IN-VITRO HUMAN IMMUNODEFICIENCY VIRUS-1 INFECTION OF PURIFIED HEMATOPOIETIC PROGENITORS IN SINGLE-CELL CULTURE

Uni- or multi-lineage suppression of hematopoiesis is observed in the majority of acquired immunodeficiency syndrome (AIDS) patients. The me chanism(s) underlying these abnormalities is not understood: particula rly, the human immunodeficiency virus (HIV) infection of hematopoietic progenitor and stem cells (HPCs/HSCs) is highly controversial. We rep ort that CD34(+) HPCs from adult peripheral blood (PB) are in part CD4 (+) and susceptible to in vitro HIV infection. Primitive CD34(+) HPCs were similar to 80% purified from PB. Double labeling for CD34 and CD4 membrane antigens was shown for 5% to 20% of the purified cells, thus suggesting their potential susceptibility to HIV-1 infection. The enr iched HPC population, challenged with purified or unpurified HIV-1 str ains, was cloned in unicellular methylcellulose culture. The single co lonies generated by erythroid burst-forming units (BFU-E), granulocyte -macrophage colony-forming units (CFU-GM), and granulocyte-erythroid-m acrophage-megakaryocyte colony-forming units (CFU-GEMM) were analyzed for the presence of HIV, ie, for gag DNA, tat mRNA, and p24 protein by PCR, reverse transcription PCR (RT-PCR), and enzyme-linked immunosorb ent assay, respectively. In the first series of experiments incubation of HPCs with HIV-1 at multiplicities of infection (MOI) ranging from 0.01 to 10 TCID50/cell consistently yielded an 11% to 17% infection ef ficiency of BFU-E-generated colonies, thus indicating the sensitivity of HPCs to in vitro HIV infection. An extensive series of experiments was then performed on HPCs challenged with HIV at 0.1 MOI level. In th e initial studies proviral gag sequences were detected in 9.2% of 121 analyzed CFU-GM colonies. In further experiments fat mRNA was monitore d in 17% and 23% of BFU-E and CFU-GM colonies, respectively, but never in CFU-GEMM clones. Finally, 12% of CFU-GM clones and rare erythroid bursts were shown to be positive for the p24 viral protein. In control studies, purified HPCs grown in liquid suspension culture were induce d to terminal unilineage erythroid, monocytic, or granulocytic differe ntiation: monocytes were consistently HIV-infected, whereas mature-ter minal erythroblasts and granulocytes were not. Our observations indica te that a minority of primitive HPCs, but not of the multipotent type, is susceptible to in vitro HIV infection. These observations may refl ect on the in vivo hematopoietic impairment in AIDS patients; more imp ortant, they provide an experimental model for studies on HIV hematopo ietic infection and in vitro tests for anti-HIV HSC gene therapy. (C) 1995 by The American Society of Hematology.