Study On The Anti-hepatocarcinoma Effect Of Cinocobalamin And Its Components

Objective:Chinese herbal medicines and extract preparations of Traditional Chinese Medicine?TCM?are widely used for cancer treatment.Based on the analysis of the components of Huachansu extract concentrate solution and Cinobufacini injection,this project used the models?in vitro and in vivo?to evaluate the absorption of Huachansu and its components?Cinobufacini injection,Bufalin,Resibufogenin,Cinobufagin,Bufothionine etc?and its pharmacological studies on anti-hepatocarcinogenesis.Methods:PA software was adopted to assess the clinical application of dried toad skins,Cinobufacini injection,and Huachansu capsules.The components in the extract concentrate solution and the Cinobufacini injection were analyzed by HPLC method.The Caco-2 model was established and combined the HPLC method to obtain the data of the transfer volume per unit time for evaluation the absorption characteristics.In in vitro pharmacodynamic study,CCK8 assay was performed to investigate the inhibitory effects of huachansu and its chemical constituents on the growth of SMMC-7721hepatoma cells.Combined with Hochest assay to detect apoptosis fluorescence of hepatocellular carcinoma cells;Western blot was employed to study its anti-tumor effect and pharmacological mechanism.In in vivo pharmacodynamic study,H₂₂ tumor-bearing mouse models were established;HE staining method was carried to observe the therapeutic effects of Huachansu and its components on pathological tissues;ELISA and Western blot experiments were performed to study and verify the relevant signaling pathways for anti-hepatocarcinogenesis.Results:PA analysis results showed that dried toad skins,cinobufacini injection,and Huachansu capsules were widely applied in the oncology department of Longhua Hospital.Our HPLC results showed that resibufogenin,cinobufagin and bufothionine existed in cinobufacini injection and Huachansu extract concentrate solution while the concentration of bufothionine in cinobufacini injection was relatively high.In in vitro study,Caco-2 model demonstrated the single pure compounds in Huachansu could be well absorbed,verifying the feasibility of Huachansu oral drugs.Meanwhile,Huachansu and its components could significantly inhibit the growth of SMMC-7721hepatoma cells under different safe concentrations while the pro-apoptotic effect of Bufothionine was weak.However,in the western blot experiments,bufothionine and cinobufacin injection could significantly increase the expressions of LC3II and autophagy-related proteins,as ATG5 and ATG7,promoting the autophagy effects on hepatoma cells.In in vivo study,ELISA and western blot experiments suggested that Huachansu,cinobufacini injection and their components?bufalin,bufothionine?could inhibit the expression of IL-6/JAK/STAT3 inflammatory pathway,reduce the incidence of inflammatory response and ameliorate the prognosis of cancer treatment.Conclusions:Huacansu,cinobufacini injection and their ingredients?bufalin,bufothionine?could reduce the incidence of tumor inflammatory response and promote the autophagy.As the main component in cinobufacin injection,anti-apoptosis function of bufothionine might be weak,but its pro-autophagy function might be strong.The anti-hepatocarcinoma effects of Huachansu,cinobufacini injection and their components involved in the interaction of apoptosis,autophagy and inflammatory mechanisms and deserved further study.