Syk、JNK在1型糖尿病大鼠心肌中的作用机制研究

Studies on the mechanism of Syk and JNK in the heart tissue of type 1 diabetic rats

目的分析糖尿病心肌病SD大鼠心肌组织中脾酪氨酸激酶(Syk)、c-Jun氨基末端激酶(JNK)和核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)炎症小体的表达水平,并探讨三者的作用关系。方法清洁级雄性SD大鼠随机分为对照(Ctrl)组和模型(DCM)组,DCM组采用链脲佐菌素(STZ)一次性腹腔注射方法造模,Ctrl组注射同等剂量的柠檬酸缓冲液。每周监测2组随机血糖和体质量,直至STZ注射后20周,处死取其心脏。大鼠H9c2心肌细胞随机分为正常糖处理(NG)组、高糖处理(HG)组、Syk抑制剂对照(BAY)组,Syk抑制剂高糖(HG+BAY)组。Western blot方法在蛋白水平检测大鼠心肌组织和H9c2心肌细胞中Syk、JNK的磷酸化及NLRP3的表达;逆转录聚合酶链反应(RT-PCR)方法在mRNA水平检测NLRP3、半胱氨酸天门冬氨酸特异性蛋白酶1(caspase-1)及白细胞介素(IL)-1β的表达。结果与Ctrl组相比,DCM组大鼠的随机血糖水平显著升高,体质量显著降低(P<0.05),DCM组大鼠心肌组织中p-Syk、p-JNK及NLRP3的蛋白表达明显增加(P<0.05),且NLRP3、caspase-1和IL-1βmRNA的表达也显著增加(P<0.05)。BAY处理后显著抑制了高糖诱导的H9c2心肌细胞中NLRP3、caspase-1、IL-1βmRNA的表达和p-JNK、NLRP3蛋白的表达(P<0.05)。结论 Syk诱导的JNK磷酸化及NLRP3炎症小体的活化在糖尿病心肌病的发病机制中发挥关键作用。

Objective To investigate the expressions of spleen tyrosine kinase(Syk),c-Jun amino terminal kinase(JNK)and nucleotide binding oligomerization domain-like receptor protein3(NLRP3)inflammasome in the heart tissue inSD rat model of diabetic cardiomyopathy,and to explore the relationship between Syk,JNK and NLRP3.Methods Cleanmale SD rats were randomly divided into the control(Ctrl)group and diabetic cardiomyopathy model(DCM)group.Rats ofDCM group were treated with a single intraperitoneal injection of streptozotocin(STZ),while rats of Ctrl group were injectedwith the same dose of citrate buffer.The random blood glucose level and body weight were monitored every week until20weeks after STZ or citrate buffer injection,then all the rats were killed and their hearts were obtained.Rat H9c2cardiomyocytes were randomly divided into normal glucose treatment(NG)group,high glucose treatment(HG)group,Sykinhibitor control(BAY)group and Syk inhibitor high glucose(HG+BAY)group.The Syk and JNK phosphorylations andNLRP3protein expression were detected by Western blot assay in the heart tissue of SD rats and H9c2cardiomyocytes.TheNLRP3,cysteine-containing aspartate specific protease1(caspase-1)and interleukin(IL)-1βexpressions at mRNA levelwere detected by reverse transcription-polymerase chain reaction(RT-PCR).Results The random blood glucose level wassignificantly increased(P<0.05)and the body weight was significantly decreased(P<0.05)in DCM group compared withthose of Ctrl group.The expressions of cardiac p-Syk,p-JNK and NLRP3at protein level were significantly increased inDCM group compared with those of Ctrl group(P<0.05).Furthermore,the mRNA levels of NLRP3,caspase-1and IL-1βwere significantly up-regulated(P<0.05).BAY treatment significantly inhibited the high glucose-induced NLRP3,caspase-1and IL-1βmRNA expressions and p-JNK,NLRP3protein expressions in H9c2cardiomyocytes(P<0.05).Conclusion JNK phosphorylation and NLRP3inflammasome activation induced by Syk play an important role in thepathogenesis of diabetic cardiomyopathy.