Detection of human T-lymphotropic virus type I (HTLV-I) infection during coculture of HTLV-I infected and uninfected cells using inverse long PCR.
OBJECTIVE
Identifying the new integration of human T-lymphotropic virus type I (HTLV-I) proviral genome into initially uninfected cells after cocultivation with HTLV-I infected cells is important for clarifying the process of infection. We examined the usefulness of inverse long polymerase chain reaction (IL-PCR) for this purpose.


METHODS
An experimental system using IL-PCR was applied to detect the transmission of HTLV-I between irradiated HTLV-I infected cells (HUT102) and uninfected targed cells (MOLT4, K562) after short-term and long-term coculturing.


RESULTS
In every coculture experiment with irradiated HTLV-I infected cells and uninfected cells, the new integration of HTLV-I was easily identified by IL-PCR. Oligoclonal proliferation of HTLV-I-positive cells was shown among MOLT4 cells even 4 months after the cocultivation; however, no evidence of viral replication was observed by indirect immunofluorescence assay or reverse transcription-PCR. We also used IL-PCR to assess the inhibitory effects of azidothymidine, anti-gp46, anti-vascular cell adhesion molecule-1 and anti-heat shock cognate protein 70 (HSC70) monoclonal antibody. Integration of HTLV-I provirus was inhibited in all of these cases except for anti-HSC70.


CONCLUSIONS
This experimental method enabled the detection of cell-to-cell transmission of HTLV-I directly and was useful for studying the mechanisms of cell-associated HTLV-I infection.
Ishizaki
Junzo
J
Department of Internal Medicine II, Miyazaki Medical College, Kiyotake, Japan.
Okayama
Akihiko
A
Kuroki
Masayuki
M
Tsubouchi
Hirohito
H
D002460:Cell Line; D002461:Cell Line, Transformed; D018920:Coculture Techniques; D004279:DNA, Viral / Q000032:analysis*; D015490:HTLV-I Infections / Q000635:transmission* / Q000821:virology; D015368:Human T-lymphotropic virus 1 / Q000302:isolation & purification*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011533:Proviruses / Q000302:isolation & purification; D013601:T-Lymphocytes / Q000821:virology*; D013997:Time Factors; D016662:Virus Integration; D014779:Virus Replication
D004279:DNA, Viral