磷脂酶Cε1在1型糖尿病大鼠病理性神经痛中的作用初探  

Role of Phospholipase C epsilon 1 in Neuropathic Pain of Rats with Type 1 Diabetes

作　　者：郭欣[1] 赖薇 李艳华[2] 陈韵如 叶傲然 唐顺松 罗静[2] GUO Xin;LAI Wei;LI Yan-hua;CHEN Yun-ru;YE Ao-ran;TANG Shun-song;LUO Jing(Dept.of Anesthesiology,The First People’s Hospital of Yunnan Province,Kunming Yunnan 650032;Dept.of Pain Management,The 1st People’s Hospital of Yunnan Province,Kunming Yunnan 650032,China)

机构地区：[1]云南省第一人民医院麻醉科,云南昆明650032 [2]云南省第一人民医院疼痛科,云南昆明650032

出　　处：《昆明医科大学学报》2021年第2期23-28,共6页Journal of Kunming Medical University

基　　金：云南省科技厅-昆明医科大学应用基础研究联合专项基金资助项目[2019FE001(-284)];云南省卫生科技计划基金资助项目(2018NS0243)。

摘　　要：目的观察糖尿病病理性神经痛(DNP)大鼠脊髓磷脂酶Cε1(PLCE1)表达变化及RhoA抑制剂C3胞外酶对其活性影响,阐明PLCE1在DNP发生中的作用。方法24只8周龄、健康雄性SD大鼠随机分为4组(n=6):正常对照组(C组),单纯RhoA抑制剂组(E组),1型糖尿病DNP模型组(DNP组)和RhoA抑制剂干预组(EG组);静脉注射1%链脲佐菌素(STZ 65 mg/kg)用于建立1型糖尿病DNP大鼠模型。对E组和EG组大鼠进行每天1次为期1周的RhoA抑制剂C3胞外酶(10 pg/10μL)鞘内注射治疗;C组和DNP组在相应时间点鞘内给予同等容积的溶剂;应用葡萄糖测定仪测定空腹血糖浓度(mmol/L);采用von Frey细丝法和冷热板测痛仪测定大鼠后爪机械刺激缩足反射阈值(MWT)和热刺激缩足反射潜伏期(TWL)作为痛阈观察指标;Westernblot检测脊髓活化RhoA蛋白(RhoA-GTP)、总RhoA蛋白和PLCE1蛋白表达,用脊髓PLCE1蛋白表达水平和RhoA-GTP/总RhoA比值分别评价PLCE1和RhoA活性。ELISA检测脊髓TNF-α和IL-6含量以评价炎症反应。结果单纯应用RhoA抑制剂C3胞外酶对非DNP大鼠各项检测指标无明显影响;1型糖尿病DNP大鼠脊髓组织RhoA和PLCE1活性均显著增加(P<0.05),TNF-α和IL-6含量明显增多,差异有统计学意义(P<0.05),TWL及MWT显著降低,差异有统计学意义(P<0.05);C3胞外酶治疗在显著抑制1型糖尿病DNP大鼠脊髓组织RhoA表达(P<0.05)的同时,伴有脊髓PLCE1活性的显著降低,差异有统计学意义(P<0.05)、炎症因子TNF-α和IL-6生成的明显减少,差异有统计学意义(P<0.05),并显著缓解1型糖尿病DNP大鼠的机械痛敏和热痛敏。结论PLCE1在1型糖尿病大鼠DNP中发挥重要作用,其活性受RhoA调控。Objective To elucidate the pathogenic role of phospholipase C epsilon 1(PLCE1)in neuropathic pain of rats with type 1 diabetes by observing the effect of intrathecal injection of exoenzyme C3(a RhoA inhibitor)on the activity of PLCE1 and inflammatory response in the spinal cord.Methods Twenty-four 8-weekold healthy male Sprague-Dawley(SD)rats were randomly allocated to healthy rats plus vehicle group(Group C),healthy rats plus exoenzyme C3 group(Group E),type 1 diabetic neuropathic pain rats induced by streptozotocin(STZ)plus vehicle group(Group DNP)and type 1 diabetic neuropathic pain rats plus exoenzyme C3 group(Group EG).The diabetic neuropathic pain model of STZ-induced type 1 diabetes in rats was established by a single intravenous injection of STZ(65 mg/kg).In Group E and Group EG,10μL exoenzyme C3(1 pg/μL)was injected intrathecally once a day for 7 consecutive days,whereas vehicle was injected intrathecally in Group C and Group DNP.Accu-Chek Compact Plus glucose meter was used to measure fasting blood glucose concentration(mmol/L)in the tail vein once weekly.Western blot was used to detect the expressions of total RhoA and PLCE1 in the spinal cord in rats,while the RhoA activity detection kit was used to measure the expression of RhoA-GTP protein.The expression of PLCE1 protein and the ratio of RhoA-GTP/total RhoA were used to estimate the activities of PLCE1 and RhoA,respectively.Enzyme-linked immunosorbent assay(ELISA)was used to measure the contents of TNF-αand IL-6 in spinal cord to evaluate proinfammatory cytokines-induced the inflammation.The severity of DNP was evaluated by thermal withdrawal latency(TWL)and mechanical withdrawal threshold(MWT).Results No significant difference was detected between group C and group E.Rats with DNP had a decreased pain threshold(P<0.05),up-regulated activities of RhoA and PLCE1(P<0.05)and increased production of TNF-αand IL-6 in spinal cord(P<0.05)as compared to the group C and group E.Intrathecal injection of exoenzyme C3 rather than vehicle decreased the activ

关 键 词：糖尿病病理性神经痛 磷脂酶Cε1 RHOA C3胞外酶 炎症因子 

分 类 号：R614[医药卫生—麻醉学]