Inhibition of hepatitis B virus expression and replication by RNA interference in HepG2.2.15.
OBJECTIVE
To observe the inhibition of hepatitis B virus replication and expression by transfecting vector-based small interference RNA (siRNA) pGenesil-HBV X targeting HBV X gene region into HepG2.2.15 cells.


METHODS
pGenesil-HBV X was constructed and transfected into HepG2.2.15 cells via lipofection. HBV antigen secretion was determined 24, 48, and 72 h after transfection by time-resolved immunofluorometric assays (TRFIA). HBV replication was examined by fluorescence quantitative PCR, and the expression of cytoplasmic viral proteins was determined by immunohistochemistry.


RESULTS
The secretion of HBsAg and HBeAg into the supernatant was found to be inhibited by 28.5% and 32.2% (P < 0.01), and by 38.67% (P < 0.05) and 42.86% (P < 0.01) at 48 h and 72 h after pGenesil-HBV X transfection, respectively. Immunohistochemical staining for cytoplasmic HBsAg showed a similar decline in HepG2.2.15 cells 48 h after transfection. The number of HBV genomes within culture supernatants was also significantly decreased 48 h and 72 h post-transfection as quantified by fluorescence PCR (P < 0.05).


CONCLUSIONS
In HepG2.2.15 cells, HBV replication and expression is inhibited by vector-based siRNA pGenesil-HBV X targeting the HBV X coding region.
Zhao
Zhong-Fu
ZF
Institute of Hepatology, Changzhi Medical College, Changzhi 046000, Shanxi Province, China. zhaozf_1226@163.com
Yang
Hui
H
Han
De-Wu
DW
Zhao
Long-Feng
LF
Zhang
Guo-Ying
GY
Zhang
Yun
Y
Liu
Ming-She
MS
D045744:Cell Line, Tumor; D004261:DNA Replication / Q000235:genetics* / Q000502:physiology; D004279:DNA, Viral / Q000235:genetics; D005786:Gene Expression Regulation / Q000235:genetics / Q000502:physiology; D015967:Gene Expression Regulation, Viral / Q000235:genetics / Q000502:physiology; D005822:Genetic Vectors; D006511:Hepatitis B Antigens / Q000235:genetics / Q000378:metabolism; D006514:Hepatitis B Surface Antigens / Q000235:genetics / Q000378:metabolism; D006513:Hepatitis B e Antigens / Q000235:genetics / Q000378:metabolism; D006515:Hepatitis B virus / Q000235:genetics* / Q000378:metabolism; D018197:Hepatoblastoma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D010957:Plasmids / Q000235:genetics; D034622:RNA Interference / Q000502:physiology*; D034741:RNA, Small Interfering / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D054334:Viral Regulatory and Accessory Proteins
D004279:DNA, Viral; D006511:Hepatitis B Antigens; D006514:Hepatitis B Surface Antigens; D006513:Hepatitis B e Antigens; D034741:RNA, Small Interfering; D012367:RNA, Viral; D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein