RESUMO
Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide. However, therapies against HCC to date have not been completely effective. Sinomenine hydrochloride (SH), an antiarthritis drug applied in clinical practice, has been reported to have in vitro antineoplastic activity in various cancer cells. Whether SH inhibits HCC remains unknown. For this purpose, in this study, MTT assay was used to determine cell growth. Flow cytometry, Hoechst staining, DNA fragmentation, western blot analysis, immunohistochemisty and TUNEL staining were performed to investigate the mechanisms involved. The in vivo activity of SH was determined using a mouse xenograft model. SH inhibited the growth of various types of human HCC cells in vitro. We found that SH promoted cell cycle arrest in the G1 phase and subG1 formation, associated with the increased p21/WAF1/Cip1 expression. Additionally, SH induced caspasedependent apoptosis, which involved the disruption of mitochondrial membrane potential, the increased release of cytochrome c and Omi/HtrA2 from the mitochondria into the cytoplasm, the downregulation of Bcl2 and the upregulation of Bax, the activation of a caspase cascade (caspase8, -10, -9 and -3) and PARP, as well as the decreased expression of survivin. The SHsuppressed growth of human HCC xenografts in vivo occurred due to the decrease in proliferation and the induction of apoptosis, implicating the activation of caspase3, the upregulation of p21 and the downregulation of survivin. These findings suggest that SH exhibits anticancer efficacy in vitro and in vivo involving cell cycle and caspasedependent apoptosis and may serve as a potential drug candidate against HCC.
Assuntos
Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/patologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias Hepáticas/patologia , Morfinanos/farmacologia , Animais , Western Blotting , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/metabolismo , Caspases/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Citocromos c/metabolismo , Citometria de Fluxo , Humanos , Técnicas In Vitro , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/metabolismo , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Poli(ADP-Ribose) Polimerases/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Células Tumorais Cultivadas , Proteína X Associada a bcl-2/metabolismoRESUMO
Cyclooxygenase-2 (COX-2) and CCAAT/enhancer binding protein ß (C/EBPß) have been shown to be involved in inflammation and carcinogenesis, and our previous study revealed that they were co-overexpressed in human hepatocellular carcinoma (HCC) tissue and a positive correlation was found. Saikosaponin-d (SSD), a triterpene saponin extracted from Bupleurum falcatum L. (Umbelliferae), is known to exert inhibitory effects on COX-2 expression, together with inflammation and hepatic fibrosis. These findings prompted us to investigate the chemopreventive potential of SSD against hepatocarcinogenesis and its possible molecular mechanism in vivo. An experimental model with diethylinitrosamine (DEN)-treated Sprague Dawley rats was used in the present study. DEN (50 mg/kg body weight) and SSD (2 mg/kg body weight) were intraperitoneally injected weekly and daily, respectively. Administration of SSD alone had no side effects. The liver nodule formation, tumorous invasion to surrounding organs and increased cellular atypia induced by DEN were markedly reduced by SSD in the SSD + DEN group compared with the DEN group. On the other hand, immunohistochemical staining demonstrated that the expression of COX-2 and C/EBPß proteins was significantly increased in tumor cells and macrophages of liver tissue from DEN-treated rats, whereas the expression of the two proteins was markedly lowered in the SSD + DEN group. Overall, our results suggest that SSD prevents DEN-induced hepatocarcinogenesis in rats through inhibition of C/EBPß and COX-2, providing indispensable experimental evidence for the clinical application of SSD as a novel chemopreventive agent against HCC in the future.