Study On The Effect Of Fuyuan Xingnao Decoction On Promoting Angiogenesis After Cerebral Infarction And Its LncRNA/mRNA Expression Profil

ObjectiveBased on the HIF-1α-VEGF and its receptors /PI3K/AKT signaling pathway and the regulatory of inflammatory factors in angiogenesis,the effect mechanism of Fuyuan Xingnaotang on promoting angiogenesis after cerebral infarction was explored through the method of high-throughput sequencing of the whole transcriptome and so on.MethodsThe SD rats were selected and used to prepare the MCAO model,divided into the normal group 、 the sham operation group 、 the model group 、 the Fuyuan Xingnao Decoction group and the Nimodipine group randomly.General conditions and neurological deficit scores were assessed.Blood lipid levels,liver and kidney functions were measured.The volume of cerebral infarction and morphological indexes of rats were measured by the method of TTC and HE staining.The density of new blood vessels angiogenesis was detected by IHC,the protein and gene expression level of HIF-1α,VEGF,VEGFR1,VEGFR2,PI3 K,and AKT were detected by Western Blot and RT-q PCR.The IL-1β,IL-6 and TNF-α determined through ELISA.In addition,Cortical ischemic penumbra tissues of rats in the sham operation group 、 the model group、the Fuyuan Xingnao Decoction group and the Nimodipine group,were collected for whole transcriptome high throughput sequencing,aiming to identify relevant lncRNAs and mRNAs that were associated with the development of focal cerebral ischemia and intervention of Fuyuan Xingnao Decoction.Furthermore,potential targets of identified lncRNAs and mRNAs were predicted,and the ceRNA relationship axis constructed by combining the predicted miRNA was explored.Their enriched functions and ceRNA networks were explored.Finally,the target ceRNA relationship pair was verified by RT-q PCR.Results1.Baseline Bederson scores and Garcia JH scores of rats were comparable among groups(P > 0.05).From postoperative day 3 to day 7,neurological functions of rats in Fuyuan Xingnao Decoction group gradually recovered,manifested as lower Bederson scores and higher Garcia JH scores than those of MCAO group(P < 0.05).2.The percentage of cerebral infarct volume in the rats of Fuyuan Xingnao Decoction group was significantly lower than that of MCAO group(P < 0.01).3.Compared with those of MCAO group,relative levels of CHOL,TG,ALT,AST,UREA and UA were significantly reduced in Fuyuan Xingnao Decoction group(P <0.05).4.Pathological examinations on cerebral ischemic tissues of rats in Fuyuan Xingnao Decoction group showed mild edema,local nerve cell necrosis,aggregation of glial cells and vacuolation of local brain tissue.Notably,cerebral angiogenesis in the cortical ischemic penumbra was significantly stimulated by the intervention of Fuyuan Xingnao Decoction(P < 0.01).5.Protein and mRNA levels of VEGFA,VEGFR1,VEGFR2/p-VEGFR2,PI3K/p-PI3 K,p-AKT and HIF-1α were significantly higher than those of MCAO group to promote angiogenesis.The levels of IL-1,IL-6 and TNF-α in Fuyuan Xingnao Decoction group were significantly lower than those in the model group(P < 0.01)to inhibit inflammatory response.6.A total of 3,419 mRNAs in rat brains were differentially expressed between MCAO group and Fuyuan Xingnao Decoction group,including 2,699 upregulated and 720 downregulated.There were 1,228 differentially expressed lncRNAs between groups,including 879 upregulated and 349 downregulated.Their main functions were enriched in angiogenesis、immune and inflammation.7.Compared with the model group,the expression level of v WF,Efna4,Col9a2,Col6a1 and NONRATT008923.2 NONRATT017194.2,NONRATT017622.2,NONRATT015773.2,NONRATT011877.2,NONRATT005625.2,NONRATT012637.2,NONRATT012636.2,NONRATT022702.2,NONRATT003533.2,NONRATT014478.2,NONRATT017578.2 NONRATT018232.2 NONRATT003524.2,NONRATT003501.2raised in Fuyuan Xingnao Decoction group(P < 0.05).8.Compared with the model group,the ceRNA of NONRATT017194.2/NONRATT017622.2/NONRATT015773.2 / NONRATT011877.2/ NONRATT005625.2-rno-mi R-328a-5P-v WF related indicators at the genetic level raised(P < 0.05).Conclusions1.Fuyuan Xingnao Decoction can significantly restore brain morphology,stimulate angiogenesis and suppress inflammation in fecal cerebral ischemia rats.2.The mechanism of Fuyuan Xingnaotang in the promotion of angiogenesis in cerebral infarction may be related to the regulating differentially expression of mRNA and lncRNA.3.The effect mechanism of Fuyuan Xingnaotang in promoting angiogenesis after cerebral infarction,it may be related to the regulation axis of ceRNA network.