肉桂醛对0℃冷应激大鼠背根神经节神经细胞胞浆内Ca^(2+)变化及TRPA1和TRPM8 mRNA表达的影响

Effect of Cinnamaldehyde on Ca^(2+) Change and TRPA1/TRPM8 mRNA Expressions in Dorsal Root Ganglia Cells of Rats with 0 ℃ Cold Stress

目的:观察肉桂醛对0℃冷应激大鼠背根神经节神经细胞胞浆内Ca^(2+)浓度变化,并探讨其对瞬时受体电位通道蛋白A1(TRPA1)和M8(TRPM8)mRNA表达的影响。方法:取大鼠原代背根节神经细胞置于0℃冷应激状态,噻唑蓝(MTT)法检测不同时长冷应激状态下细胞的活性;加入肉桂醛,采用激光共聚焦显微镜观察细胞内外Ca^(2+)分布的变化;另以实时荧光定量PCR(Real-time PCR)检测其TRPA1,TRPM8 mRNA的表达。结果:不同时长的冷应激状态下,细胞存活度无明显差异;冷应激可降低背根神经节(DRG)神经细胞胞浆内Ca^(2+)浓度(P<0.05),一定量的肉桂醛则能使Ca^(2+)浓度显著升高(P<0.01)。Real-time PCR检测显示,长时间冷应激TRPA1 mRNA表达上调(P<0.05);长时间冷应激TRPM8 mRNA表达明显要高于短时冷应激(P<0.05);适当浓度的肉桂醛使冷应激大鼠DRG神经细胞TRPA1 mRNA表达升高(P<0.05,P<0.01),对TRPM8 mRNA的影响尚不明显。结论:冷应激能影响TRPA1,TRPM8 mRNA的表达,降低胞浆内Ca^(2+)浓度,作用机制可能为肉桂醛通过活化TRPA1通道,升高细胞浆内Ca^(2+)浓度。

Objective： To investigate the effects of cinnamaldehyde on Ca^2＋ concentration change in dorsal root ganglia cells of rats with 0 ℃ cold stress,and mRNA expressions of transient receptor potential cation channel,subfamily A,member 1（TRPA1） and transient receptor potential cation channel,subfamily M,member8（TRPM8）. Method： The rat dorsal root ganglion neuron cells were put in 0 ℃ cold stress to detect the activity of cells under the different time length of cold stress by the methylthiazolyldiphenyl-tetrazolium bromide（MTT）assay. Cinnamonaldehyde was added to detect the changes in intracellular Ca^2＋ distribution by the laser confocal microscopy. Furthermore,Real-time PCR method was used to measure the mRNA expression of TRPA1 and TRPM8. Result： There was no significant difference in cell viability under the different time length of cold stress.Cold stress decreased cytosolic Ca^2＋ concentration within dorsal root ganglia（DRG） neurons（P〈0. 05）,while a certain amount of cinnamicaldehyde could make it significantly rise（P〈0. 01）. Real-time PCR method showed that under long-time old stress,the mRNA expression of TRPA1 was up-regulated（P〈0. 05）. And the TRPM8 mRNA expression under long-time cold stress was clearly higher than that under short-time cold stress（P〈0. 05）.Cinnamaldehyde at the appropriate concentration can up-regulate the TRPA1 mRNA expression of rat DRG neurons under cold stress（P〈0. 05,P〈0. 01）. The effect of cinnamaldehyde on TRPM8 was unclear yet. Conclusion：The cold stress can impact TRPA1 and TRPM8, and decrease the cytosolic Ca^2＋ concentration. Its action mechanism is to activate TRPA1 channel and them increase the cytosolic Ca^2＋ concentration.