普伐他汀对病毒性心肌炎小鼠缝隙连接蛋白43的调节

Effects of pravastatin on myocardial connexin 43, IFN-γ and IL-10 expressions in a murine model of viral myocarditis induced by Coxsackievirus B3

目的通过比较普伐他汀不同剂量干预组和模型对照组小鼠的缝隙连接蛋白43（connexin43，Cx43）及细胞因子干扰素（IFN）-γ、白细胞介素（IL）-10的表达水平，探讨普伐他汀对急性病毒性心肌炎及其心律失常的作用，不同剂量疗效的差别及其作用机制。方法取雄性4周龄Balb／c小鼠，分为模型对照组、普伐他汀40mg·kg^-1·d^-1组、普伐他汀80mg·kg^-1·d^-1组[以上3组均予以腹腔接种柯萨奇病毒B组3型（CVB3）]，空白对照组和毒性实验组（以上2组均予以Eagle＇s液腹腔注射）。接种病毒当天（第0天），即开始给药。模型对照组和空白对照组给予生理盐水灌胃，毒性实验组用药同普伐他汀80mg·kg^-1·d^-1组，持续2周。第14天处死存活鼠。ELISA、实时定量PCR法和免疫组织化学法检测Cx43、IFN-γ和IL-10改变。结果模型对照组Cx43 mRNA明显少于普伐他汀40mg·kg^-1·d^-1组和普伐他汀80mg·kg^-1·d^-1组（1．000±0．127比1．320±0．096和1．550±0．126，P均〈0．05），免疫组织化学染色结果基本一致（0．16±0．06比4．55±0．73和5．21±0．42，P均〈0．01）。普伐他汀40mg·kg^-1·d^-1组、普伐他汀80mg·kg^-1·d^-1组IL-10 mRNA表达明显高于模型对照组（1．810±0．029和2．140±0．032比1．000±0．031，P均〈0．05），而IFN-γ mRNA模型对照组则高于普伐他汀40mg·kg^-1·d^-1组、普伐他汀80mg·kg^-1·d^-1组（1．000±0．061比0．603±0．063和0．333±0．071，P均〈0．01），血清IFN-γ、IL-10蛋白水平的改变与其核酸水平的改变基本一致。结论普伐他汀对病毒性心肌炎有一定的疗效。普伐他汀能减轻Cx43基因及蛋白表达的降低，维持IFN-γ、IL-10基因的平衡表达，表明它可通过调节细胞因子的平衡和维持缝隙连接蛋白的正常表达发挥抗病毒性心肌炎及抗心律失常的作用。

Objective To observe the effects of pravastatin on myocardial connexin 43 （ Cx43 ）, IFN-γ and IL-10 expressions in a murine model of viral myocarditis （ VMC ） induced by Coxsackievirus B3. Methods Four-week-old male Balb/c mice were inoculated intraperitonealty with Coxsackievirus B3 and then randomly received saline （group A, n = 12）, low dose pravastatin （40 mg·kg^-1·d^-1 , group B, n = 12） and high dose pravastatin （80 mg·kg^-1·d^-1 , group C, n = 12） for 14 days. Another group of mice were injected with Eagle＇s solution and treated with saline （ group D, n = 12） or high dose pravastatin （80 mg·kg^-1·d^-1 , group E, n = 12）. After 14 days treatments, serum IFN-γ, IL-10 were assayed using ELISA, myocardium IFN-γ, IL-10 and Cx43 mRNA levels were measured by real-time PCR, myocardium expression of Cx43 was also determined by immunohistochemistry staining method on cardiac myocytes. Results Pravastatin dose-dependently upregulated Cx43 proteins on membrane of myocardial cells （groupA：0.16±0.06, group B： 4.55 ±0.73 and group C： 5.21 ±0.42, P〈0.01 vs. group A） and Cx43 mRNA expression（ group A ： 1. 000 ± 0. 127, group B ： 1. 320 ± 0. 096, group C ： 1. 550 ± 0. 126, P 〈 0. 05 vs. group A）, IL-10 mRNA expression（group A： 1. 000 ±0. 031, group B：1. 810 ±0. 029,group C： 2. 140 ± 0. 032, P 〈 0.05 vs. group A） while down-regulated IFN-γ mRNA（ group A ： 1. 000 ± 0. 061, group B： 0. 603 ± 0. 063,group C： 0. 333 ± 0. 071, P 〈 0.01 vs. group A）. Serum IFN-γ,IL-10 concentrations changed in the same direction as myocardial mRNA levels of IFN-γ, IL-10 post pravastatin treatments. Conclusion Pravastatin treatments upregulated expressions of Cx43 at mRNA and protein levels and improved the INF-γ/IL-10 balance which might be the working mechanisms for pravastatin-mediated anti-VMC and anti-arrhythmia effects in this VMC model.