We previously described a method for isolating murine hematopoietic stem cells capable of reconstituting lethally irradiated recipients, which depends solely on dual-wavelength flow cytometric analysis of murine bone marrow cells stained with the fluorescent DNA-binding dye Hoechst 33342. This method, which appears to rely on the differential ability of stem cells to efflux the Hoechst dye, defines an extremely small and homogeneous population of cells (termed SP cells). We show here that dual-wavelength analysis of Hoechst dye-stained human, rhesus and miniature swine bone marrow cells reveals a small, distinct population of cells that efflux the dye in a manner identical to murine SP cells. Like the murine SP cells, both human and rhesus SP cells are primarily CD34-negative and lineage marker-negative. In vitro culture studies demonstrated that rhesus SP cells are highly enriched for long-term culture-initiating cells (LTC-ICs), an indicator of primitive hematopoietic cells, and have the capacity for differentiation into T cells. Although rhesus SP cells do not initially possess any hematopoietic colony-forming capability, they acquire the ability to form colonies after long-term culture on bone marrow stroma, coincident with their conversion to a CD34-positive phenotype. These studies suggest the existence of a hitherto unrecognized population of hematopoietic stem cells that lack the CD34 surface marker classically associated with primitive hematopoietic cells.

It has recently been shown that rapid and profound CD4؉ T-cell depletion occurs almost exclusively within the intestinal tract of simian immunodeficiency virus (SIV)-infected macaques within days of infection. Here we demonstrate (by three-and four-color flow cytometry) that this depletion is specific to a definable subset of CD4 ؉ T cells, namely, those having both a highly and/or acutely activated (CD69 ؉ CD38 ؉ HLA-DR ؉ ) and memory (CD45RA ؊ Leu8 ؊ ) phenotype. Moreover, we demonstrate that this subset of helper T cells is found primarily within the intestinal lamina propria. Viral tropism for this particular cell type (which has been previously suggested by various studies in vitro) could explain why profound CD4 ؉ T-cell depletion occurs in the intestine and not in peripheral lymphoid tissues in early SIV infection. Furthermore, we demonstrate that an acute loss of this specific subset of activated memory CD4 ؉ T cells may also be detected in peripheral blood and lymph nodes in early SIV infection. However, since this particular cell type is present in such small numbers in circulation, its loss does not significantly affect total CD4 ؉ T cell counts. This finding suggests that SIV and, presumably, human immunodeficiency virus specifically infect, replicate in, and eliminate definable subsets of CD4 ؉ T cells in vivo.

Kaposi's sarcoma-associated herpesvirus (KSHV) K3 and K5 proteins dramatically downregulate MHC class I molecules. However, although MHC class I downregulation may protect KSHV-infected cells from cytotoxic T lymphocyte recognition, these cells become potential targets for natural killer (NK) cell-mediated lysis. We now show that K5 also downregulates ICAM-1 and B7-2, which are ligands for NK cell-mediated cytotoxicity receptors. As a consequence, K5 expression drastically inhibits NK cell-mediated cytotoxicity. Conversely, de novo expression of B7-2 and ICAM-1 resensitizes the K5-expressing cells to NK cell-mediated cytotoxicity. This is a novel viral immune evasion strategy where KSHV K5 achieves immune avoidance by downregulation of cellular ligands for NK cell-mediated cytotoxicity receptors.