ngb可通过激活PI3K/Akt通路抑制大鼠脊髓神经细胞的凋亡,从而对SCI发挥神经保护作用。Objective To observe the effect of overexpression of neuroglobin on neurological function after spinal cord injury(SCI)in rats,and investigate the possible mechanism. Methods Ninety SD rats were randomly divided into sham operation,SCI model(SCI),SCI with AAV as negative control(NC),SCI with AAV-Ngb(Ngb)and SCI with AVV-Ngb +LY249002(Ngb + LY)groups,18 for each. Modified Allen method was used for the establishment of SCI model,while only the back side of T10 spinal cord was exposed in sham operation. The rats in sham operation group were untreated,while those in SCI,NC,Ngb and Ngb + LY groups were injected with physiological saline,AAV-Con,AAV-Ngb and AAV-Ngb + LY294002(5 μmol/L) into the upper,middle and lower sites of T10 spinal cord segments,respectively.The motor ability of hind limbs of rats in various groups was evaluated by Basso Beattie Bresnaha(BBB)scoring method1,3,7,14,21 and 28 d after SCI. The apoptosis of spinal cord cells was observed by Tunel staining. The activities of caspase 3 and caspase 9 in spinal cord tissue were determined by the corresponding kit. The expressions of caspase 3,caspase 9 and T(p)-Akt in spinal cord tissue were determined by Western blot. Results Compared with those in sham operation group,the BBB scores for motor function of hind limbs in SCI and NC groups decreased significantly(P < 0. 01),while the apoptotic cells increased significantly(P < 0. 01),and the activities and expression levels of caspase 3 and caspase 9,which were closely associated with apoptosis,also increased significantly(P < 0. 01). However,the expression level of p-Akt protein decreased significantly(P < 0. 01). After injection with Ngb,the BBB scores 7,14,21 and 28 d after SCI increased significantly(P < 0. 01),while the number of apoptotic cells as well as activities and expression levels of caspase 3 and caspase 9 decreased significantly(P < 0. 01). However,the expression level of p-Akt increased significantly(P < 0. 01)in a time-dependent manner(P < 0. 01). After injection with Akt inhibitor LY24900

关 键 词：脑红蛋白 脊髓损伤 细胞凋亡 PI3K/AKT通路 

分 类 号：R683.2[医药卫生—骨科学]