{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,9]],"date-time":"2026-09-09T05:08:55Z","timestamp":1788930535890,"version":"build-2803163510"},"reference-count":33,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,9,9]],"date-time":"2026-09-09T00:00:00Z","timestamp":1788912000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Immunol."],"abstract":"<jats:p>\n                    Porcine vascular endothelial cells are primary targets of humoral and cellular immune responses in pig-to-primate organ xenotransplantation. Although porcine aortic endothelial cells (PAECs) are widely used to evaluate primate anti-pig immune responses, their limited proliferative capacity poses a significant obstacle to repeated and standardized experimentation. To overcome this challenge, we established immortalized PAEC lines (iPAECs) derived from WT, GTKO, and GTKO\/CMAHKO\/B4GALNT2KO (triple-knockout, TKO) donor pigs using stable transduction of the SV40 large T antigen. All iPAEC lines expanded stably beyond passage 20, maintained high CD31 expression, and preserved genotype-specific xenoantigen profiles. Anti-pig IgM\/IgG binding and complement-dependent cytotoxicity (CDC) against these cells were measured with flow cytometry using pooled rhesus monkey sera (n=20) or human sera (n=20). Compared with WT iPAECs, GTKO iPAECs demonstrated significantly reduced IgM\/IgG binding and CDC with human and rhesus sera. Additional deletion of Sda and Neu5Gc in TKO iPAECs markedly reduced human serum reactivity, whereas rhesus sera retained relatively high antibody binding and CDC against TKO iPAECs. Notably, the humoral immune responses remained consistent across primary cells and passages P5, P10, and P20, validating the long-term efficacy of iPAECs as an experimental tool. Preformed anti-pig antibodies in 30 rhesus monkeys were further evaluated using TKO iPAECs and TKO peripheral blood mononuclear cells (PBMCs). The results demonstrated a significant correlation between the two target cell types, supporting their interchangeability for humoral immune assessment. Furthermore, xenogeneic MLR assays confirmed that iPAECs could stimulate human CD4\n                    <jats:sup>+<\/jats:sup>\n                    and CD8\n                    <jats:sup>+<\/jats:sup>\n                    T-cell proliferation, especially following porcine IFN-\u03b3\u2013induced upregulation of SLA-I and SLA-II expression. Collectively, these findings establish donor-derived iPAECs as a reliable, reproducible, and genetically defined\n                    <jats:italic>in vitro<\/jats:italic>\n                    platform to comprehensively evaluate humoral and cellular xenogeneic immune responses to genetically modified porcine endothelium.\n                  <\/jats:p>","DOI":"10.3389\/fimmu.2026.1936313","type":"journal-article","created":{"date-parts":[[2026,9,9]],"date-time":"2026-09-09T04:40:30Z","timestamp":1788928830000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["Evaluation of primate anti-pig xenoreactive immune responses using immortalized aortic endothelial cells from genetically modified pigs"],"prefix":"10.3389","volume":"17","author":[{"given":"Man","family":"Zhang","sequence":"first","affiliation":[{"name":"Institute of Organ Transplantation, Tongji Hospital, Tongji Medical College, Huazhong University of Science 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China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Junlong","family":"Wu","sequence":"additional","affiliation":[{"name":"Institute of Organ Transplantation, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology","place":["Wuhan, China"]},{"name":"Key Laboratory of Organ Transplantation, Ministry of Education","place":["Wuhan, China"]},{"name":"National Health Commission (NHC) Key Laboratory of Organ Transplantation","place":["Wuhan, China"]},{"name":"Key Laboratory of Organ Transplantation, Chinese Academy of Medical Sciences","place":["Wuhan, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jie","family":"Lian","sequence":"additional","affiliation":[{"name":"Department of Pathogen Biology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology","place":["Wuhan, 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Sciences","place":["Wuhan, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lan","family":"Zhu","sequence":"additional","affiliation":[{"name":"Institute of Organ Transplantation, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology","place":["Wuhan, China"]},{"name":"Key Laboratory of Organ Transplantation, Ministry of Education","place":["Wuhan, China"]},{"name":"National Health Commission (NHC) Key Laboratory of Organ Transplantation","place":["Wuhan, China"]},{"name":"Key Laboratory of Organ Transplantation, Chinese Academy of Medical Sciences","place":["Wuhan, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shaoping","family":"Deng","sequence":"additional","affiliation":[{"name":"Chengdu Clonorgan Biotechnology Co., Ltd.","place":["Chengdu, China"]},{"name":"Xenotransplantation and Regeneration Key Laboratory of Sichuan Province, Sichuan ClonOrgan Biotechnology Co., Ltd.","place":["Neijiang, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hong","family":"Wu","sequence":"additional","affiliation":[{"name":"Laboratory of Hepatic AI Translation, Frontiers Science Center for Disease-Related Molecular Network, West China Hospital, Sichuan University","place":["Chengdu, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiaxiang","family":"Du","sequence":"additional","affiliation":[{"name":"Chengdu Clonorgan Biotechnology Co., Ltd.","place":["Chengdu, China"]},{"name":"Xenotransplantation and Regeneration Key Laboratory of Sichuan Province, Sichuan ClonOrgan Biotechnology Co., Ltd.","place":["Neijiang, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dengke","family":"Pan","sequence":"additional","affiliation":[{"name":"Chengdu Clonorgan Biotechnology Co., Ltd.","place":["Chengdu, China"]},{"name":"Xenotransplantation and Regeneration Key Laboratory of Sichuan Province, Sichuan ClonOrgan Biotechnology Co., 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