title: S-phase arrest after vincristine treatment may promote hepatitis B virus replication.
abstract: OBJECTIVE
  To observe the effect of vincristine on hepatitis B virus (HBV) replication in vitro and to study its possible mechanisms.
  
  
  METHODS
  Vincristine was added to the cultures of two cell lines stably expressing HBV. Then, the levels of hepatitis B surface antigen (HBsAg), hepatitis B e antigen (HBeAg), and hepatitis B core antigen (HBcAg) in the supernatants or cytoplasm were examined using by enzyme-linked immunosorbent assay and Western blot. The HBV pregenome RNA (pgRNA) was detected using reverse transcription-PCR and real-time fluorescent quantitative PCR (RT-qPCR), and viral DNA was detected using Southern blot and RT-qPCR. Cell proliferation after drug treatment was detected using the BrdU incorporation test and the trypan blue exclusion assay. Cell cycle and cell apoptosis were examined using flow cytometry and Western blot.
  
  
  RESULTS
  Vincristine up-regulated HBV replication directly in vitro in a dose-dependent manner, and 24-h exposure to 0.1 μmol/L vincristine induced more than 4-fold and 3-fold increases in intracellular HBV DNA and the secretion of viral DNA, respectively. The expression of HBV pgRNA, intracellular HBsAg and HBcAg, and the secretion of HBeAg were also increased significantly after drug treatment. Most importantly, vincristine promoted the cell excretion of HBV nucleocapsids instead of HBV Dane particles, and the nucleocapsids are closely related to the HBV pathogenesis. Furthermore, vincristine inhibited the proliferation of cells stably expressing HBV. The higher the concentration of the drug, the more significant the inhibition of the cell proliferation and the stronger the HBV replication ability in cells. Flow cytometry indicated that cell cycle arrest at S-phase was responsible for the cell proliferation inhibition.
  
  
  CONCLUSIONS
  Vincristine has a strong stimulatory effect on HBV replication and induces cell cycle arrest, and cell proliferation inhibition may be conducive to viral replication.
authors: Lei Xu, Zeng Tu, Ge Xu, Jie-Li Hu, Xue-Fei Cai, Yu-Wei Wang, Yuan Huang, Juan Chen, Ai-Long Huang, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing 400016, China.
authors: Lei
authors: L
authors: Xu
authors: Lei Xu, Zeng Tu, Ge Xu, Jie-Li Hu, Xue-Fei Cai, Yu-Wei Wang, Yuan Huang, Juan Chen, Ai-Long Huang, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing 400016, China.
authors: Zeng
authors: Z
authors: Tu
authors: Lei Xu, Zeng Tu, Ge Xu, Jie-Li Hu, Xue-Fei Cai, Yu-Wei Wang, Yuan Huang, Juan Chen, Ai-Long Huang, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing 400016, China.
authors: Ge
authors: G
authors: Xu
authors: Lei Xu, Zeng Tu, Ge Xu, Jie-Li Hu, Xue-Fei Cai, Yu-Wei Wang, Yuan Huang, Juan Chen, Ai-Long Huang, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing 400016, China.
authors: Jie-Li
authors: JL
authors: Hu
authors: Lei Xu, Zeng Tu, Ge Xu, Jie-Li Hu, Xue-Fei Cai, Yu-Wei Wang, Yuan Huang, Juan Chen, Ai-Long Huang, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing 400016, China.
authors: Xue-Fei
authors: XF
authors: Cai
authors: Lei Xu, Zeng Tu, Ge Xu, Jie-Li Hu, Xue-Fei Cai, Yu-Wei Wang, Yuan Huang, Juan Chen, Ai-Long Huang, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing 400016, China.
authors: Xing-Xing
authors: XX
authors: Zhan
authors: Lei Xu, Zeng Tu, Ge Xu, Jie-Li Hu, Xue-Fei Cai, Yu-Wei Wang, Yuan Huang, Juan Chen, Ai-Long Huang, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing 400016, China.
authors: Yu-Wei
authors: YW
authors: Wang
authors: Lei Xu, Zeng Tu, Ge Xu, Jie-Li Hu, Xue-Fei Cai, Yu-Wei Wang, Yuan Huang, Juan Chen, Ai-Long Huang, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing 400016, China.
authors: Yuan
authors: Y
authors: Huang
authors: Lei Xu, Zeng Tu, Ge Xu, Jie-Li Hu, Xue-Fei Cai, Yu-Wei Wang, Yuan Huang, Juan Chen, Ai-Long Huang, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing 400016, China.
authors: Juan
authors: J
authors: Chen
authors: Lei Xu, Zeng Tu, Ge Xu, Jie-Li Hu, Xue-Fei Cai, Yu-Wei Wang, Yuan Huang, Juan Chen, Ai-Long Huang, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing 400016, China.
authors: Ai-Long
authors: AL
authors: Huang
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004261:DNA Replication / Q000187:drug effects; D004279:DNA, Viral / Q000096:biosynthesis; D004305:Dose-Response Relationship, Drug; D056945:Hep G2 Cells; D006512:Hepatitis B Core Antigens / Q000378:metabolism; D006514:Hepatitis B Surface Antigens / Q000378:metabolism; D006513:Hepatitis B e Antigens / Q000378:metabolism; D006515:Hepatitis B virus / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development / Q000276:immunology; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology* / Q000821:virology*; D012367:RNA, Viral / Q000096:biosynthesis; D059807:S Phase Cell Cycle Checkpoints / Q000187:drug effects*; D013997:Time Factors; D014162:Transfection; D014750:Vincristine / Q000494:pharmacology*; D014775:Virus Activation / Q000187:drug effects; D014779:Virus Replication / Q000187:drug effects*
chemical_list: D000970:Antineoplastic Agents; D004279:DNA, Viral; D006512:Hepatitis B Core Antigens; D006514:Hepatitis B Surface Antigens; D006513:Hepatitis B e Antigens; D012367:RNA, Viral; D014750:Vincristine
keywords: Cell cycle arrest; Cytotoxic chemotherapy; Hepatitis B virus; Proliferation; Viral reactivation