Heparin-Binding Epidermal-like Growth Factor (HB-EGF) Reduces Cell Death in an Organoid Model of Retinal Damage

Organoids Pub Date : 2024-07-05 DOI:10.3390/organoids3030010
M. N. H. Tang, M. Moosajee, Najam A Sharif, G. A. Limb, K. Eastlake
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Abstract

In zebrafish and various mammalian species, HB-EGF has been shown to promote Müller glia proliferation and activation of repair mechanisms that have not been fully investigated in human retina. In the current study, 70- to 90-day-old human retinal organoids were treated with 20 μM 4-hydroxytamoxifen (4-OHT), and CRX, REC, NRL, PAX6, VIM, GFAP, and VSX2 gene and protein expression were assessed at various times points after treatment. Organoids with or without 4-OHT-induced damage were then cultured with HB-EGF for 7 days. We showed that 20 μM 4-OHT caused a reduction in the number of recoverin-positive cells; an increase in the number of TUNEL-positive cells; and downregulation of the photoreceptor gene markers CRX, NRL, and REC. Culture of organoids with HB-EGF for 7 days after 4-OHT-induced damage caused a marked reduction in the number of TUNEL-positive cells and small increases in the number of Ki67-positive cells and PAX6 and NOTCH1 gene expression. The current results suggest that treatment of human ESC-derived retinal organoids with 4-OHT may be used as a model of retinal degeneration in vitro. Furthermore, HB-EGF treatment of human retinal organoids increases proliferating Müller cells, but only after 4-OHT induced damage, and may be an indication of Muller reactivity in response to photoreceptor damage. Further studies will aim to identify factors that may induce Müller cell-mediated regeneration of the human retina, aiding in the development of therapies for retinal degeneration.
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肝素结合表皮样生长因子(HB-EGF)可减少视网膜损伤类器官模型中的细胞死亡
在斑马鱼和各种哺乳动物中,HB-EGF 被证明能促进 Müller 胶质增殖和修复机制的激活,而在人类视网膜中,HB-EGF 尚未得到充分研究。在本研究中,用 20 μM 4-hydroxytamoxifen (4-OHT) 处理 70 至 90 天大的人类视网膜器官组织,并在处理后的不同时间点评估 CRX、REC、NRL、PAX6、VIM、GFAP 和 VSX2 基因和蛋白的表达。然后用 HB-EGF 培养有或无 4-OHT 诱导损伤的器官组织 7 天。我们发现,20 μM 4-OHT 会导致恢复素阳性细胞数量减少;TUNEL 阳性细胞数量增加;感光基因标记 CRX、NRL 和 REC 下调。在 4-OHT 诱导的损伤后用 HB-EGF 培养器官组织 7 天,TUNEL 阳性细胞的数量明显减少,Ki67 阳性细胞的数量以及 PAX6 和 NOTCH1 基因的表达略有增加。目前的研究结果表明,用4-OHT处理人ESC衍生的视网膜器官组织可用作体外视网膜变性的模型。此外,用HB-EGF处理人视网膜器官组织可增加增殖的Müller细胞,但仅在4-OHT诱导的损伤之后,这可能是Muller对感光细胞损伤反应的一种迹象。进一步的研究旨在确定可能诱导Müller细胞介导的人类视网膜再生的因素,从而帮助开发治疗视网膜变性的疗法。
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