Hypoxia Stimulates Low-Density Lipoprotein Receptor–Related Protein-1 Expression Through Hypoxia-Inducible Factor-1α in Human Vascular Smooth Muscle Cells

Abstract

Objective-Hypoxia is considered a key factor in the progression of atherosclerotic lesions. Low-density lipoprotein receptor-related protein (LRP1) plays a pivotal role in the vasculature. The aim of this study was to investigate the effect of hypoxia on LRP1 expression and function in vascular smooth muscle cells (VSMC) and the role of hypoxia-inducible factor-␣ (HIF-1␣). Methods and Results-Real-time polymerase chain reaction and Western blot analysis demonstrated that hypoxia (1% O 2 ) time-dependently induced LRP1 mRNA (maximum levels at 1 to 2 hours) and protein expression (maximum levels at 12 to 24 hours). The delayed hypoxic upregulation of LRP1 protein versus mRNA may be explained by the long half-life of LRP1 protein. Luciferase assays demonstrated that hypoxia and HIF-1␣ overaccumulation induced LRP1 promoter activity and that 2 consensus hypoxia response element sites located at Ϫ1072/Ϫ1069 and Ϫ695/Ϫ692 participate in the induction. Chromatin immunoprecipitation showed the in vivo binding of HIF-1␣ to LRP1 promoter in hypoxic VSMC. Hypoxia effects on LRP1 protein expression were functionally translated into an increased cholesteryl ester (CE) accumulation from aggregated low-density lipoprotein (agLDL) uptake. The blockade of HIF-1␣ expression inhibited the upregulatory effect of hypoxia on LRP1 expression and agLDL-derived intracellular CE overaccumulation, suggesting that both LRP1 overexpression and CE overaccumulation in hypoxic vascular cells are dependent on HIF-1␣. Immunohistochemical analysis showed the colocalization of LRP1 and HIF-1␣ in vascular cells of human advanced atherosclerotic plaques. H ypoxia plays a pivotal role in the pathophysiology of cancer, myocardial infarction, and atherosclerosis. Arterial wall thickness associated with atherosclerotic plaque progression reduces the oxygen supply in certain areas of the vascular intima. 1,2 Hypoxia is present in human advanced atherosclerotic lesions and correlates with the presence of angiogenesis and thrombus. 3,4 Most of the cells in hypoxic areas of atherosclerotic plaques respond by altering the expression of genes involved in vasculogenesis, angiogenesis, inflammation, and lipid deposition. [5][6][7][8][9][10][11] These genes are activated through hypoxia-inducible factor-1 complex (HIF-1). The HIF-1␣ subunit is subjected to quick oxygendependent proteasomal degradation through hydroxylation of proline residues by hydroxylases. [12][13][14] Under low oxygen tensions, HIF-1␣ accumulates and heterodimerizes with the constitutively expressed beta subunit, allowing the HIF-1 complex to bind to hypoxia response element (HRE) sequences in target promoters. 15 It has been reported that HIF-1␣ accumulation in macrophages promotes foam cell formation and atherosclerosis. 16 In experimental models of hypercholesterolemia and hypertension, HIF-1␣ is also associated with vascular smooth muscle cells (VSMC). 17 30 -32 It is unknown whether hypoxia may modulate LRP1 expression in human atherosclerotic plaques and, if so, which transcription ...

Hypoxia Induces Metalloproteinase-9 Activation and Human Vascular Smooth Muscle Cell Migration Through Low-Density Lipoprotein Receptor–Related Protein 1–Mediated Pyk2 Phosphorylation

“…Hypoxia, however, strongly increased pPyk2 staining in hVSMCs ( Figure 1E). As previously shown in hVSMCs, 16 HIF-1α levels were significantly increased at 4 and 8 hours of mVSMCs ( Figure IE …”

“…Previous studies from our group showed that hypoxia strongly upregulated LRP1 expression in hVSMCs. 16 Here, we show that LRP1 expression and Pyk2 phosphorylation were concomitantly upregulated by hypoxia for ≤32 hours in hVSMCs. According to our results, hypoxia modulates LRP1 and pPyk2 levels in a specific manner depending on species and cell type.…”

“…We previously showed that hypoxia-induced LRP1 overexpression is mediated by HIF-1α in hVSMCs and HL-1 cells. 16,17 HIF-1α expression depends on intracellular calcium concentrations that rapidly and persistently increase in response to hypoxia. 31,32 Hypoxia-induced Pyk2 phosphorylation has been associated to a hypoxia-induced increase in calcium cytosolic levels.…”

High glucose facilitated endothelial heparanase transfer to the cardiomyocyte modifies its cell death signature

“…HIF-1a is a known factor that can induce LRP1 expression in cardiomyocytes 42 and in other cell types. [43][44][45][46] These effects were lost following chronic diabetes, and could contribute to the development of cardiomyopathy in these animals. The disappearance of LRP1, with prolonged duration of diabetes, may be related to a further attenuation of myocytes pooled from four independent experiments.…”

Aggregated Low-Density Lipoprotein Induces LRP1 Stabilization Through E3 Ubiquitin Ligase CHFR Downregulation in Human Vascular Smooth Muscle Cells

“…[5][6][7] Uptake of agLDL through LRP1 allows high-intracellular CE accumulation not only because of its high capacity to bind and internalize agLDL, but also because LRP1 is transcriptionally upregulated by intracellular cholesterol. Our group reported that LRP1 is upregulated at transcriptional level by hypercholesterolemia through sterol regulatory element-binding proteins (SREBP) downregulation in human VSMC, [8][9][10] and by hypoxia through hypoxia-inducible factor 1α upregulation in human VSMC 11 and cardiomyocytes. 12,13 Other groups have reported that insulin promotes the presence of LRP1 in the plasma membrane without influencing mRNA expression levels.…”