摘要
背景:研究表明血管内皮细胞受损与创伤性脑损伤的血脑屏障功能障碍密切相关,类表皮生长因子域7(EGF-likedomain7,EGFL7)是促进血管生成的重要因子,尚不可知EGFL7能否通过激活脑血管新生从而促进大脑创伤修复。目的:探讨EGFL7促进脑损伤后大鼠损伤脑组织中血管新生的具体机制。方法:健康成年雄性SD大鼠42只,随机分为2组,颅脑损伤组(n=36)运用改良Feeney法撞击大鼠脑硬膜建立颅脑损伤模型,假手术组(n=6)仅暴露完整脑硬膜;依照损伤后取标本时间分为颅脑损伤1,6,24h以及3,7,14d共6个亚组,每亚组各6只。经过神经功能损伤评分可用后,取挫伤灶及周围脑组织进行后续实验。另取18只SD大鼠随机分为3组,建立颅脑损伤模型,颅脑损伤1h后运用立体定向注射仪分别向大鼠脑室内注射腺病毒rAd-vector、rAd-EGFL7及rAd-EGFL7+PI3K/Akt通路抑制剂LY294002,每组6只。干预12h后取材检测。结果与结论:①RT-PCR、Westernblot及免疫组织化学染色检测显示:颅脑损伤大鼠脑组织中EGFL7、CD34表达显著上调,且随损伤后时间的增加而上调,EGFL7在颅脑损伤3d达到最大(P<0.05),EGFL7与CD34两者的表达趋势相一致,微血管生成数目在颅脑损伤7d到达最高;②大鼠颅脑损伤后1h颅内rAd-EGFL7微量注射12h后,损伤脑组织中EGFL7、CD34表达显著上调(P<0.05),同时pAkt、cyclinD1显著增加,FOXO1显著下降(P<0.05);③大鼠颅脑损伤后颅内同时注射rAd-EGFL7和PI3K/Akt信号通路抑制剂12h后,EGFL7表达变化不明显,但CD34表达显著受抑制(P<0.05),同时微血管生成数目显著减少(P<0.05);④结果证实,EGFL7在脑损伤组织中表达上调,其高表达可能与损伤脑组织中血管生成相关。EGFL7过表达激活PI3K/Akt信号通路从而促进损伤脑组织中的血管新生。
BACKGROUND: Vascular endothelial cell damage is shown to be closely related to blood brain barrier function disorder after traumatic brain injury. Epidermal growth factor-like domain 7(EGFL7) is an important factor to promote angiogenesis. Whether EGFL7 can promote traumatic brain injury repair by activating angiogenesis remains unknown. OBJECTIVE: To explore the mechanisms by which EGFL7 promotes angiogenesis after traumatic brain injury in rats. METHODS: Forty-two healthy adult male rats were randomly divided into craniocerebral injury group(n=36, craniocerebral injury model was established by impacting rat dura mater with modified Feeney method) and sham group(n=6, only complete dura mater was exposed) and the rats were divided into six subgroups according to the sample time after injury: 1 hour, 6 hours, 24 hours, 3 days, 7 days and 14 days, 6 rats in each subgroup. The contusion lesion and the surrounding brain tissue were obtained after the neurological injury score was available. Another 18 rats were selected and randomized into three groups(n=6/group) to establish craniocerebral injury model. rAd-vector, rAd-EGFL7 and combined PI3K/Akt pathway inhibitor rAd-EGFL7+LY294002 was respectively injected into the ventricles of rats at 1 hour after injury. Samples were obtained at 12 hours later. RESULTS AND CONCLUSION:(1) RT-PCR, western blot assay and immumohistochemical staining results showed that the EGFL7 and CD34 were significantly up-regulated in rat brain after craniocerebral injury(P < 0.05). The expression of EGFL7 peaked on day 3(P < 0.05), and the expression of CD34 had the same trend with EGFL7. The number of angiogenesis peaked on day 7.(2) After 12 hours of the rAd-EGFL7 microinjection in rat brain at 1 hour post-injury, the expression levels of EGFL7, CD34, pAkt and cyclin D1 were significantly up-regulated and the FOXO1 expression was significantly down-regulated(all P < 0.05).(3) Microinjection of rAd-EGFL7 and PI3K/Akt inhibitor in rat brain at 1 hour post-injury, the EGFL7 expression was not significantly changed, but the CD34 expression was significantly inhibited(P < 0.05), and the number of angiogenesis was significantly decreased(P < 0.05).(4) To conclude, up-regulated EGFL7 in the injured brain tissue may be related to angiogenesis after traumatic brain injury. Overexpression of the EGFL7 promotes the angiogenesis in the injured brain tissue through activating PI3K/Akt signaling.
作者
徐鹏翔
李强
许琼冠
罗孟亚男
成凯
谢镇明
付宙锋
Xu Pengxiang;Li Qiang;Xu Qiongguan;Luo Mengyanan;Cheng Kai;Xie Zhenming;Fu Zhoufeng(Department of Neurosurgery, the Second Affiliated Hospital of Hainan Medical University, Haikou 570311, Hainan Province, China;Department of Radiotherapy, the Second Affiliated Hospital of Hainan Medical University, Haikou 570311, Hainan Province, China)
出处
《中国组织工程研究》
CAS
北大核心
2019年第35期5619-5625,共7页
Chinese Journal of Tissue Engineering Research
基金
海南省自然科学基金项目(817332)，项目负责人：徐鹏翔~~
