Expression of HLA-G1 and G5 Enables Human Mesenchymal Stem Cells To Engraft at High Levels across Xenogeneic Barriers Following Transplantation into Immunocompetent Recipients.
Authors: Yamagami Takashi , Sanada Chad , Wiendl Heinz , Zanjani Esmail D. , Porada Christopher D. , Almeida-Porada Graça 
Published: October 2019

Abstract
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-γ 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 (MSC-G1) or G5 (MSC-G5) would render these cells 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. First, we performed mixed lymphocyte reactions using unmodified MSC (unMSC), MSC-G1 or MSC-G5 as stimulators, and lymphocytes from allogeneic human or sheep donors as responders. While neither unMSC nor MSC-G1 or -G5 elicited a significant proliferative response from human lymphocytes, sheep lymphocytes proliferated 2-6 fold more when cultured with unMSC than with MSC-G1 or MSC-G5. Next, we tested whether HLA-G1 and HLA-G5 expression would enable the engraftment of human MSC in fetal sheep later 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 (55 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 Tx at 55 days resulted in similar levels of engraftment for all cell types (MSC:4±0.9; MSC-G1:6±0.3; MSC-G5:5±0.3%). In contrast, while unMSC engrafted at very low levels at 82d and 104d, Tx 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 (82d:7±1;18±2.% and 104d:12±2;16±2%). 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 at later time points. However, even at the early Tx time point of 55days, MSC-G1 and -G5 gave rise to, respectively, 5 times and 2 times more donor-derived hepatocytes than their unMSC counterpart. Our 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 diseases in which an underlying MSC defect precludes the use of the patient’s own MSC.