| Although mesenchymal stem cells (MSC) have been shown to be fairly non-immunogenic, recent studies demonstrated that MSC are capable of activating and becoming targets of NK-mediated lysis. Furthermore, the ability of IFN-gamma to induce MSC expression of HLA-II molecules shows that MSC are not completely immuno-inert. Expression of HLA-G has been associated with the maintenance of fetomaternal tolerance during pregnancy through its inhibition of the cytolytic functions of NK and cytotoxic T cells and of dendritic cell maturation. Here we investigated whether transduction of human MSC with a retroviral vector encoding HLA-G1 or G5 would render MSC and their differentiated progeny undetectable by the recipient's immune system, and thereby allow us to efficiently transplant these cells into immuno-competent xenogeneic recipients. To this end, we started by evaluating the ability of unmodified MSC (unMSC), or MSC transduced with HLA-G1 or G5 to elicit an immune response in vitro from allogeneic human or sheep lymphocytes. While neither unMSC nor MSC-G1 or -G5 elicited a significant proliferative response from human lymphocytes, sheep lymphocytes proliferated significantly more (2-6 fold) when cultured with unMSC than with MSC-G1 or MSC-G5. Furthermore NK lysis assays also showed a significant decrease in NK cell activity when cultured in the presence of MSC transduced with both HLA-G1 and G5. Next, we tested whether HLA-G1 and HLA-G5 expression by human MSC would enable the engraftment of these cells in fetal sheep, at later time points in gestation, when donor cells are normally rejected due to the presence of a competent immune system. To this end, 105 unMSC, MSC-G1, or MSC-G5 were transplanted (Tx) into fetal sheep recipients during the pre-immune period (56 days n=9), or after immunocompetence was achieved, at 82 days (n=10) or 104 days (n=8) of gestation. Evaluation of the recipients' hematopoietic system at 42 days post-transplant for the presence of human cell engraftment, using a panel of antibodies specific to human blood cells, revealed that transplantation at 56 days resulted in similar levels of engraftment for all blood cell types. In contrast, while unMSC engrafted at very low levels at 82d and 104d, transplant of MSC-G1 and -G5 at these later time points not only overcame the immune barriers, but resulted in levels of engraftment that were considerably higher than those achieved during the pre-immune period. A similar outcome was seen with liver engraftment and hepatic differentiation with MSG-1 and -G5 giving rise to 3-5 times more hepatocytes than unMSC when transplanted at later time points in gestation. However, even at the early time point of 56 days, MSC-G1 and -G5 gave rise to, respectively, 5 times and 2 times more donor-derived hepatocytes than their unMSC counterpart. We also investigated the contribution of MSC-G1 and -G5 to type II alveolar cells/Clara cells in the lung of the transplanted animals and found that more human (donor) derived Type II alveolar cells were found in animals transplanted later on in gestation. These studies demonstrate that the forced expression of HLA-G1 or G5 enables MSC to evade a competent recipient immune system and engraft at significant levels at times in gestation when donor cells are normally rejected. These studies may allow the broadening of the use of MSC to enhance the efficacy of stem cell therapies for diseases in which an underlying MSC defect precludes the use of the patient's own MSC. |
