Immortalization of caprine fibroblasts permissive for replication of small ruminant lentiviruses

Abstract

Objective: To establish immortalized caprine fibroblastic cell lines permissive for replication of small ruminant lentiviruses.

Animals: Carpal synovial membrane explants collected aseptically from a surgically delivered fetus of a lentivirus-seronegative goat.

Procedure: Immortalization of goat embryonic fibroblasts was performed by DNA transfection with plasmids coding for simian virus 40 large T antigen. The generated cell lines were phenotypically characterized. Cytogenetics, growth pattern, and sensitivity to viral infection were studied.

Results: 3 cell lines, designated TIGEF, mMTSV-54, and mMTSV-93, were generated. They had a more rapid doubling time than did fibroblasts from which they were derived, and retained morphologic and phenotypic fibroblastic characteristics. They were immortalized but not transformed because tumor formation was not promoted after their s.c. injection into athymic nude mice. The 3 cell lines were susceptible to caprine arthritis-encephalitis virus and visna-maedi virus infections, and supported a complete virus replication cycle.

Conclusions: Cultured caprine fibroblastic cells were immortalized, using simian virus 40 large T antigen. The TIGEF, mMTSV-54, and mMTSV-93 immortalized cell lines were permissive to in vitro small ruminant lentivirus replication.

Clinical relevance: Because lentivirus detection, as well as detailed studies of host-lentivirus interactions, are hampered by differences in viral susceptibility of each primary culture, permanent cell lines are essential tools for such analysis.