Abstract
While impairment of vascular homeostasis induced by hypercholesterolemia is the first step of cardiovascular diseases, the molecular mechanism behind such impairment is not well known. Here, we reported that high-cholesterol diet (HCD) induced defective vessel sprouting in zebrafish larvae. Electron transfer flavoprotein subunit α (ETFα) (encoded by the ETFA gene), a protein that mediates transfer of electrons from a series of mitochondrial flavoenzymes to the respiratory chain, was downregulated in HCD-fed zebrafish and in endothelial cells treated with oxidized low-density lipoprotein. Knockdown of ETFα with morpholino antisense oligonucleotides reproduced vascular sprouting defects in zebrafish larvae, while replenishing with exogeneous ETFA mRNA could successfully rescue these defects. ETFA knockdown in endothelial cells reduces cell migration, proliferation, and tube formation in vitro. Finally, knockdown of ETFA in endothelial cells also reduced fatty acid oxidation, oxygen consumption rate, and hypoxia-inducible factor-1α (HIF1α) protein levels. Taken together, we demonstrate that downregulation of ETFα is involved in hypercholesterolemia-induced defective vessel sprouting in zebrafish larvae via inhibition of endothelial proliferation and migration. The molecular mechanism behind this phenomenon is the decrease of HIF1α induced by downregulation of ETFα in endothelial cells. This work suggests that disturbance of ETFα-mediated oxygen homeostasis is one of the mechanisms behind hypercholesterolemia-induced vascular dysfunction.
Highlights
Hypercholesterolemia has been linked to cardiovascular disease (CVD) for decades, which could lead to premature atherosclerosis-based cardiovascular diseases and death, but the underlying mechanism is still far away from elucidation [1, 2]
Mitochondria are in the central position of endothelial metabolic homeostasis through supplying energy, signaling, and material needed for biosynthesis [11]
Our work found that persistent hypercholesterolemia in zebrafish larvae could induce downregulation of several mitochondrion- and metabolic-related enzymes, ETF α is one of them, and knockdown of it resulted to defective vessel growth in vivo and vitro
Summary
Hypercholesterolemia has been linked to cardiovascular disease (CVD) for decades, which could lead to premature atherosclerosis-based cardiovascular diseases and death, but the underlying mechanism is still far away from elucidation [1, 2]. The severity of deformities like vascular defects was positively correlated with the increase of cholesterol concentration in diet, which suggests that hypercholesterolemia compromises the vessel development. ITRAQ analysis of normal- and HCD-fed zebrafish larvae found a total of 63 differentially expressed proteins [3]. More than 12.2% of those 63 candidates were metabolic activities related, and 14.3% of them were mitochondrion located, which suggest that mitochondria are probably related to hypercholesterolemia and deformities. To reveal the relationship between mitochondrion function and hypercholesterolemia-related vascular deformities, we pick electron transfer flavoprotein-α (ETFα), one of the differentially expressed mitochondrial proteins in iTRAQ analysis, to carry out our investigation. To mimic the milder downregulation of ETFα induced by hypercholesterolemia but not severely demolished like MADD, we use morpholino to knockdown ETFA in zebrafish embryos to leave residual ETFA function. This work may shed light on the underlying link between hypercholesterolemia and CVD
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