Study On The Mechanism Of Structural Equation Model Of Coagulation-fibrinolysis Imbalance In Chronic Schistosome Hepatic Disease And Cantharidin Inhibiting The Growth Of Liver Cancer Cells

First part: Study on the structural equation model of coagulation-fibrinolysis imbalance in chronic schistosome hepatic diseaseObjectiveThrough structural equation model verification in patients with chronic schistosomiasis of coagulation-anticoagulant system imbalance and fibrinolysis system abnormalities, to elucidate mechanism of D-dimer(D-D) level elevated in the advanced chronic schistosome hepatic disease.MethodsIt was randomly selected 150 cases of patients with chronic schistosomiasis(slow blood group), 90 cases of patients with advanced chronic schistosomiasis(night blood group) and 69 cases of healthy control group,by enzyme linked immunosorbent assay(ELISA) detecting the content of plasma tissue type fiber plasminogen activator(tPA), urokinase type plasminogen activator mitogen activated agents(uPA), plasmin-anti-plasmin complex(PAP), plasminogen(PLG), antithrombin(AT), plasminogen activator inhibitor 1(PAI1), and immune turbidity assay of D-D levels, solidification method detecting factor VIII(F VIII) activity, chromogenic substrate method was used to detect AT-III, PLG, protein S, protein C activity.It was analyzed situation in different groups of patients with coagulation-anticoagulation system imbalance and fiber dissolution system abnormal by combining with liver function, coagulation, liver fibrosis and blood routine data. Analysis and verificate the path mechanism of structural equation model about " chronic schistosomiasis disease stage-(coagulation-fibrinolysis)-D-D" by Amos software.ResultsThe levels of FVIII, PAP, DD, TPA and uPA in slow blood group was significantly higher than that of normal control group, and late blood group was significantly higher than that of slow blood group. But the levels of AT-III, protein C, protein S, AT, PLG and PAI-1 was significantly lower than that of normal control group, and late blood group was significantly lower than that of slow blood group,it was different of traditional accounting significance(P<0.05). According to chronic schistosomiasis patients with stages and the progress of the disease, hypercoagulable state induced by coagulation-anticoagulation imbalance, thereby causing fiber soluble compensatory hyperfunction, eventually leading to patients with advanced produce high levels of D-D, we speculated and established the structure equation model path of a "chronic schistosomiasis disease stage-(coagulation-fibrinolysis)-D-D". The path coefficient was estimated by the method of maximum likelihood. By AMOS software simulation and calculation, we found that parameters with chronic schistosomiasis disease stage has good correlation, and the chi square test(χ2=28.768, P=0.274), quasi analysis degree analysis(NFI=0.957) and path coefficient of significantly(P<0.001) results show that the structural equation model has the degree of convergence and high degree of fitting and reasonable parameters.ConclusionPatients of chronic schistosomiasis, especially of advanced schistosomiasis due to blood coagulation-anti coagulation system and fibrinolysis-antifibrinolytic system imbalance format hypercoagulable state, result in intravascular thrombosis, thereby cause fibrinolysis hyperfunction, elevated levels of D-D, to maintain the body’s blood circulation. It reveals the mechanism of elevated D-D level of chronic schistosomiasis, namely chronic schistosomiasis with staging and progression by coagulation-fibrinolysis system mediated upregulation of D-D levels and fibrinolysis system in the upregulation of D-D occupied a dominant position, status of body fibrinolysis can reflect the stage of chronic schistosomiasis in a certain extent.Second part: Study on the action mechanism of cantharidin inhibiting the growth of liver cancer cellsObjectiveWith the deterioration of chronic schistosomiasis, patients may develop into liver cancer. Cantharidin has antitumor activity. This study aims to reveal the mechanism of cantharidin inhibiting hepatic cancer stem cells(HCSCs) from the prime of hepatocellular carcinoma cell line HepG2 at cell proliferation, self-renewal capacity and cell cycle arrest and apoptosis, to provide theoretical basis of liver cancer target treatment.MethodsIn this paper, the CD133+ HCSCs was isolated from the prime of hepatocellular carcinoma cell line Hep G2 by using the micro bead kit, and the CD133+ HCSCs was tested by FCM method, and the serum free DMEM medium was used to culture the cells. Secondly, the cell proliferation was detected by MTT method, and the in vitro assay was used to detect the self-renewal, WB method was used to detect the expression of β-catenin and cyclin D1, to evaluate the ability of proliferation and self-renewal. Thirdly, WB method was used to detect the expression of H2 AX, H3,Myt1,cyclin A2,cyclin B1,p53 and cdc2(Tyr 15), and the cell growth cycle distribution was evaluated by FCM.Finally, using V/PI Annexin double staining, FCM and immunofluorescence were used to detect the apoptosis state of CD133+HCSCs.ResultsCantharidin can inhibit CD133 + HCSCs cell viability and there is presence of concentration and time dependent(P<0.05).Cantharidin can inhibit CD133+ HCSCs cell proliferation and self-renewal capacity. After 48 h process of containing cantharid in cultivation, when 5 μM and 20 μM concentration of cantharidin inhibited the expression of the most obvious to CD133+ HCSCs and parental respectively. Cantharidin can regulate Wnt/β-catenin via downregulation of β-catenin and cyclin D1.Compared with the parental cells, CD133+ HCSCs by 48 h treatment of cantharidin could increase the regulation of cell cycle protein expression and phosphorylation for H2 AX, Myt1, cyclin, A2, cyclin, B1, p53 and cdc2(Tyr 15).Using FCM, It found cantharidin in 5uM time can be significantly improved in the G2/M phase of the amount of cells and decreased in the G1 phase of the amount of cells compared with the parental cells,and can also significantly improve the proportion of apoptosis CD133+ HCSCs.ConclusionCantharidin can inhibit the proliferation, self-renewal, promote cell arrest in G2/M phase and induce apoptosis of CD133 + HCSCs in extraction of the prime of hepatocellular carcinoma cell line HepG2. It can provide sufficient theoretical basis to cantharidin on liver cancer target treatment.