高强度间歇运动降低2型糖尿病小鼠海马神经元坏死性凋亡及炎症反应的研究

High intensity interval exercise reduced necroptosis and inflammation response of hippocampal neurons in type 2 diabetic mice

目的:观察高强度间歇运动(high intensity interval exercise,HIIT)对2型糖尿病(type 2 diabete mellitus,T2DM)小鼠海马神经元坏死性凋亡及神经炎症的影响。方法:雄性C57BL/6小鼠高脂饲养10周后腹腔注射链脲佐菌素(streptozotocin,STZ),制备T2DM小鼠模型。小鼠分组如下:健康对照组(CON,n=10),T2DM模型组(T2DM,n=10),T2DM运动组(HIIT,n=10)。HIIT组小鼠进行7周跑台训练。实验结束后,乳酸脱氢酶(lactate dehydrogenase,LDH)含量和碘化丙啶(propidium iodide,PI)染色,观察各组小鼠海马神经元细胞坏死情况;免疫荧光染色观察各组小鼠Nod样受体蛋白3(the NOD-like receptor protein3,NLRP3)水平;应用q RT-PCR(quantitative real-time polymerase chain reaction)检测各组小鼠海马组织肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-1β(interleukin-1β,IL-1β)mRNA表达。Western Blot观察各组小鼠海马组织坏死性凋亡及炎症因子相关蛋白的表达情况。结果:与CON组相比,T2DM组小鼠海马LDH释放水平显著增加(P<0.0001),PI染色阳性细胞数量、坏死性凋亡相关蛋白表达水平增加;NLRP3荧光强度增加(P<0.05),NLRP3、caspase-1、Cleaved caspase-1蛋白及炎症因子i NOS、IL-6、TNF-α、IL-1β表达明显增加(P<0.05)。7周HIIT干预后,与T2DM组相比,HIIT组小鼠海马LDH释放水平显著降低(P<0.0001),PI染色阳性细胞数量减少,坏死性凋亡相关蛋白表达水平降低;NLRP3荧光强度降低(P<0.05)、NLRP3、caspase-1、Cleaved caspase-1蛋白及促炎因子的表达水平明显降低(P<0.05)。结论:T2DM小鼠海马组织中存在坏死性凋亡,HIIT可抑制T2DM小鼠海马坏死性凋亡,降低海马组织炎症反应。

Objective:To investigate the effect of high-intensity interval exercise(HIIT)on necroptosis and neuroinflammation in hippocampus of type 2 diabetes mellitus(T2DM)mice.Method:Male C57BL/6 mice fed with high-fat for ten weeks and then injected with streptozotocin(STZ)to establish the model of T2DM mice.Mice were divided into normal control group(CON,n=10),T2DM model group(T2DM,n=10),and T2DM exercise group(HIIT,n=10).Mice in HIIT group received treadmill training for 7 weeks.At the end of the experiment,the necroptosis of hippocampal neurons was determined by lactate dehydrogenase(LDH)content and propidium iodide(PI)staining.The NOD-like receptor protein 3(NLRP3)level was examined by immunofluorescence staining.The mRNA expressions of tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)in hippocampus were evaluate by quantitative real-time polymerase chain reaction(qRT-PCR).The level of necroptosis and inflammatory cytokines related proteins in hippocampus were measured by the Western Blot.Result:Compared with the CON group,the LDH release level was significantly increased(P<0.0001),the number of PI-stained positive cells and the expression level of necroptosis related proteins were increased in T2DM group.The fluorescence intensity of NLRP3,the expression of NLRP3,caspase-1,cleaved caspase-1 protein and inflammatory cytokines iNOS,IL-6,TNF-αand IL-1βwere significantly increased(P<0.05)in T2DM group.After 7 weeks of the HIIT intervention,the hippocampal LDH release level of HIIT group was significantly lower than T2DM group(P<0.0001),the number of PI-stained positive cells and the expressions level of necroptosis related proteins was also decreased.The fluorescence intensity of NLRP3 and the expression of NLRP3,caspase-1,Cleaved caspase-1 protein and the expression levels of proinflammatory factor were all decreased in HIIT group compared to the T2DM group(P<0.05).Conclusion:Necroptosis was found in the hippocampal tissue of T2DM mice.HIIT can inhibit the necroptosis and eventually reduce the inflammatory response in the hippocampal tissue of T2DM mice.