温经通脉法对奥沙利铂致外周神经毒性大鼠瞬时受体电位通道的影响

Effect of warming the meridians and unblocking the vessels on transient receptor potential channels in rats with peripheral neurotoxicity caused by oxaliplatin

目的探讨温经通脉法对奥沙利铂致外周神经毒性大鼠的神经保护作用及对背根神经节瞬时受体电位(TRP)通道的影响。方法将50只SPF级Wistar大鼠按体重随机分为空白组、模型组、甲钴胺组、温经通脉低剂量组、温经通脉高剂量组,每组10只。除空白组正常饲养外,其余组大鼠给予奥沙利铂腹腔注射,2次/周(每周的前2 d),连续4周。造模期间,甲钴胺组同时给予甲钴胺0.135 mg/kg腹腔注射,2次/周(每周的前2 d),连续4周;温经通脉低、高剂量组分别给予温经通脉汤剂6.21 g/kg、12.42 g/kg灌胃,1次/d,连续4周。记录各组大鼠实验前及实验第7,14,21,28天的体重和机械性缩足阈值(MWT);实验结束后,应用免疫组化法检测各组大鼠背根神经节中TRPA1、TRPM8阳性表达情况;每组随机选取5只大鼠分离背根神经节细胞,免疫荧光法鉴定后采用Western blot法和RT-PCR法检测背根神经节细胞中TRPA1、TRPM8、TRPV1、TRPV2、TRPV4蛋白和mRNA表达情况。结果模型组大鼠实验第14,21,28天的体重及MWT均明显低于空白组(P均<0.05),温经通脉低、高剂量组和甲钴胺组大鼠实验第21,28天的体重和实验第14,21,28天的MWT均明显高于同期模型组(P均<0.05)。实验结束后,模型组大鼠背根神经节中TRPA1、TRPM8阳性表达量和背根神经节细胞中TRPA1、TRPM8、TRPV1、TRPV2、TRPV4蛋白及mRNA相对表达量均明显高于空白组(P均<0.05),温经通脉低、高剂量组和甲钴胺组大鼠各指标均明显低于模型组(P均<0.05)。结论温经通脉法可有效减轻奥沙利铂外周神经毒性,其作用机制可能与抑制TRP通道激活相关。

Objective It is to explore the neuroprotective effect of warming the meridians and unblocking the vessels on oxaliplatin-induced peripheral neurotoxicity in rats and effect on transient receptor potential(TRP)channels in the dorsal root ganglion.Methods Fifty SPF-grade Wistar rats were randomly divided into blank group,model group,methylcobalamin group,low and high dose groups of warming the meridians and unblocking the vessels according to their body weight,10 rats in each group.Except for the rats of blank group which were fed normally,the rats in the remaining groups were given oxaliplatin intraperitoneally twice per week(the first 2 d of each week),continuously treated for 4 weeks.During the modeling period,the methylcobalamin group was also given methylcobalamin 0.135 mg intraperitoneally twice per week(the first 2 d of each week),continuously treated for 4 weeks;the low and high dose groups of warming the meridians and unblocking the vessels were respectively given 6.21 g and 12.42 g decoction for warming the meridians and unblocking the vessels by gavage,once per day,continuously treated for 4 weeks.The body weight and mechanical withdrawal threshold(MWT)of the rats in each group were recorded before the experiment and on the 7th,14th,21st and 28th days of the experiment;at the end of the experiment,the positive expression of TRPA1 and TRPM8 in the dorsal root ganglion of the rats in each group was detected by immunohistochemistry;5 rats in each group were randomly selected to isolate their dorsal root ganglion cells,and the expression of TRPA1,TRPM8,TRPV1,TRPV2 and TRPV4 proteins and mRNAs in the dorsal root ganglion cells were respectively detected by Western blot and RT-PCR after immunofluorescence identification.Results The body weight and MWT of the rats in the model group were significantly lower than those in the blank group on the 14th,21st and 28th days of experiment(all P<0.05),while the body weight and MWT of the rats in the low and high dose groups of warming the meridians and unblocking the vessels on the 21st and 28th days of experiment were significantly higher than those in the model group during the same period(all P<0.05).At the end of the experiment,the positive expression of TRPA1 and TRPM8 in the dorsal root ganglion and the relative expression of TRPA1,TRPM8,TRPV1,TRPV2,TRPV4 protein and mRNA in the dorsal root ganglion cells of the rats in the model group were significantly higher than those in the blank group(P<0.05),and the indexes of the rats in the low and high dose groups and the mecobalamin group were significantly lower than those in the model group(all P<0.05).Conclusion Warming the meridians and unblocking the vessels can effectively reduce the peripheral neurotoxicity of oxaliplatin,its action mechanism may be related to inhibiting TRP channel activation.