Effect Of LncRNA CRNDE On Hepatocyte Proliferation And Apoptosis And Mechanism Of Jiedu Huayu Granule On Hepatic Failure

Objective:1.To observe the expression of lncRNA CRNDE in patients with liver failure and the effect of Jiedu Huayu Granule on its expression.2.To observe the expression of lncRNA CRNDE and PI3K/Akt/mT OR signaling pathway in ALF rats and the mechanism of Jiedu Huayu Granule in regulating this pathway.Methods:1.Clinical study:48 hospitalized patients with liver failure will be included,including 38 patients with chronic and acute liver failure,3 patients with subacute liver failure,2 patients with acute liver failure,and 5 patients with chronic liver failure.They will be randomly divided into a Western medicine control group and a Jiedu Huayu Granule group(referred to as JDHY group),with 24 patients in each group.At the same time,10 healthy subjects were included and named as normal group.The western medicine control group was given the comprehensive treatment of western medicine,while the JDHY group was given the second prescription of detoxicating and removing stasis on the basis of the comprehensive treatment of western medicine,with a course of 4 weeks.The normal group was not given any other intervention except for the detection of serum CRNDE content.(1)The patients in the western medicine control group and JDHY group were given blood collection once at admission,and the subjects in the normal group were arranged to collect blood once,and the serum was collected for qRT-PCR to detect the expression of CRNDE.(2)The serum CRNDE content,liver function and coagulation function were detected in the western medicine control group and JDHY group at admission and after 4 weeks of treatment,and the clinical symptom scores were calculated.2.Experimental study:Thirty-four SD rats,male,weighing 250 ± 30g,were randomly divided into three groups after adaptive feeding for one week:10 in the normal group,12 in the model group and 12 in the JDHY group,and D-GalN(600mg/kg)+LPS(lOμg/kg)was injected intraperitoneally to form a film in the model group and JDHY group.The JDHY group was given intragastric perfusion 7 days before the intraperitoneal injection,with a dose of 2mL/100g,twice in the morning and evening every day(with an interval of 12h),until 24h after the membrane formation.The normal group and model group were given the same amount of normal saline every day.After the rat model was successfully established for 24 hours,the blood and liver tissue of the abdominal aorta were collected.HE staining was used to observe the pathology of the liver tissue.The content and expression of CRNDE in the liver tissue were measured by qRT-PCR,and the expression of PI3K,Akt and CyclinD1 protein in the liver tissue were detected by Western blot,and ELISA to detect the expression levels of mTOR in liver tissues.Automated analyzer was used to detect TBil,ALT and AST indexes,and TUNEL staining was used to observe apoptosis of hepatocytes.Results:1.Clinical studies:(1)Compared with the normal group,the expression of serum CRNDE content in the western medicine control group and JDHY group decreased significantly before treatment(P<0.05);However,there was no difference between the western medicine control group and JDHY group(P>0.05).(2)Compared with before treatment,the expression of serum CRNDE in the western medicine control group and JDHY group increased after treatment,and the increase in JDHY group was more significant(P<0.05),with a statistically significant difference.(3)Compared with before treatment,TBiL,ALT,AST and PT in western medicine control group and JDHY group decreased after treatment,while ALB and PTA increased compared with before treatment,and symptom score decreased(P<0.05),with statistically significant difference;Compared with the western medicine control group,TBiL,ALT,AST and PT in JDHY group decreased,ALB and PTA increased,and symptom score decreased(P<0.05),with statistically significant difference.2.Experimental study:(1)HE staining:In the normal group,the structure of hepatic lobules in the liver tissue of rats is complete,the arrangement of hepatic cords is regular,and the arrangement is clear.The hepatocytes are radially distributed around the central vein.There is no swelling,degeneration and necrosis in the hepatocytes.The structure of hepatic sinuses is complete,and the endothelial cells of hepatic sinuses are regular and orderly arranged.There is no endothelial cell proliferation,no obvious inflammatory cell infiltration in the field of vision,and no cholestasis in the capillaries.In the model group,the liver tissue of rats was massive necrosis,almost no complete hepatic lobule structure was seen,the hepatic cord was disordered,large areas of liver cells were swollen,degenerated and necrotic,only sporadic normal liver cells were left,some liver cells were dissolved,the cell boundary was blurred,the hepatic sinus fissure was significantly expanded,the vascular endothelial cells fell off,inflammatory cells were infiltrated,and a large amount of bile was blocked in the capillaries.In JDHY group,the liver tissue of rats showed patchy necrosis,the structure of hepatic lobule was incomplete,the liver cord was still clear,the liver cells showed swelling and degeneration in varying degrees,a small number of liver cells were necrotic,a small amount of inflammatory cells were seen in the portal area,and a small amount of cholestasis was seen in the capillaries.(2)CRNDE content expression:Compared with the normal group,the expression of CRNDE content in the model group and JDHY group decreased(P<0.05),the difference was statistically significant;Compared with the model group,the expression of CRKDE in JDHY group increased significantly(P<0.05).(3)Protein expressions of PI3K,Akt,mTOR and CyclinDl:Compared with the normal group,the protein contents of PI3K,Akt,mTOR and CyclinD1 in the liver tissue of rats in the model group were decreased(P<0.05),and the differences were statistically significant;Compared with the model group,the contents of PI3K,Akt,mTOR and CyclinDl protein in the liver tissue of rats in JDHY group increased significantly(P<0.05).(4)TBiL,ALT,AST:Compared with the normal group,the TBiL,ALT,AST values in the model group were significantly higher(P<0.05),the difference was statistically significant;Compared with the model group,the TBiL,ALT and AST values in JDHY group decreased significantly(P<0.05).(5)TUNEL staining:Compared with the normal group,the number and rate of apoptosis in liver tissue of model group and JDHY group increased(P<0.05);Compared with the model group,the number and rate of apoptosis in liver tissue of JDHY group decreased(P<0.05).Conclusion(s):1.Clinical research:(1)The expression of serum CRNDE in patients with liver failure was significantly decreased,which was negatively correlated with TBil,PT and PTA.CRNDE has the effect of antagonizing liver failure.(2)Jiedu Huayu Granule can increase the expression of serum CRNDE in patients with liver failure,reduce the levels of TBil,ALT and AST,increase the level of ALB,shorten PT,increase PTA,improve the liver function,coagulation function and clinical symptoms of patients,and improve clinical efficacy.2.Experimental studies:(1)CRNDE expression decreased in liver tissues of ALF rats and positively correlated with PI3K,Akt,mTOR and CyclinD1 proteins,which promoted hepatocyte proliferation,inhibited hepatocyte apoptosis and antagonized ALF in ALF rats.(2)Detoxification and chemophoresis granules promoted hepatocyte proliferation and inhibited hepatocyte apoptosis in ALF rats through upregulating CRNDE and PI3K,Akt,mTOR and CyclinDl expression,promoting hepatocyte proliferation and inhibiting hepatocyte apoptosis in ALF rats,thus exerting antagonistic effects on liver failure.