藏红花素通过调控cGMP/PKG通路对癫痫大鼠的神经保护及对海马神经元兴奋性的影响

The neuroprotection of crocin on the rats with epilepsy and its effect on the excitability of hippocampal neurons by regulating cGMP/PKG pathway

目的观察藏红花素对癫痫大鼠的神经保护及对海马神经元兴奋性的影响。方法取45只大鼠,随机选取10只设为健康组,剩余35只建立癫痫模型,成功30只。随机分为癫痫组(10只)、藏红花素组(10只)、抑制剂组(10只);藏红花素组腹腔注射藏红花素(50 mg/kg);抑制剂组腹腔注射藏红花素(50 mg/kg),髓鞘注射ODQ[1H(1,2,4)oxadiazolo(4,3α)quinoxaline 1 one][环磷酸鸟苷(Cyclic guanosine monophosphate,cGMP)/cGMP依赖性蛋白激酶(cGMP-dependent protein kinase,PKG)通路抑制剂](10μg/只);健康组、癫痫组腹腔注射等体积生理盐水;1次/d,干预14 d;检测血清氧化应激反应水平指标;检测神经元钙离子水平;用原位末端标记法(Terminal deoxynucleoitidy transferase mediated nick end labeling,TUNEL)检测海马神经元凋亡率;用免疫印迹法检测海马组织cGMP,PKG、磷酸化-PKG(Phosphorylation-PKG,p-PKG)蛋白表达水平。结果与癫痫组比较,藏红花素组血清丙二醛(Malondialdehyde,MDA)水平、海马神经元内钙离子水平、海马神经元凋亡率降低,血清超氧化物歧化酶(Superoxide dismutase,SOD)活性水平、海马组织cGMP蛋白表达水平、p-PKG/PKG升高(P<0.05);与藏红花素组比较,抑制剂组血清MDA水平、海马神经元内钙离子水平、海马神经元凋亡率升高,血清SOD活性水平、海马组织cGMP蛋白表达水平、p-PKG/PKG降低(P<0.05)。结论藏红花素可抑制海马组织氧化应激,降低海马神经元兴奋性,保护神经功能,推测其作用机制可能与激活cGMP/PKG信号通路有关。

Objective To observe the neuroprotection of crocin on epileptic rats and its effect on the excitability of hippocampal neurons.Methods 45 rats were randomly divided into,healthy group(10 mice)and epilepsy model(35 mice).30 rats of the epilepsy model group were successfully modeling,and the 31 mice were randomly divided into the epilepsy group(10 rats),the crocin treatment group(10 rats),and the crocin-inhibitor treatment group(10 rats).The crocin group was intraperitoneally injected with crocin(50 mg/kg).In the crocin-inhibitor group,crocin(50 mg/kg)was injected intraperitoneally,and ODQ(cyclic guanosine phosphate(cGMP)/cGMP-dependent protein kinase(PKG)pathway inhibitor)(10μg each rat)was injected into the myelin sheath.The healthy group and the epilepsy group were intraperitoneally injected with an equal volume of normal saline(once/day for 14 days).Serum oxidative stress indicators,the neuronal calcium ion concentration was measured.Terminal deoxynucleotidyl transferase dUTP nick end labeling(TUNEL)was used to detect the apoptosis rate of hippocampal neurons.Western blotting was used to detect the expression of cGMP,PKG,and p-PKG in the hippocampus.Results Compared with the epilepsy group,serum malondialdehyde(MDA)level,calcium ion concentration in hippocampal neurons,the apoptosis rate of hippocampal neurons were decreasedin the crocin treatment group(P<0.05).Serum superoxide dismutase(SOD)activity level,hippocampal cGMP expression,and p-PKG/PKG were increased in the crocin group(P<0.05).Compared with the crocin group,serum MDA level,calcium ion concentration in hippocampal neurons,and the apoptosis rate of hippocampal neurons were increased;the serum SOD activity level,the hippocampal cGMP and p-PKG/PKG expression were decreased in the crocin-inhibitor treatment group(P<0.05).Conclusion Crocetin can inhibit oxidative stress in the hippocampus,reduce the excitability of hippocampal neurons,and protect nerve function.It is speculated that its mechanism is related to the activation of cGMP/PKG signaling pathway.