诱导多能干细胞来源的视网膜色素上皮细胞对 T 细胞激活的抑制作用。

Inhibition of T-cell activation by retinal pigment epithelial cells derived from induced pluripotent stem cells.

机构信息

Laboratory for Retinal Regeneration, Center for Developmental Biology, RIKEN, Kobe, Japan.

Laboratory for Retinal Regeneration, Center for Developmental Biology, RIKEN, Kobe, Japan Department of Physiology, Keio University School of Medicine, Tokyo, Japan.

出版信息

Invest Ophthalmol Vis Sci. 2015 Jan 20;56(2):1051-62. doi: 10.1167/iovs.14-15619.

DOI:10.1167/iovs.14-15619

PMID:25604685

Abstract

PURPOSE

The purpose of this study was to determine whether human retinal pigment epithelial (RPE) cells from induced pluripotent stem (iPS) cells could inhibit T-cell activation in vitro.

METHODS

Cultured iPS-derived RPE (iPS-RPE) cells were established from fresh skin tissues or dental pulp cells obtained from healthy donors or a retinal patient after informed consent was obtained. To confirm expression of the specific markers on iPS and iPS-RPE cells, immunohistochemistry, quantitative RT-PCR (qRT-PCR), and flow cytometry were performed. Target T cells were obtained from peripheral blood mononuclear cells of healthy donors. Target T cells were assessed for proliferation by incorporation of bromodeoxyuridine or carboxyfluorescein succinimidyl ester for production of cytokines such as IFN-γ. Expression of TGFβ and other candidate molecules by iPS-RPE cells was evaluated with flow cytometry, ELISA, multiplex cytokine array, immunohistochemistry, and qRT-PCR.

RESULTS

The RPE cells we established from iPS cells had many characteristics of mature RPE cells but no characteristics of pluripotent stem cells. Cultured iPS-RPE cells inhibited cell proliferation and production of IFN-γ by activated CD4(+) T cells. In some bystander T cells, iPS-derived RPE cells induced CD25(+)Foxp3(+) regulatory T cells in vitro. Induced pluripotent stem-RPE cells constitutively expressed TGFβ and suppressed activation of T cells via soluble TGFβ, because TGFβ-downregulated iPS-RPE cells did not inhibit this T-cell activation.

CONCLUSIONS

Cultured iPS-derived retinal cells fully suppress T-cell activation. Transplantation of iPS-RPE cells into the eye might be a therapy for retinal disorders.

摘要

目的

本研究旨在确定诱导多能干细胞（iPS）来源的人视网膜色素上皮（RPE）细胞是否能在体外抑制 T 细胞的活化。

方法

从健康供体或视网膜患者的新鲜皮肤组织或牙髓细胞中建立 iPS 衍生的 RPE（iPS-RPE）细胞培养物。为了确认 iPS 和 iPS-RPE 细胞上特定标志物的表达，进行了免疫组织化学、定量 RT-PCR（qRT-PCR）和流式细胞术。靶 T 细胞取自健康供体的外周血单核细胞。通过掺入溴脱氧尿苷或羧基荧光素琥珀酰亚胺酯来评估靶 T 细胞的增殖，以产生细胞因子如 IFN-γ。通过流式细胞术、ELISA、多重细胞因子阵列、免疫组织化学和 qRT-PCR 评估 iPS-RPE 细胞中 TGFβ和其他候选分子的表达。

结果

我们从 iPS 细胞建立的 RPE 细胞具有许多成熟 RPE 细胞的特征，但没有多能干细胞的特征。培养的 iPS-RPE 细胞抑制激活的 CD4(+)T 细胞的细胞增殖和 IFN-γ的产生。在一些旁观者 T 细胞中，iPS 衍生的 RPE 细胞在体外诱导 CD25(+)Foxp3(+)调节性 T 细胞。诱导多能干细胞-RPE 细胞持续表达 TGFβ，并通过可溶性 TGFβ抑制 T 细胞的活化，因为 TGFβ 下调的 iPS-RPE 细胞不能抑制这种 T 细胞的活化。