Generation of Mesenchymal Stromal Cells with Low Immunogenicity from Human PBMC-Derived β2 Microglobulin Knockout Induced Pluripotent Stem Cells

Mesenchymal stromal cells (MSCs) are viewed as immune-privileged cells and have been broadly applied in allogeneic adoptive cell transfer for regenerative medicine or immune-suppressing purpose. However, the surface expression of human leukocyte antigen (HLA) class I molecules on MSCs could still possibly induce the rejection of allogeneic MSCs from the recipients. Here, we disrupted the β2 microglobulin (B2M) gene in human peripheral blood mononuclear cell-derived induced pluripotent stem cells (iPSCs) with two clustered regulatory interspaced short palindromic repeat (CRISPR)-associated Cas9 endonuclease-based methods. The B2M knockout iPSCs did not express HLA class I molecules but maintained their pluripotency and genome stability. Subsequently, MSCs were derived from the HLA-negative iPSCs (iMSCs). We demonstrated that B2M knockout did not affect iMSC phenotype, multipotency, and immune suppressive characteristics and, most importantly, reduced iMSC immunogenicity to allogeneic peripheral blood mononuclear cells. Thus, B2M knockout iPSCs could serve as unlimited off-the-shelf cell resources in adoptive cell transfer, while the derived iMSCs hold great potential as universal grafts in allogeneic MSC transplantation.