Laboratory for Retinal Regeneration, Center for Biosystems Dynamics Research, RIKEN, Kobe, Hyogo, Japan.
Department of Ophthalmology, Keio University School of Medicine, Tokyo, Japan.
Invest Ophthalmol Vis Sci. 2018 Aug 1;59(10):4198-4209. doi: 10.1167/iovs.18-24769.
To determine whether human induced pluripotent stem (iPS) cell-derived retinal pigment epithelial (RPE) cells (iPS-RPE) can express complement factors.
To confirm expression of complement factors in human iPS-RPE cells, we performed flow cytometry, immunohistochemistry, ELISA, and quantitative RT-PCR for the following: C3, C5, CFB (Factor B), C5b-9 (membrane attack complex [MAC]), CFH (Factor H), CFI (Factor I), CD46, CD55, CD59, clusterin, and vitronectin. We also prepared iPS-RPE cells in the presence of recombinant IFN-γ, recombinant TNF-α, lipopolysaccharide, supernatants of naïve T cells, and T helper 1 (Th1) cells. For the transplantation, after preparation of iPS-RPE cells from cynomolgus monkeys, the iPS-RPE cells (allografts) were transplanted into the subretinal space in monkeys. After surgery, monkeys were euthanized for IHC evaluation of the retinal section and determination of complement factors (C3, C5, CFB, MAC, and C1q), cytokines, and immunoglobulin G (IgG).
Human iPS-RPE cells expressed complement activators and inhibitors. iPS-RPE cells highly expressed complement factors during inflammatory conditions, especially IFN-γ exposure including Th1 cell supernatants. In immune attack eyes after allogeneic iPS-RPE cell transplantation, complement activators such as C3, CFB, C5, and MAC were detected around the host RPE layer, grafted RPE cells, inflammatory retinal lesions, and transplanted subretinal space. In addition, we observed a large number of C1q and IgG double positive and IFN-γ positive inflammatory cells in the retinal sections.
iPS-derived RPE cells greatly expressed complement factors. Thus, RPE cells might be activated and produce complement factors after exposure to infiltrating inflammatory cells in the eye.
确定人诱导多能干细胞（iPS）衍生的视网膜色素上皮（RPE）细胞（iPS-RPE）是否可以表达补体因子。
为了确认人 iPS-RPE 细胞中补体因子的表达，我们通过流式细胞术、免疫组织化学、ELISA 和定量 RT-PCR 检测了以下因子：C3、C5、CFB（因子 B）、C5b-9（膜攻击复合物[MAC]）、CFH（因子 H）、CFI（因子 I）、CD46、CD55、CD59、簇蛋白和玻连蛋白。我们还在存在重组 IFN-γ、重组 TNF-α、脂多糖、幼稚 T 细胞和辅助性 T 细胞 1（Th1）细胞上清液的情况下制备 iPS-RPE 细胞。为了进行移植，在从食蟹猴制备 iPS-RPE 细胞后，将 iPS-RPE 细胞（同种异体移植物）移植到猴的视网膜下腔。手术后，通过免疫组织化学评估视网膜切片和确定补体因子（C3、C5、CFB、MAC 和 C1q）、细胞因子和免疫球蛋白 G（IgG）来处死猴子。
人 iPS-RPE 细胞表达补体激活剂和抑制剂。在炎症条件下，iPS-RPE 细胞高度表达补体因子，尤其是在 IFN-γ暴露包括 Th1 细胞上清液的情况下。在同种异体 iPS-RPE 细胞移植后的免疫攻击眼中，在宿主 RPE 层、移植的 RPE 细胞、炎症性视网膜病变和移植的视网膜下腔周围检测到补体激活剂，如 C3、CFB、C5 和 MAC。此外，我们在视网膜切片中观察到大量 C1q 和 IgG 双阳性和 IFN-γ 阳性的炎症细胞。
iPS 衍生的 RPE 细胞大量表达补体因子。因此，RPE 细胞在暴露于眼内浸润性炎症细胞后可能被激活并产生补体因子。
