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Abstract：
BACKGROUND: Glucose-6-phosphate dehydrogenase (G6PD) plays an important role in vascular smooth muscle cell (VSMC) phenotypic switching, which is an early pathogenic event in various vascular remodeling diseases (VRDs). However, the underlying mechanism is not fully understood.
METHODS: An IP‒LC‒MS/MS assay was conducted to identify new binding partners of G6PD involved in the regulation of VSMC phenotypic switching under platelet-derived growth factor-BB (PDGF-BB) stimulation. Co-IP, GST pull-down, and immunofluorescence colocalization were employed to clarify the interaction between G6PD and voltage-dependent anion-selective channel protein 1 (VDAC1). The molecular mechanisms involved were elucidated by examining the interaction between VDAC1 and apoptosis-related biomarkers, as well as the oligomerization state of VDAC1.
RESULTS: The G6PD level was significantly elevated and positively correlated with the synthetic characteristics of VSMCs induced by PDGF-BB. We identified VDAC1 as a novel G6PD-interacting molecule essential for apoptosis. Specifically, the G6PD-NTD region was found to predominantly contribute to this interaction. G6PD promotes VSMC survival and accelerates vascular neointimal hyperplasia by inhibiting VSMC apoptosis. Mechanistically, G6PD interacts with VDAC1 upon stimulation with PDGF-BB. By competing with Bax for VDAC1 binding, G6PD reduces VDAC1 oligomerization and counteracts VDAC1-Bax-mediated apoptosis, thereby accelerating neointimal hyperplasia.
CONCLUSIONS: Our study showed that the G6PD-VDAC1-Bax axis is a vital switch in VSMC apoptosis and is essential for VSMC phenotypic switching and neointimal hyperplasia, providing mechanistic insight into early VRDs.
METHODS: An IP‒LC‒MS/MS assay was conducted to identify new binding partners of G6PD involved in the regulation of VSMC phenotypic switching under platelet-derived growth factor-BB (PDGF-BB) stimulation. Co-IP, GST pull-down, and immunofluorescence colocalization were employed to clarify the interaction between G6PD and voltage-dependent anion-selective channel protein 1 (VDAC1). The molecular mechanisms involved were elucidated by examining the interaction between VDAC1 and apoptosis-related biomarkers, as well as the oligomerization state of VDAC1.
RESULTS: The G6PD level was significantly elevated and positively correlated with the synthetic characteristics of VSMCs induced by PDGF-BB. We identified VDAC1 as a novel G6PD-interacting molecule essential for apoptosis. Specifically, the G6PD-NTD region was found to predominantly contribute to this interaction. G6PD promotes VSMC survival and accelerates vascular neointimal hyperplasia by inhibiting VSMC apoptosis. Mechanistically, G6PD interacts with VDAC1 upon stimulation with PDGF-BB. By competing with Bax for VDAC1 binding, G6PD reduces VDAC1 oligomerization and counteracts VDAC1-Bax-mediated apoptosis, thereby accelerating neointimal hyperplasia.
CONCLUSIONS: Our study showed that the G6PD-VDAC1-Bax axis is a vital switch in VSMC apoptosis and is essential for VSMC phenotypic switching and neointimal hyperplasia, providing mechanistic insight into early VRDs.
摘要：
背景：葡萄糖-6-磷酸脱氢酶（G6PD）在血管平滑肌细胞（VSMC）表型转换中起重要作用，这是各种血管重塑疾病（VRD）的早期致病事件。然而,潜在的机制还没有完全理解。
方法：进行了IP-LC-MS/MS测定，以鉴定在血小板衍生生长因子-BB（PDGF-BB）刺激下参与调节VSMC表型转换的G6PD的新结合配偶体。共同IP,GST下拉式,和免疫荧光共定位用于阐明G6PD与电压依赖性阴离子选择性通道蛋白1（VDAC1）之间的相互作用。通过检查VDAC1与凋亡相关生物标志物之间的相互作用，阐明了所涉及的分子机制。以及VDAC1的低聚状态。
结果：G6PD水平显着升高，并与PDGF-BB诱导的VSMC的合成特性呈正相关。我们将VDAC1鉴定为凋亡必需的新型G6PD相互作用分子。具体来说,发现G6PD-NTD区域主要促成这种相互作用。G6PD通过抑制VSMC凋亡促进VSMC存活并加速血管新生内膜增生。机械上,在用PDGF-BB刺激时,G6PD与VDAC1相互作用。通过与Bax竞争VDAC1结合，G6PD减少VDAC1寡聚化，并抵消VDAC1-Bax介导的细胞凋亡，从而加速新生内膜增生。
结论：我们的研究表明，G6PD-VDAC1-Bax轴是VSMC凋亡的重要开关，对于VSMC表型转换和新内膜增生至关重要，提供对早期VRD的机械洞察。
方法：进行了IP-LC-MS/MS测定，以鉴定在血小板衍生生长因子-BB（PDGF-BB）刺激下参与调节VSMC表型转换的G6PD的新结合配偶体。共同IP,GST下拉式,和免疫荧光共定位用于阐明G6PD与电压依赖性阴离子选择性通道蛋白1（VDAC1）之间的相互作用。通过检查VDAC1与凋亡相关生物标志物之间的相互作用，阐明了所涉及的分子机制。以及VDAC1的低聚状态。
结果：G6PD水平显着升高，并与PDGF-BB诱导的VSMC的合成特性呈正相关。我们将VDAC1鉴定为凋亡必需的新型G6PD相互作用分子。具体来说,发现G6PD-NTD区域主要促成这种相互作用。G6PD通过抑制VSMC凋亡促进VSMC存活并加速血管新生内膜增生。机械上,在用PDGF-BB刺激时,G6PD与VDAC1相互作用。通过与Bax竞争VDAC1结合，G6PD减少VDAC1寡聚化，并抵消VDAC1-Bax介导的细胞凋亡，从而加速新生内膜增生。
结论：我们的研究表明，G6PD-VDAC1-Bax轴是VSMC凋亡的重要开关，对于VSMC表型转换和新内膜增生至关重要，提供对早期VRD的机械洞察。
