HLA-G Expression in Human Embryonic Stem Cells and Preimplantation Embryos

Abstract

Human leukocyte Ag-G, a tolerogenic molecule that acts on cells of both innate and adaptive immunity, plays an important role in tumor progression, transplantation, placentation, as well as the protection of the allogeneic fetus from the maternal immune system. We investigated HLA-G mRNA and protein expression in human embryonic stem cells (hESC) derived from the inner cell mass (ICM) of blastocysts. hESC self-renew indefinitely in culture while maintaining pluripotency, providing an unlimited source of cells for therapy. HLA-G mRNA was present in early and late passage hESC, as assessed by real time RT-PCR. Protein expression was demonstrated by flow cytometry, immunocytochemistry, and ELISA on an hESC extract. Binding of HLA-G with its ILT2 receptor demonstrated the functional active status. To verify this finding in a physiologically relevant setting, HLA-G protein expression was investigated during preimplantation development. We demonstrated HLA-G protein expression in oocytes, cleavage stage embryos, and blastocysts, where we find it in trophectoderms but also in ICM cells. During blastocyst development, a downregulation of HLA-G in the ICM cells was present. This data might be important for cell therapy and transplantation because undifferentiated hESC can contaminate the transplant of differentiated stem cells and develop into malignant cancer cells.

Human Parthenogenetic Embryonic Stem Cell-Derived Neural Stem Cells Express HLA-G and Show Unique Resistance to NK Cell-Mediated Killing

“…Moreover, there is no neutralizing antibody for free HLA-G available (compare patent application WO2014072534A1/EP 2730588 A1, published May 14, 2014). Western blotting revealed a predominance of HLA-G1 in both N and PG NSCs as reported previously for N hESCs (31). The elevated HLA-G levels in PG compared with N NSCs may be caused by parentof-origin-specific mechanisms.…”

“…One report concluded that HLA-G is not expressed on N hESCs or differentiated progeny (16). Others showed that functionally active HLA-G is expressed (30,31). However, these discrepant results were generated using different hESC lines and different HLA-G antibodies.…”

“…A small number of HLA-G-expressing cells is sufficient to maintain an antiinflammatory milieu in the central nervous system (CNS) (29). Whether N hESCs express HLA-G remains unclear, as disparate results have been reported (16,30,31).…”

Immunogenicity and Immune-Modulating Properties of Human Stem Cells

“…Human embryonic tissues possess a range of proteins and mechanisms that efficiently counteract and prevent maternal cytotoxic T cell attack and thereby provide protection and immune privilege to the fetus (Clark, 2005;Fändrich, 2002;Mor & Abrahams, 2009;Parhar et al 1989;Petroff & Perchellet, 2010;Rebmann et al, 2010;Rizzo et al, 2011a;Verloes et al, 2011). Knowledge of the immunological tolerance mechanisms taking place at the feto-maternal interface in mouse and rat models have contributed to extend such findings to human cells.…”

“…Among known immune tolerance molecules (section 5.2), ICM-derived hESC expressed the tolerogenic HLA-G molecule at the mRNA and protein levels, but it underwent downregulation in ICM cells during blastocyst growth (Rizzo et al, 2011a;Verloes et al, 2011). Several immune-modulating proteins found in hMSC are also expressed in hESC (section 5.2; Nehlin et al, Isa et al; Sabir et al, in preparation).…”