TRPV1与CaMKII相互调控在局麻药神经毒性中的作用与机制

It’s report that local anesthetics(LA) can induce calcium overload and lead to apoptosis. It’s convinced that LA can activate TRPV1 channel protein and result in the increase of intracellular calcium ion. TRPV1 is involved in nerves injury induced by LA. New findings indicate that CaMKII, downstream substrate of TRPV1, can phosphorylate TRPV1 and regulate the function of TRPV1. The phosphorylation of Ser-502 and Ser-800 can restart the inactivation TRPV1 channel. Accordingly, it’s presumed that LA induce calcium ion inflow by activating TRPV1 channel protein and activate CaMKII. The feedback control of CaMKII on TRPV1 by the phosphorylation of Ser-502 and Ser-800 makes further increase of intracellular calcium ion concentration. On the one hand the intracellular calcium stores release generous calcium and result in calcium overload by calcium induced calcium mechism(CICR), on the other hand p38 MAPK is activated sequentially, leading to apoptosis. This study will intend to use gene recombination technique to up-regulate and silence TRPV1 and CaMKII gene, detect calcium ion concentration, the situation of phosphorylation of Ser-502 and Ser-800, cell apoptosis and so on, and investigate whether LA targets cell apoptosis by mutual regulation of TRPV1 and CaMKII, in order to offer the basis of neurovirulent prevention and cure of LA.

研究报道局麻药（LA）使细胞内钙超载导致凋亡，LA激活TRPV1通道使细胞内钙浓度增高，参与LA诱发的神经损伤。最新发现，TRPV1下游底物CaMKII可磷酸化TRPV1并调控其功能，TRPV1的2个位点Ser-502和 Ser-800被磷酸化后使失活的TRPV1通道重新开放。因此，推测LA激活TRPV1后使钙离子内流，使CaMKII活化后通过磷酸化TRPV1的Ser-502和 Ser-800位点反馈调控TRPV1功能，进一步增高钙浓度，一方面通过钙诱导钙释放机制（CICR）使细胞内钙库释放大量钙离子造成钙超载，另一方面激活p38 MAPK导致细胞凋亡。本研究拟用基因重组技术，上调和沉默TRPV1、CaMKII基因，检测LA损伤时钙浓度、TRPV1 Ser-502和 Ser-800磷酸化情况、细胞凋亡等，研究LA是否通过TRPV1-CaMKII相互调控触发凋亡，为LA神经毒性防治提供依据。

研究报道局麻药（LA）使细胞内钙超载导致凋亡，LA激活TRPV1通道使细胞内钙浓度增高，参与LA诱发的神经损伤。最新发现，TRPV1下游底物CaMKII可磷酸化TRPV1并调控其功能，TRPV1的2个位点Ser-502和 Ser-800被磷酸化后使失活的TRPV1通道重新开放。因此，推测LA激活TRPV1后使钙离子内流，使CaMKII活化后通过磷酸化TRPV1的Ser-502和 Ser-800位点反馈调控TRPV1功能，进一步增高钙浓度，一方面通过钙诱导钙释放机制（CICR）使细胞内钙库释放大量钙离子造成钙超载，另一方面激活p38 MAPK导致细胞凋亡。本研究采用基因重组技术，上调和沉默TRPV1、CaMKII基因，检测LA损伤时钙浓度、TRPV1 Ser-502和 Ser-800磷酸化情况、细胞凋亡等，研究LA是否通过TRPV1-CaMKII相互调控触发凋亡，结果成功构建了上调和沉默TRPV1基因病毒载体和CaMKⅡ低表达载体，在转染U87-MG细胞后，发现其可抑制利多卡因使细胞内钙离子浓度增高的效应，并且发现利多卡因激活的CaMKII使TRPV1离子通道磷酸化，并诱导U87-MG细胞钙超载和上调caspase-3 mRNA的表达，从而引发细胞死亡。研究结果提示TRPV1、CaMKII介导了利多卡因的神经毒性作用。