Hepatitis C virus in human B lymphocytes transformed by Epstein-Barr virus in vitro by in situ reverse transcriptase-polymerase chain reaction.
OBJECTIVE
To study persistence and replication of hepatitis C virus (HCV) in patients' peripheral blood mononuclear cells (PBMC) cultured in vitro.


METHODS
Epstein Barr virus (EBV) was used to transform the hepatitis C virus from a HCV positive patient to permanent lymphoblastoid cell lines (LCL). Positive and negative HCV RNA strands of the cultured cells and growth media were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) each month. Core and NS5 proteins of HCV were further tested using immunohistochemical SP method and in situ RT-PCR.


RESULTS
HCV RNA positive strands were consistently detected the cultured cells for one year. The negative-strand RNA in LCL cells and the positive-strand RNA in supernatants were observed intermittently. Immunohistochemical results medicated expression of HCV NS3 and C proteins in LCL cytoplasm mostly. The positive signal of PCR product was dark blue and mainly localized to the LCL cytoplasm. The RT-PCR signal was eliminated by overnight RNase digestion but not DNase digestion.


CONCLUSIONS
HCV may exist and remain functional in a cultured cell line for a long period.
Cheng
J L
JL
Institute of Hepatology, People's Hospital, Medical Center of Beijing, University of Beijing, China.
Liu
B L
BL
Zhang
Y
Y
Tong
W B
WB
Yan
Z
Z
Feng
B F
BF
D001402:B-Lymphocytes / Q000166:cytology / Q000821:virology*; D002478:Cells, Cultured; D005260:Female; D016174:Hepacivirus / Q000235:genetics* / Q000254:growth & development / Q000302:isolation & purification; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D016133:Polymerase Chain Reaction; D012367:RNA, Viral / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014170:Transformation, Genetic; D014758:Viral Core Proteins / Q000032:analysis; D017361:Viral Nonstructural Proteins / Q000032:analysis; D014779:Virus Replication; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D012367:RNA, Viral; D014758:Viral Core Proteins; D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C069130:nucleocapsid protein, Hepatitis C virus