Wei Jing, Ouyang Xunli, Tang Yawei, Li Han, Wang Bing, Ye Yunshan, Jin Minli, Al Azab Mahmoud, Li Weiping, Li Xia
Department of Immunology, College of Basic Medical Science, Dalian Medical University, Liaoning, China.
Department of Hematology, The Second Affiliated Hospital of Dalian Medical University, Liaoning, China.
Mod Rheumatol. 2020 May;30(3):509-516. doi: 10.1080/14397595.2019.1651446. Epub 2019 Aug 20.
To analyze the further immunomodulatory effects of endoplasmic reticulum (ER)-stressed umbilical cord-derived mesenchymal stem cells MSCs (UC-MSCs) on rheumatoid arthritis (RA) CD4CXCR5ICOS T (follicular helper-like T, Tfh) cells. MSCs were isolated from umbilical cord and surface markers were identified by flow cytometry. CD4 T cells were purified from RA patients' peripheral blood mononuclear cells (PBMCs) using immunomagnetic beads. Thapsigargin (Tg)-stimulated or unstimulated MSCs were co-cultured with RA CD4 T cells. CD4CXCR5ICOS T cells were analyzed with fluorescence activating cell sorter (FACS) and major soluble factors secreted by MSCs were detected by qRT-PCR as well as ELISA. Receptors of prostanoid E2 (PGE2), known as EP1-4, on CD4 T cells were tested with RT-PCR and FACS. Proportion of CD4CXCR5ICOS T cells was determined after EP2/EP4 antagonists and anti-IL-6R antibody was added into co-cultured system, respectively. ER-stressed MSCs further down-regulated peripheral CD4CXCR5ICOS T cells compared with Tg-stimulated MSCs and CD4 T co-cultured group. PGE2 and IL-6 increased obviously in the supernatants. EP2/EP4 could be detected on CD4 T cells and frequencies of CD4CXCR5ICOS T cells were upregulated when EP2 and/or EP4 antagonists rather than anti-IL-6R antibody were added. ER-stressed MSCs exhibited better inhibition effect on RA CD4CXCR5ICOS T cells by releasing PGE2, indicating the immunosuppressive effect of MSCs could be enhanced by induction of ER stress.
分析内质网（ER）应激的脐带间充质干细胞（UC-MSCs）对类风湿关节炎（RA）CD4⁺CXCR5⁺ICOS⁺ T（滤泡辅助样T细胞，Tfh）细胞的进一步免疫调节作用。从脐带中分离出间充质干细胞，并通过流式细胞术鉴定其表面标志物。使用免疫磁珠从RA患者外周血单个核细胞（PBMCs）中纯化CD4⁺ T细胞。将毒胡萝卜素（Tg）刺激或未刺激的间充质干细胞与RA CD4⁺ T细胞共培养。用荧光激活细胞分选仪（FACS）分析CD4⁺CXCR5⁺ICOS⁺ T细胞，并用qRT-PCR和ELISA检测间充质干细胞分泌的主要可溶性因子。用RT-PCR和FACS检测CD4⁺ T细胞上前列腺素E2（PGE2）的受体EP1-4。分别向共培养体系中加入EP2/EP4拮抗剂和抗IL-6R抗体后，测定CD4⁺CXCR5⁺ICOS⁺ T细胞的比例。与Tg刺激的间充质干细胞和CD4⁺ T细胞共培养组相比，ER应激的间充质干细胞进一步下调外周CD4⁺CXCR5⁺ICOS⁺ T细胞。上清液中PGE2和IL-6明显增加。在CD4⁺ T细胞上可检测到EP2/EP4，当加入EP2和/或EP4拮抗剂而非抗IL-6R抗体时，CD4⁺CXCR5⁺ICOS⁺ T细胞的频率上调。ER应激的间充质干细胞通过释放PGE2对RA CD4⁺CXCR5⁺ICOS⁺ T细胞表现出更好的抑制作用，表明诱导ER应激可增强间充质干细胞的免疫抑制作用。
