Preliminary Findings of anin VitroHuman Spleen Mononuclear Cell Culture System for Primary Isolates of HIV Type 1

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Abstract
Acute HIV-1 infection is often manifested with a high level of viremia. The cell types and tissues/organs that contribute to the virus load are thought to be of central and peripheral lymphoreticular origin. The establishment and permissiveness of organ-based cell culture systems from spleen with laboratory strains or primary isolates of HIV-1 have not been reported. We studied unseparated splenic mononuclear cells (SMCs) and adherent cells derived from human spleen and liver in comparison with blood monocyte-derived macrophages (MDMs). Unstimulated, SMCs were highly permissive to primary lymphotropic HIV-1 and dual/macrophagetropic isolates (which are able to replicate in both MDMs and PBMCs). Furthermore, SMCs were found to replicate virus to high titer in a rapid log-phase manner and exhibited a prolonged stationary phase of virus production, unlike PBMCs, which required conventional activation with mitogens and exhibited a much shorter period of virus production. Interestingly, the SMCs maintained themselves as a mixed phenotype of nested lymphocytes with complex and well-differentiated macrophage(s) for extended periods of time. In addition, splenic macrophages readily purified by adherence were highly permissive to a dual/macrophage-tropic primary isolate, HIV-1ADA, intermediate with two laboratory strains, HIV-1RF and HIV-1HB3, and least permissive to the lymphotropic primary isolate HIV-1Mr452 and two other laboratory strains, HIV-1cc and HIV-1MN. The replication of HIV-1ADA as measured by extracellular p24 was sustained for up to 7 weeks and similar to the replication patterns observed with adherent hepatic macrophages and blood-derived MDMs. This study demonstrates that exogenous stimulation is not required for infection of these cells; either adherence-isolated and/or mixed lymphoid populations can be studied together, and viable stocks can be readily prepared and cryopreserved. In addition, these cells could be used for isolating new and/or other variants of HIV-1. Thus, the use of the SMC primary in vitro cell culture system for future studies involving HIV-1 is warranted.