Susceptibility of human fetal mesencyhmal stem cells to Kaposi sarcoma-associated herpesvirus

Abstract

IntroductionThe etiologic agent of Kaposi sarcoma (KS), KS-associated herpesvirus (KSHV), 1 infects a number of cell types within KS lesions, including endothelial cells, monocyte-derived cells, and characteristic "spindle cells" that help define these tumors. 2,3 Within infected individuals, a variety of circulating bone marrowderived cells also can harbor KSHV DNA. Some of these cells can transform to spindle-shaped cells displaying morphology and cell surface characteristics similar to KS spindle cells. [4][5][6] Recent observations further implicate KSHV infection of bone marrow cells as a potential cause of pancytopenia, marrow engraftment failure, and hemophagocytic lymphohistiocytosis. [7][8][9][10] However, the precise identity of the infected marrow cells and whether they may serve as reservoirs for subsequent viral dissemination and the clinical manifestations of KSHV infection remain unknown.Human adult and fetal bone marrow contain a rare (0.01%-0.001%) but critical population of stroma-based cells-mesenchymal stem cells (MSCs)-that regulate the differentiation and proliferation of hematopoietic cells within bone marrow. 11,12 Although the mechanisms by which MSCs exert these effects are unclear, in vitro and in vivo data indicate that certain cell lineages that they influence also can be targets for KSHV infection in human patients, including CD34 ϩ progenitor cells. 12 In this study, we provide the first report of direct KSHV infection of primary human bone marrow cells, demonstrating susceptibility of fetal-derived MSCs to KSHV infection in culture. Our results raise the possibility that MSCs may play a role in the development of KSHV-related pathology and suggest the potential of this cell culture system for studying the long-term effects of KSHV infection on relevant human hematopoietic precursor cells. Study design Isolation of mesenchymal stem cellsIsolation and enrichment of adherent stem cells from human fetal bone marrow was undertaken with commercially available reagents (RosetteSep and MesenCult enrichment media, StemCell Technologies, Vancouver, BC, Canada), following the manufacturer's recommendations. MSCs were identified by flow cytometry (FACS Calibur) using a panel of fluorochromeconjugated monoclonal cell surface antibodies ( Figure 1) from Caltag (Burlingame, CA). KSHV infection of MSCsThe KSHV-infected primary effusion lymphoma (PEL) cell line, BCBL-1, was chemically induced to support lytic KSHV replication, as described previously. 13 MSCs were incubated for 2 hours with KSHV concentrated from BCBL-1 supernatants, or for 16 hours with induced BCBL-1 cells. 14 KSHV-infected MSCs were identified with an immunofluorescence assay that detects expression of the KSHV latency-associated nuclear antigen (LANA) in the nuclei of fixed cells. 14 Quantification of input KSHVThe concentration of encapsidated viral genomes from each viral preparation was determined using a fluorescently labeled KSHV DNA probe complementary to ORF73. 14 Unless otherwise stated, all cell-free infections were c...

Human but not murine multipotent mesenchymal stromal cells exhibit broad-spectrum antimicrobial effector function mediated by indoleamine 2,3-dioxygenase

“…We observed a profound dose-dependent inhibitory effect of IFNg on toxoplasma growth, which was completely reversed by addition of supplementary tryp (Figure 1c) confirming that IDO-mediated tryp depletion indeed contributes to MSC-mediated antiparasitic effector function. As recent reports have demonstrated that MSCs might be target of infections with human herpesviruses representing prominent pathogens in the setting of allogeneic hematopoietic stem cell transplantation, 8,16 we next investigated the potential impact of IDO-expression on replication of human CMV in MSCs. With increasing amounts of IFNg, we detected a 42.5-log decrease in CMV replication in MSC monolayer cultures as determined by quantitative PCR for CMV genome copy number after a 3-day culture period (Figure 1d).…”

Mesenchymal stem cells are susceptible to human herpesviruses, but viral DNA cannot be detected in the healthy seropositive individual

“…Therefore, MSC in general are not susceptible to EBV. Parsons et al 42 recently described that fetal MSC are susceptible to human herpesvirus 8/Kaposi's sarcomaassociated herpesvirus (HHV8/KSHV) belonging to the same subfamily of gammaherpesvirus as EBV. Mesenchymal stem cells are infected by both cell-free and cell-bound HHV8/KSHV, but supernatants could not transmit infection to naı¨ve cells.…”

Emerging roles for multipotent, bone marrow–derived stromal cells in host defense

“…86 Yet attention must be paid to the direct susceptibility of BMSCs to infectious pathogens. Fetal BMSCs have been shown to be in vitro targets for Kaposi sarcoma virus, 87 which may explain hematopoietic deficiencies and BM failures in these patients given the role of MSCs in hematopoiesis as previously reviewed. BMSCs have also been shown to harbor and transmit parvovirus B19 to hematopoietic cells in vitro.…”