Based On ALK1-LDL Pathway To Explore The Mechanism Of Gualou Xiebai Banxia Decoction In Alleviating Ox-LDL-Induced Vascular Endothelial Cell Injury

Objective: To explore the mechanism of Gualou Xiebai Banxia Decoction in alleviating endothelial cell damage and inhibiting the pathological process of AS from the cellular,molecular and genetic level more accurately by using the method of serum pharmacology,to provide experimental basis for the clinical application of Gualou Xiebai Banxia Decoction to prevent and treat coronary heart disease.Methods: Using ox-LDL to infiltrate human umbilical vein endothelial cells(HUVEC)to induce the establishment of a vascular endothelial cell injury model.Western Blot was used to detect the protein expression of LDL,ALK1 and pro-apoptotic genes Bax,protease Caspase-9,and Caspase-3.Polymerase chain reaction(PCR)technology was used to detect the m RNA expression of ALK1 and pro-apoptotic genes Bax,protease Caspase-9 and Caspase-3.To explore the mechanism of Gualou Xiebai Banxia Decoction alleviating endothelial cell injury.Results:1.Model evaluation: Flow cytometry was used to detect the apoptosis level of endothelial cells after ox-LDL infiltrated HUVEC for 24 hours.The results showed that the apoptosis level was significantly increased,and the difference was statistically significant(P<0.01).Based on this result,it can be considered that the endothelial cell injury model has been established successfully.2.Apoptosis level: Compared with the model group,the apoptosis level of endothelial cells in the drug-containing serum group,the silenced ALK1 group,and the silenced ALK1+drug-containing serum group was significantly reduced,and the difference was statistically significant(P<0.01).3.Expression levels of LDL intake,ALK1 protein and ALK1 m RNA: Silenced ALK1+drug-containing serum group had the best effect in down-regulating LDL intake and ALK1 protein expression,followed by silence ALK1 group,and drug-containing serum group had the third best effect.Compared with the model group,The expression of ALK1 m RNA in the drug serum group,the silenced ALK1 group and the silenced ALK1+drug-containing serum group were significantly reduced,and the difference was statistically significant(P<0.01);compared with the drug-containing serum group,the ALK1 m RNA expression in the silenced ALK1+drug-containing serum group was reduced,and the difference was statistically significant(P<0.05);compared with the silenced ALK1+drug-containing serum group,the ALK1 m RNA expression of the silenced ALK1 group increased,and the difference was statistically significant(P<0.05).4.Expression level of Bax,Caspase-9,Caspase-3 protein and m RNA: Silenced ALK1+drug-containing serum group had the best effect in down-regulating the protein expression of Bax,Caspase-9,and Caspase-3,followed by silenced ALK1 group,and drug-containing serum group had the third best effect.Bax: Compared with the model group,the m RNA expression of Bax in the drug serum group,the silenced ALK1 group,and the silenced ALK1+drug serum group were significantly reduced.And the difference was statistically significant(P<0.01);compared with the drug-containing serum group,the m RNA expression of Bax in the silenced ALK1+drug-containing serum group was reduced,and the difference was statistically significant(P<0.05);compared with the silenced ALK1+drug-containing serum group,the expression of Bax m RNA in the ALK1 silenced group increased,and the difference was statistically significant(P<0.05).Caspase-9: Compared with the model group,the Caspase-9 m RNA expression of the drug-containing serum group,the silenced ALK1 group and the silenced ALK1+drug-containing serum group was significantly reduced,and the difference was statistically significant(P<0.01);compared with the drug-containing serum group,the m RNA expression of Caspase-9in the silenced ALK1+drug-containing serum group was significantly reduced,and the difference was statistically significant Significance(P<0.01);Compared with the silenced ALK1+drug-containing serum group,the Caspase-9 m RNA expression in the silenced ALK1 group was significantly increased,and the difference was statistically significant(P<0.01).Caspase-3: Compared with the model group,the m RNA expression of Caspase-3 in the drug-containing serum group,the silenced ALK1 group and the silenced ALK1+drug-containing serum group were significantly reduced,and the difference was statistically significant(P<0.01);Compared with the drug-containing serum group,Caspase-3 m RNA expression in the silenced ALK1+drug-containing serum group was significantly reduced,and the difference was statistically significant(P<0.01);compared with the silenced ALK1+drug-containing serum group,the Caspase-3 m RNA expression in the silenced ALK1 group was significantly increased,and the difference was statistically significant(P<0.01).Conclusion: Gualou Xiebai Banxia Decoction can inhibit endothelial cell damage induced by ox-LDL.Part of its mechanism may be related to down-regulating the expression of ALK1 protein,inhibiting the process of combined endocytosis mediated by LDL through the ALK1 pathway,and reducing LDL intake in endothelial cells.It is related to reducing the expression levels of pro-apoptotic genes Bax and protease Caspase-9 and Caspase-3 in injured endothelial cells.In view of the fact that endothelial cell apoptosis is the central link in the occurrence of coronary heart disease,it is suggested that Gualou Xiebai Banxia Decoction can inhibit the pathological process of AS by controlling endothelial cell damage,which may be one of its important mechanisms of prevention and treatment of coronary heart disease.