Endothelial Drp1 Couples VEGF-induced Redox Signaling with Glycolysis Through Cysteine Oxidation to Drive Angiogenesis.

Angiogenesis plays a vital role for postnatal development and tissue repair following ischemia. Reactive oxygen species (ROS) generated by NADPH oxidases (NOXes) and mitochondria act as signaling molecules that promote angiogenesis in endothelial cells (ECs) which mainly relies on aerobic glycolysis for ATP production. However, the connections linking redox signaling with glycolysis are not well understood. The GTPase Drp1 is a member of the dynamin superfamily that moves from cytosol to mitochondria through posttranslational modifications to induce mitochondrial fission. The role of Drp1 in ROS-dependent VEGF signaling and angiogenesis in ECs has not been previously described. Here, we identify an unexpected function of endothelial Drp1 as a redox sensor, transmitting VEGF-induced H 2 O 2 signals to enhance glycolysis and angiogenesis. Loss of Drp1 expression in ECs inhibited VEGF-induced angiogenic responses. Mechanistically, VEGF rapidly induced the NOX4-dependent sulfenylation (CysOH) of Drp1 on Cys 644 , promoting disulfide bond formation with the metabolic kinase AMPK and subsequent sulfenylation of AMPK at Cys 299 / 304 via the mitochondrial fission-mitoROS axis. This cysteine oxidation of AMPK, in turn, enhanced glycolysis and angiogenesis. In vivo , mice with EC-specific Drp1 deficiency or CRISPR/Cas9-engineered "redox-dead" (Cys to Ala) Drp1 knock-in mutations exhibited impaired retinal angiogenesis and post-ischemic neovascularization. Our findings uncover a novel role for endothelial Drp1 in linking VEGF-induced mitochondrial redox signaling to glycolysis through a cysteine oxidation-mediated Drp1-AMPK redox relay, driving both developmental and reparative angiogenesis.

Single-cell transcriptome analyses reveal microglia types associated with proliferative retinopathy.

Pathological angiogenesis is a major cause of irreversible blindness in individuals of all age groups with proliferative retinopathy (PR). Mononuclear phagocytes (MPs) within neovascular areas contribute to aberrant retinal angiogenesis. Due to their cellular heterogeneity, defining the roles of MP subsets in PR onset and progression has been challenging. Here, we aimed to investigate the heterogeneity of microglia associated with neovascularization and to characterize the transcriptional profiles and metabolic pathways of proangiogenic microglia in a mouse model of oxygen-induced PR (OIR). Using transcriptional single-cell sorting, we comprehensively mapped all microglia populations in retinas of room air (RA) and OIR mice. We have unveiled several unique types of PR-associated microglia (PRAM) and identified markers, signaling pathways, and regulons associated with these cells. Among these microglia subpopulations, we found a highly proliferative microglia subset with high self-renewal capacity and a hypermetabolic microglia subset that expresses high levels of activating microglia markers, glycolytic enzymes, and proangiogenic Igf1. IHC staining shows that these PRAM were spatially located within or around neovascular tufts. These unique types of microglia have the potential to promote retinal angiogenesis, which may have important implications for future treatment of PR and other pathological ocular angiogenesis-related diseases.

Protein kinase N2 connects blood flow with NO production in a double AKT.

Shear stress is an important regulator of blood flow, and luminal endothelial cells (ECs) sense increases in frictional forces and respond with an appropriate release of vasoactive mediators. In this issue of the JCI, Jin et al. identified a mechanism by which ECs respond to shear stress with endothelial NOS (eNOS) activation and NO release. The authors showed that PKN2 was activated by fluid shear stress and contributed to eNOS activation via a double play - indirect phosphorylation at serine 1177 (S1177) via AKT and direct phosphorylation of the S1179 site. Phosphorylation of both sites individually increased eNOS activity, but together they had an additive effect. In sum, these findings reveal exciting details about how shear stress regulates eNOS and have important implications for blood flow and blood pressure.