Abnormal Lysosomal Positioning and Small Extracellular Vesicle Secretion in Arterial Stiffening and Calcification of Mice Lacking Mucolipin 1 Gene

Abstract

Recent studies have shown that arterial medial calcification is mediated by abnormal release of exosomes/small extracellular vesicles from vascular smooth muscle cells (VSMCs) and that small extracellular vesicle (sEV) secretion from cells is associated with lysosome activity. The present study was designed to investigate whether lysosomal expression of mucolipin-1, a product of the mouse Mcoln1 gene, contributes to lysosomal positioning and sEV secretion, thereby leading to arterial medial calcification (AMC) and stiffening. In Mcoln1 −/− mice, we found that a high dose of vitamin D (Vit D; 500,000 IU/kg/day) resulted in increased AMC compared to their wild-type littermates, which was accompanied by significant downregulation of SM22-α and upregulation of RUNX2 and osteopontin in the arterial media, indicating a phenotypic switch to osteogenic. It was also shown that significantly decreased co-localization of lysosome marker (Lamp-1) with lysosome coupling marker (Rab 7 and ALG-2) in the aortic wall of Mcoln1 −/− mice as compared to their wild-type littermates. Besides, Mcoln1 −/− mice showed significant increase in the expression of exosome/ sEV markers, CD63, and annexin-II (AnX2) in the arterial medial wall, accompanied by significantly reduced co-localization of lysosome marker (Lamp-1) with multivesicular body (MVB) marker (VPS16), suggesting a reduction of the lysosome-MVB interactions. In the plasma of Mcoln1 −/− mice, the number of sEVs significantly increased as compared to the wild-type littermates. Functionally, pulse wave velocity (PWV), an arterial stiffening indicator, was found significantly increased in Mcoln1 −/− mice, and Vit D treatment further enhanced such stiffening. All these data indicate that the Mcoln1 gene deletion in mice leads to abnormal lysosome positioning and increased sEV secretion, which may contribute to the arterial stiffness during the development of AMC.

The Protective Effects of the Autophagic and Lysosomal Machinery in Vascular and Valvular Calcification: A Systematic Review

“…However, lysosomal function was also shown to promote VCN by degrading the anti-calcifying protein granzyme B. Knocking out Mcoln1 enhanced VCN both, in vivo and in vitro by impairing lysosomal trafficking, and consequentially enhancing secretion of pro-calcific small extracellular vesicles (sEVs) ( Table 1 ) [ 79 ]. Asah1 gene deletion also aggravated VCN and decreased TRPML1 activity, leading to impaired lysosomal trafficking and enhanced pro-calcific sEV secretion [ 80 ].…”

“…Knocking-out Mcoln1 , which is the gene that encodes TRPML1, resulted in increased VCN both in vitro and in vivo. Moreover, increased secretion of pro-calcific sEVs was reported in the absence of TRPML1 [ 79 ]. sEVs contain a variety of cargoes such as proteins and lipids that can chelate Pi and calcium.…”

The Role of Vitamin D in Modulating Mesenchymal Stem Cells and Endothelial Progenitor Cells for Vascular Calcification

“…Contrary evidence also demonstrated the vitamin D might stimulate vascular calcification by modulating the expression of parathyroid hormone-related peptide or the receptor activator of nuclear factor kappa-B ligand/osteoprotegerin of VSMC [42,43]. The pharmacologic or the supraphysiologic concentrations of active or nutritional vitamin D might contribute to the vascular calcification in vivo studies [44,45]. Therefore, vitamin D has rather complex effects on calcification from the aspect of VSMC, and more advanced studies are needed to elucidate the role of vitamin D in vascular calcification.…”

The Progress of Advanced Ultrasonography in Assessing Aortic Stiffness and the Application Discrepancy between Humans and Rodents

“…They found that PWV was significantly increased with age in these mice [ 57 ]. PWV was also found to be increased dramatically in Mcoln1 -/- mice compared to their wild-type littermates, and Vitamin D treatment further enhanced such stiffening in these mice [ 11 ].…”

“…It is also known as an independent risk factor of cardiovascular complications and has been considered as a predictor of cardiovascular outcomes in clinical [ 8 , 9 , 10 ]. As a non-invasive method, ultrasonography has been used in clinical diagnosis for aortic stiffness in many aortic disorders such as atherosclerotic degeneration and aortic aneurysms through measuring the wall-thickness, diameter, structural defects, blood flow velocity, and other pathological changes in the aorta [ 7 , 11 , 12 , 13 ]. The developed techniques used in patients also provide powerful tools for basic science research in different animal models, particularly the large animal models such as monkeys, swine, canines, and horses [ 14 , 15 , 16 , 17 , 18 , 19 ].…”