丙戊酸对大鼠创伤性颅脑损伤后脑水肿和神经功能的影响

The impact of valproic acid on brain edema and neuroprotection after traumatic brain injury in rats

目的 观察丙戊酸对大鼠创伤性颅脑损伤后脑水肿的影响及磷脂酰肌醇3激酶/蛋白激酶B（PI3K/Akt）信号通路在其中的作用.方法 采用Feeney法建立大鼠创伤性脑损伤模型.72只健康雄性成年SD大鼠,采用随机数字表法分为4组（n=18）：假手术组（Sham组）、创伤性颅脑损伤组（TBI组）、TBI＋丙戊酸处理组（TBI＋ VPA组）和TBI＋ PI3K特异性抑制剂LY29400组（TBI＋ LY组）.分别于伤后1、3、7d进行神经行为学评分（mNSS）;采用干湿重法测损伤区脑组织脑水含量;免疫荧光染色测定脑组织中组蛋白去乙酰化酶1（HDAC1）、基质金属蛋白酶-9（MMP-9）蛋白表达;Westerrn blot检查脑组织HDAC1、MMP-9和PI3K/Akt信号通路相关蛋白（PI3K、p-PI3K、Akt和p-Akt）表达水平的变化.结果 建模后第1、3、7天,与Sham组比较,TBI、TBI＋ VPA、TBI＋ LY组的mNSS评分显著升高,脑水含量显著增高;TBI、TBI＋ VPA、TBI＋ LY组脑组织HDAC1、MMP-9和PI3K/Akt通路相关蛋白表达均显著上调（P＜0.01）.模型建立后第3、7天,与TBI组比较,TBI＋ VPA组和TBI＋ LY组的mNSS评分显著低于TBI（TBI＋ VPA组：10.83±0.48比12.04 ±0.51、P＜0.05,8.55±0.29比10.73±0.42、P＜0.05;TBI＋ LY组：10.24±0.38比12.04±0.51,P＜0.05,7.83±0.26比10.73±0.42,P＜0.01）;脑组织含水量显著降低[TBI＋VPA组：（80.01±0.61）％比（82.87±0.69）％,P ＜0.05,（75.88±0.52）％比（78.33±0.41）％,P ＜0.05;TBI＋ LY组：（79.59±0.57）％比（82.87±0.69）％,P＜0.05,（74.94±0.43）％比（78.33±0.41）％,P＜0.01].模型建立后第3天,与TBI组比较,TBI＋ VPA组和TBI＋ LY组脑组织HDAC1和MMP-9蛋白,PI3K/Akt通路中p-PI3K和p-Akt表达下调（P＜0.05）.结论 丙戊酸可减轻大鼠创伤性脑损伤后脑水肿,改善伤后神经功能,其机制可能与抑制脑组织PI3K/Akt信号通路活化,降低MMP-9的表达相关.

Objective To investigate the effect of valproic acid （VPA） treatment on brain edema and the role of the phosphatidylinositol 3-kinases/protein Kinase B （PI3K/Akt） signaling pathway after traumatic brain injury （TBI） in rats.Methods TBI animal models were established using Feeney＆#39;s method.Seventy two SD rats were randomly divided into 4 groups：sham operation group （group Sham）,traumatic brain injury group （group TBI）,VPA treatment group （group TBI ＋ VPA） and PI3K inhibitor LY29400 group （group TBI ＋ LY） （n =18）.We measured rat behavioral outcomes by modified neurologic severity score （mNSS） tests at day 1,3,and 7 after TBI.Brain water content was measured with wet-dry weight method.The edema relatived factors of Histone deacetylase-1 （HDAC1） and matrix metalloproteinase-9 （MMP-9） expressions in TBI cerebral cortex were determined by immunohistochemistry staining and western blotting,respectively.While,the PI3K/Akt signaling pathway was tested with Western blotting.Results Compared with the Sham group,TBI group,TBI ＋ VPA group and TBI ＋ LY group had significantly higher mNSS scores,and brain water content.The protein expressions of HDAC1,MMP-9 and PI3K/Akt pathway were also increased significantly （all P 〈 0.01）.Compared with the TBl group,TBI ＋ VPA group and TBI ＋ LY group had significantly lower mNSS scores on day 3 and day 7 after TBI （TBI＋VPA group：10.83 ±0.48 vs.12.04 ±0.51,P〈0.05,8.55 ±0.29 vs.10.73 ±0.42,P〈 0.05;TBI＋LYgroup：10.24±0.38 vs.12.04±0.51,P〈0.05,7.83±0.26 vs.10.73 ±0.42,P〈 0.01）,and brain water content [TBI ＋ VPA group：（80.01 ±0.61）% vs.（82.87 ±0.69）%,P〈 0.05,（75.88±0.52）% vs.（78.33±0.41）%,P〈0.05;TBI＋LY group：（79.59±0.57）% vs.（82.87 ± 0.69） %,P 〈 0.05,（74.94 ± 0.43） % vs.（78.33 ± 0.41） %,P 〈 0.01].Meanwhile,VPA suppressed protein expressions of edema relatived factors （HDAC1,MMP-9） and activation of PI3K/Akt signaling pathway （P 〈 0.05 or 0.01）.Conclusion Treatment with VPA markedly reduced brain edema and improved neurological outcomes after TBI,possibly mediated by inhibiting MMP-9 expression,which might suppress activation of PI3K/Akt signaling pathway.