Endothelial-to-mesenchymal transition (EndoMT), the process wherein endothelial cells lose endothelial identity and adopt mesenchymal-like phenotypes, constitutes a critical contributor to cardiac fibrosis. The phenotypic plasticity of endothelial cells can be intricately shaped by alteration of metabolic pathways, but how endothelial cells adjust cellular metabolism to drive EndoMT is incompletely understood. Here, we identified 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) as a critical driver of EndoMT via triggering abnormal glycolysis and compromising mitochondrial respiration. Pharmacological suppression of PFKFB3 with salvianolic acid C (SAC), a phenolic compound derived from Salvia miltiorrhiza, attenuates EndoMT and fibrotic response. PFKFB3-haplodeficiency recapitulates the anti-EndoMT effect of SAC while PFKFB3-overexpression augments the magnitude of EndoMT and exacerbates cardiac fibrosis. Mechanistically, PFKFB3-driven glycolysis compromises cytoplasmic nicotinamide adenine dinucleotide phosphate (reduced form, NADPH) production via hijacking glucose flux from pentose phosphate pathway. Efflux of mitochondrial NADPH through isocitrate/α-ketoglutarate shuttle replenishes cytoplasmic NADPH pool but meanwhile impairs mitochondrial respiration by hampering mitochondrial iron-sulfur cluster biosynthesis. SAC disrupts PFKFB3 stability by accelerating its degradation and thus maintains metabolic homeostasis in endothelial cells, underlying its anti-EndoMT effects. These findings for the first time identify the critical role of PFKFB3 in triggering EndoMT by driving abnormal glycolysis in endothelial cells, and also highlight the therapeutic potential for pharmacological intervention of PFKFB3 (with SAC or other PFKFB3 inhibitors) to combat EndoMT-associated fibrotic responses via metabolic regulation.

内皮-间充质转化 (EndoMT)，即内皮细胞失去内皮特性并采用间充质样表型的过程，是心脏纤维化的关键因素。内皮细胞的表型可塑性可以通过代谢途径的改变而错综复杂地形成，但内皮细胞如何调节细胞代谢以驱动 EndoMT 尚不完全清楚。在这里，我们确定 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) 通过触发异常糖酵解和损害线粒体呼吸作为 EndoMT 的关键驱动因素。丹酚酸 C (SAC) 对 PFKFB3 的药理学抑制作用，丹酚酸 C (SAC) 是一种源自丹参的酚类化合物, 减弱 EndoMT 和纤维化反应。PFKFB3-单倍体缺陷概括了 SAC 的抗 EndoMT 作用，而 PFKFB3-过表达增加了 EndoMT 的程度并加剧了心脏纤维化。从机制上讲，PFKFB3 驱动的糖酵解通过劫持来自戊糖磷酸途径的葡萄糖通量来损害细胞质烟酰胺腺嘌呤二核苷酸磷酸盐（还原形式，NADPH）的产生。线粒体 NADPH 通过异柠檬酸/α-酮戊二酸穿梭流出补充细胞质 NADPH 池，但同时通过阻碍线粒体铁硫簇生物合成而损害线粒体呼吸。SAC 通过加速其降解来破坏 PFKFB3 的稳定性，从而维持内皮细胞的代谢稳态，这是其抗 EndoMT 作用的基础。