Mesenchymal stem cell derived hematopoietic cells are permissive to HIV-1 infection

Abstract

Background: Tissue resident mesenchymal stem cells (MSCs) are multipotent, self-renewing cells known for their differentiation potential into cells of mesenchymal lineage. The ability of single cell clones isolated from adipose tissue resident MSCs (ASCs) to differentiate into cells of hematopoietic lineage has been previously demonstrated. In the present study, we investigated if the hematopoietic differentiated (HD) cells derived from ASCs could productively be infected with HIV-1.

Results: HD cells were generated by differentiating clonally expanded cultures of adherent subsets of ASCs (CD90+, CD105+, CD45-, and CD34-). Transcriptome analysis revealed that HD cells acquire a number of elements that increase their susceptibility for HIV-1 infection, including HIV-1 receptor/co-receptor and other key cellular cofactors. HIV-1 infected HD cells (HD-HIV) showed elevated p24 protein and gag and tat gene expression, implying a high and productive infection. HD-HIV cells showed decreased CD4, but significant increase in the expression of CCR5, CXCR4, Nef-associated factor HCK, and Vpu-associated factor BTRC. HIV-1 restricting factors like APOBEC3F and TRIM5 also showed up regulation. HIV-1 infection increased apoptosis and cell cycle regulatory genes in HD cells. Although undifferentiated ASCs failed to show productive infection, HIV-1 exposure increased the expression of several hematopoietic lineage associated genes such as c-Kit, MMD2, and IL-10.

Conclusions: Considering the presence of profuse amounts of ASCs in different tissues, these findings suggest the possible role that could be played by HD cells derived from ASCs in HIV-1 infection. The undifferentiated ASCs were non-permissive to HIV-1 infection; however, HIV-1 exposure increased the expression of some hematopoietic lineage related genes. The findings relate the importance of ASCs in HIV-1 research and facilitate the understanding of the disease process and management strategies.

Figure 1

Gene expression analysis of HD cells following hematopoietic differentiation. Compared to ASCs, the expression level of HIV receptor genes (CD4, CXCR4, CCR4, and CCR5) were up regulated in HD cells as result of differentiation (A). Expression of several genes involved in innate and adaptive immune reactions (B), and cell cycle regulators (C) were altered in HD cells. A fold change was applied to select genes (P < 0.05). All values are normalized to ASCs (X axis).

Figure 2

Morphological changes in differentiated HD cells following exposure to HIV-1. Day-0, HD cells which are on day 8^th of differentiation and before viral infection. Day-3, shows the morphological changes of HD cells following 72 hours of viral infection. Day-5, morphology of HD cells infected with HIV-1 virus following 120 hours of infection. Images were taken with Nikon Eclipse 2000. Scale bar is 100 μm.

Figure 3

Expression of HIV-1 p24 protein in HD-HIV and ASC-HIV cells. p24 antigen level was monitored following post exposure to HIV-1 for 24 hr. HIV-1 was exposed to undifferentiated, 5-HD and 8-HD cells and p24 level was measured after 1, 3 and 5 days following removal of virus (A). Data represent the compilation of three separate experiments carried out in triplicates (P < 0.0001). (B) HIV-1 gag expression in HD-HIV and HUT78-HIV cells and compared to HIV-1 exposed ASCs (P < 0.05). (C) mRNA was extracted from ASC-HIV, HD-HIV, and HUT78-HIV cells and RT-PCR was performed for Tat expression.

Figure 4

Comparison of the gene expression profiles of HIV-1 infected HD and HUT78 cells. Genes that were found to be differentially expressed in HD-HIV vs. HD cells in one set and HD-HIV vs. HUT78-HIV cells in another set were then grouped according to functional categories including genes encoding for HIV-1 receptors and ligands (A), cell cycle and apopotosis (B), and cellular factors involved in HIV-1 infection (C). A fold change was applied to select genes (P < 0.05). All values are normalized to either ASCs or HD cells (X axis).

Figure 5

Expression of hematopoietic markers in HD cells following HIV infection. Immunohistochemistry of HD-HIV cells, fluorescent images indicate the expression of CCR5, CCR4, NOS2, and CXCR4. Right panel shows the DIC images of identical fields. Images were obtained with Leica TCS SP-2 confocal microscope. Scale bar 10 μm.

Figure 6

Effects of HIV-1 exposure on gene expression of ASCs. The gene expression levels were calculated based on the dCT-values which have been standardized with the GADPH levels. The ASCs (n = 4 donors) were harvested following 3 and 5 days after exposure to HIV-1. IL10, c-Kit, and MMD2 expression increased significantly by 3 days post HIV-1 exposure (P < 0.05, * represents P < 0.01). The ASCs showed no morphological changes during the course of experiment.