普伐他汀对子痫前期样小鼠模型中长链脂肪酸氧化酶的调节作用

Regulation of pravastatin on long-chain fatty acid oxidative enzyme in pre-eclampsia-like mouse model

目的探讨普伐他汀（Pra）对子痫前期（PE）样小鼠模型中长链3-羟基酰基辅酶A脱氢酶（LCHAD）的调节作用。方法将32只C57BL／6J小鼠随机于孕7—18d每天注射亚硝基左旋精氨酸甲酯以建立PE样模型（即PE），或同期注射等剂量生理盐水为正常对照（即Con），再分别于孕8d始每天Pra灌胃（PE＋Pra、Con＋Pra组）或生理盐水灌胃（PE＋N、Con＋N组）；孕鼠共4组，每组各8只。于孕18d时收集孕鼠的肝脏及胎盘组织。通过蛋白印迹法、实时荧光定量PCR技术、免疫组化法检测，并比较LCHAD在各组孕鼠肝脏及胎盘中的表达。结果（1）PE＋N组孕鼠的动脉压自孕8d至孕18d逐渐升高，而PE＋Pra组孕鼠自孕10d始开始下降；PE＋N组孕鼠孕18d时的24h尿蛋白量[（1494±201）μg]明显高于Con＋N组[（935±128）μg]，两组比较，差异有统计学意义，（P〈0．01），也明显高于PE＋Pra组孕鼠[（981±116）μg，P〈0．01]。（2）蛋白印迹法：PE＋N组孕鼠的肝脏及胎盘LCHAD蛋白的表达水平（肝脏：0．64±0．11，胎盘：0．48±0．06）明显低于Con＋N组（肝脏：1．06±0．10，胎盘：0．60±0．10；P均〈0．01），也明显低于PE＋Pra组（肝脏：0．99±0．04，胎盘：0．60±0．08；P均〈0．01）。（3）实时荧光定量PCR技术：PE＋N组孕鼠的肝脏及胎盘LCHADmRNA的表达水平（肝脏：0．621±O．128，胎盘：0．646±0．129）明显低于Con＋N组（肝脏：1.007±0．130，胎盘：1．004±0．103；P均〈0．01），而与PE＋Pra组（肝脏：0．693±0．678，胎盘：0．662±0．183）比较，差异无统计学意义（P均〉0．05）。（4）免疫组化法：4组孕鼠肝脏组织中均有LCHAD蛋白的广泛表达，在胎盘组织中以迷路滋养层绒毛干外层绒毛滋养细胞及合体滋养细胞中表达最多。LCHAD蛋白在PE＋N组孕鼠的表达水平（肝脏：0．062±0．016，胎盘：0．147±0．018）明显低于Con＋N组（肝脏：0．126±0．013，胎盘：0．183±0．024；P均〈0．05），也明显低于PE＋Pra组（肝脏：0．111±0．017，胎盘：0．174±0．027；P均〈0．05）。结论Pra可上调PE样模型孕鼠肝脏及胎盘LCHAD蛋白的表达水平，其可能为调节PE临床表现的机制。

Objective To investigate the modulation of long-chain 3-hydroxyacyl-CoA dehydrogenase （LCHAD） expression by pravastatin in pre-eelampsia-like mouse model. Methods C57BL/6J mice were randomly injected with N-nitro-L-arginine methyl ester （L-NAME） as pre-eclampsia-like model group （PE） or saline as normal pregnaney control group （Con） respectively, from gestational the 7th to 18th day. For each group, pravastatin （PE＋Pra, Con＋Pra group） or saline （PE＋N, Con＋N Group） was given from the 8th to 18th day of gestation, respectively. Liver and placenta of pregnant mice were collected on gestational day 18. The LCHAD protein expression and mRNA levels of liver and placenta were detected through western blot, immunohistochemistry and real-time quantitative PCR. Results （1） The average arterial pressure of pregnant mice increased gradually from the 8th to 18th day in PE＋N group, but decreased in PE＋Pra group from gestational 10th day, 24 hour urinary protein levels in PE ＋ N group [（1 494 ± 201） μg] were significantly higher than that in Con＋N group [（935±128）μg, P〈0.01], and also higher than that in PE＋Pra group [（981 ± 116） μg, P〈0.01].（2） The results of western blot： the expression of LCHAD was significantly lower in PE＋N group （liver： 0.64±0.11, placenta： 0.48±0.06） than that in Con＋N group （liver： 1.06±0.10, placenta： 0.60±0.10）, and lower than that in PE＋Pra group （liver： 0.99±0.04, placenta： 0.60±0.08; all P〈 0.01）.（3）The results of real-time quantitative PCR： the levels of LCHAD mRNA in liver and placenta in PE＋ N group （liver： 0.621 ±0.128, placenta： 0.646±0.129） were significantly decreased compared with Con＋N group （liver： 1.007 ±0.130, placenta： 1.004±0.103; all P〈0.01）, but there was no significant difference between PE ＋ Pra group （liver： 0.693 ± 0.678, placenta： 0.662 ± 0.183 ; P〉0.05）. （4） LCHAD protein was expressed widely and evenly in liver. The expression in placental cytotrophoblast and syncytial trophoblast cells located in outer layer of villous in labyrinth layer was the most. The expression of LCHAD was significantly lower in PE＋N group （liver： 0.062±0.016, placenta： 0.147±0.018） than that in Con＋N group （liver： 0.126±0.013, placenta： 0.183±0.024）, and lower than that in PE＋Pra group （liver： 0.111±0.017, placenta： 0.174 ± 0.027; all P〈0.05）. Conclusion Pravastatin could upreguiate the LCHAD protein expression of liver and placenta in the pre-eclampsia-like mouse, which may be a mechanism to improve the clinical manifestations of pre-eelampsia.