PMID- 32930470 OWN - NLM STAT- MEDLINE DCOM- 20210902 LR - 20210902 IS - 1549-4918 (Electronic) IS - 1066-5099 (Linking) VI - 38 IP - 11 DP - 2020 Nov TI - Generation of hypoimmunogenic human pluripotent stem cells via expression of membrane-bound and secreted beta2m-HLA-G fusion proteins. PG - 1423-1437 LID - 10.1002/stem.3269 [doi] AB - Allogeneic immune rejection is a major barrier for the application of human pluripotent stem cells (hPSCs) in regenerative medicine. A broad spectrum of immune cells, including T cells, natural killer (NK) cells, and antigen-presenting cells, which either cause direct cell killing or constitute an immunogenic environment, are involved in allograft immune rejection. A strategy to protect donor cells from cytotoxicity while decreasing the secretion of inflammatory cytokines of lymphocytes is still lacking. Here, we engineered hPSCs with no surface expression of classical human leukocyte antigen (HLA) class I proteins via beta-2 microglobulin (B2M) knockout or biallelic knockin of HLA-G1 within the frame of endogenous B2M loci. Elimination of the surface expression of HLA class I proteins protected the engineered hPSCs from cytotoxicity mediated by T cells. However, this lack of surface expression also resulted in missing-self response and NK cell activation, which were largely compromised by expression of beta2m-HLA-G1 fusion proteins. We also proved that the engineered beta2m-HLA-G5 fusion proteins were soluble, secretable, and capable of safeguarding low immunogenic environments by lowering inflammatory cytokines secretion in allografts. Our current study reveals a novel strategy that may offer unique advantages to construct hypoimmunogenic hPSCs via the expression of membrane-bound and secreted beta2m-HLA-G fusion proteins. These engineered hPSCs are expected to serve as an unlimited cell source for generating universally compatible "off-the-shelf" cell grafts in the future. CI - (c) 2020 AlphaMed Press. FAU - Shi, Lei AU - Shi L AD - College of Animal Sciences, Zhejiang University, Hangzhou, People's Republic of China. AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Li, Wenjing AU - Li W AD - College of Animal Sciences, Zhejiang University, Hangzhou, People's Republic of China. FAU - Liu, Yang AU - Liu Y AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Chen, Zhenyu AU - Chen Z AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Hui, Yi AU - Hui Y AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Hao, Pengcheng AU - Hao P AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Xu, Xiangjie AU - Xu X AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Zhang, Shuwei AU - Zhang S AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Feng, Hexi AU - Feng H AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Zhang, Bowen AU - Zhang B AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Zhou, Shanshan AU - Zhou S AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Li, Nan AU - Li N AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. FAU - Xiao, Lei AU - Xiao L AD - College of Animal Sciences, Zhejiang University, Hangzhou, People's Republic of China. AD - Shanghai SiDanSai Biotechnology Limited Company, Shanghai, People's Republic of China. FAU - Liu, Ling AU - Liu L AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Translational Medical Center for Stem Cell Therapy, Shanghai East Hospital, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Shanghai Institute of Stem Cell Research and Clinical Translation, Shanghai, People's Republic of China. FAU - Ma, Lin AU - Ma L AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Translational Medical Center for Stem Cell Therapy, Shanghai East Hospital, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Shanghai Institute of Stem Cell Research and Clinical Translation, Shanghai, People's Republic of China. FAU - Zhang, Xiaoqing AU - Zhang X AUID- ORCID: 0000-0002-7405-1893 AD - Brain and Spinal Cord Clinical Research Center, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Key Laboratory of Neuroregeneration of Shanghai Universities, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Translational Medical Center for Stem Cell Therapy, Shanghai East Hospital, Tongji University School of Medicine, Shanghai, People's Republic of China. AD - Shanghai Institute of Stem Cell Research and Clinical Translation, Shanghai, People's Republic of China. AD - Key Laboratory of Reconstruction and Regeneration of Spine and Spinal Cord Injury, Ministry of Education, Shanghai, People's Republic of China. AD - Tsingtao Advanced Research Institute, Tongji University, Qingdao, People's Republic of China. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20200915 PL - England TA - Stem Cells JT - Stem cells (Dayton, Ohio) JID - 9304532 RN - 0 (Membrane Fusion Proteins) SB - IM MH - Animals MH - Cell Culture Techniques MH - Humans MH - Male MH - Membrane Fusion Proteins/*metabolism MH - Mice MH - Pluripotent Stem Cells/*metabolism OTO - NOTNLM OT - B2M OT - CRISPR-Cas9 OT - HLA-G OT - allogeneic OT - hypoimmunogenic OT - natural killer cell EDAT- 2020/09/16 06:00 MHDA- 2021/09/03 06:00 CRDT- 2020/09/15 08:54 PHST- 2020/03/29 00:00 [received] PHST- 2020/07/24 00:00 [revised] PHST- 2020/08/11 00:00 [accepted] PHST- 2020/09/16 06:00 [pubmed] PHST- 2021/09/03 06:00 [medline] PHST- 2020/09/15 08:54 [entrez] AID - 10.1002/stem.3269 [doi] PST - ppublish SO - Stem Cells. 2020 Nov;38(11):1423-1437. doi: 10.1002/stem.3269. Epub 2020 Sep 15.