Department of Surgery, Alberta Diabetes Institute, University of Alberta, Edmonton, AB, T6G 2E1, Canada.
Department of Biochemistry, Faculty of Science, The Maharaja Sayajirao University of Baroda, Vadodara, 390021, Gujarat, India.
Sci Rep. 2022 Dec 13;12(1):21496. doi: 10.1038/s41598-022-25405-w.
Poly(ADP-ribose) polymerase-1 (PARP1), a fundamental DNA repair enzyme, is known to regulate β cell death, replication, and insulin secretion. PARP1 knockout (KO) mice are resistant to diabetes, while PARP1 overactivation contributes to β cell death. Additionally, PARP1 inhibition (PARPi) improves diabetes complications in patients with type-2 diabetes. Despite these beneficial effects, the use of PARP1 modulating agents in diabetes treatment is largely neglected, primarily due to the poorly studied mechanistic action of PARP1 catalytic function in human β cell development. In the present study, we evaluated PARP1 regulatory action in human β cell differentiation using the human pancreatic progenitor cell line, PANC-1. We surveyed islet census and histology from PARP1 wild-type versus KO mice pancreas in a head-to-head comparison with PARP1 regulatory action for in-vitro β cell differentiation following either PARP1 depletion or its pharmacological inhibition in PANC-1-differentiated islet cells. shRNA mediated PARP1 depleted (SiP) and shRNA control (U6) PANC-1 cells were differentiated into islet-like clusters using established protocols. We observed complete abrogation of new β cell formation with absolute PARP1 depletion while its inhibition using the potent inhibitor, PJ34, promoted the endocrine β cell differentiation and maturation. Immunohistochemistry and immunoblotting for key endocrine differentiation players along with β cell maturation markers highlighted the potential regulatory action of PARP1 and augmented β cell differentiation due to direct interaction of unmodified PARP1 protein elicited p38 MAPK phosphorylation and Neurogenin-3 (Ngn3) re-activation. In summary, our study suggests that PARP1 is required for the proper development and differentiation of human islets. Selective inhibition with PARPi can be an advantage in pushing more insulin-producing cells under pathological conditions and delivers a potential for pilot clinical testing for β cell replacement cell therapies for diabetes.
聚（ADP-核糖）聚合酶 1（PARP1）是一种基本的 DNA 修复酶，已知其可调节β细胞死亡、复制和胰岛素分泌。PARP1 敲除（KO）小鼠对糖尿病具有抗性，而 PARP1 过度激活则导致β细胞死亡。此外，PARP1 抑制（PARPi）可改善 2 型糖尿病患者的糖尿病并发症。尽管具有这些有益作用，但 PARP1 调节因子在糖尿病治疗中的应用在很大程度上被忽视了，主要是因为 PARP1 催化功能在人类β细胞发育中的作用机制研究不足。在本研究中，我们使用人胰腺祖细胞系 PANC-1 评估了 PARP1 在人类β细胞分化中的调节作用。我们在头对头比较中调查了 PARP1 野生型与 KO 小鼠胰腺中的胰岛计数和组织学，并比较了 PARP1 对 PANC-1 分化的胰岛细胞体外β细胞分化的调节作用，方法是 PARP1 耗竭或其药理学抑制。通过 shRNA 介导的 PARP1 耗竭（SiP）和 shRNA 对照（U6）PANC-1 细胞使用已建立的方案分化为胰岛样簇。我们观察到，在绝对 PARP1 耗竭的情况下，新β细胞的形成完全被阻断，而使用强效抑制剂 PJ34 抑制其活性则促进了内分泌β细胞的分化和成熟。免疫组织化学和免疫印迹分析关键的内分泌分化因子以及β细胞成熟标志物突出了 PARP1 的潜在调节作用，并由于未经修饰的 PARP1 蛋白的直接相互作用引发了 p38 MAPK 磷酸化和神经基因 3（Ngn3）的再激活，增强了β细胞的分化。总之，我们的研究表明 PARP1 是人类胰岛正常发育和分化所必需的。选择性抑制 PARPi 可以在病理条件下产生更多的胰岛素产生细胞，并为糖尿病的β细胞替代细胞治疗的初步临床测试提供了潜力。
