Hydrogen sulfide prevents arterial medial calcification in rats with diabetic nephropathy

Background Arterial medial calcification (AMC) is associated with a high incidence of cardiovascular risk in patients with type 2 diabetes and chronic kidney disease. Here, we tested whether hydrogen sulfide (H2S) can prevent AMC in rats with diabetic nephropathy (DN). Methods DN was induced by a single injection of streptozotocin and high-fat diet (45% kcal as fat) containing 0.75% adenine in Sprague–Dawley rats for 8 weeks. Results Rats with DN displayed obvious calcification in aorta, and this was significantly alleviated by Sodium Hydrosulfide (NaHS, a H2S donor, 50 μmol/kg/day for 8 weeks) treatment through decreasing calcium and phosphorus content, ALP activity and calcium deposition in aorta. Interestingly, the main endogenous H2S generating enzyme activity and protein expression of cystathionine-γ-lyase (CSE) were largely reduced in the arterial wall of DN rats. Exogenous NaHS treatment restored CSE activity and its expression, inhibited aortic osteogenic transformation by upregulating phenotypic markers of smooth muscle cells SMα-actin and SM22α, and downregulating core binding factor α-1 (Cbfα-1, a key factor for bone formation), protein expressions in rats with DN when compared to the control group. NaHS administration also significantly reduced Stat3 activation, cathepsin S (CAS) activity and TGF-β1 protein level, and improved aortic elastin expression. Conclusions H2S may have a clinical significance for treating AMC in people with DN by reducing Stat3 activation, CAS activity, TGF-β1 level and increasing local elastin level.

Regulation of mineralisation in bone and vascular tissue: a comparative review

Biomineralisation, the deposition of mineral onto a matrix, can be both a physiological and pathological process. Bone formation involves the secretion of an extracellular matrix (ECM) by osteoblasts and subsequent mineralisation of that matrix. It is regulated by a number of local and systemic factors and is necessary for maintenance of normal bone health. Conversely, mineralisation (or calcification) of soft tissues, including the vasculature, is detrimental to that tissue, leading to diseases such as arterial medial calcification (AMC). The mechanisms underlying AMC development are not fully defined, though it is thought that vascular smooth muscle cells (VSMCs) drive this complex, cell-mediated process. Similarly, AMC is regulated by a variety of enzymes and molecules, many of which have already been implicated in the regulation of bone mineralisation. This review will provide an overview of the similar, and sometimes opposing effects of these signalling molecules on the regulation of bone mineralisation and AMC.

Loss of PKCα increases arterial medial calcification in a uremic mouse model of chronic kidney disease

Arterial medial calcification is an independent risk factor for mortality in chronic kidney disease. We previously reported that knock-down of PKCα expression increases high phosphate-induced mineral deposition by vascular smooth muscle cells in vitro. This new study tests the hypothesis that PKCα regulates uremia-induced medial calcification in vivo. Female wild-type and PKCα−/− mice underwent a two-stage subtotal nephrectomy and were fed a high phosphate diet for 8 weeks. X-ray micro computed tomography demonstrated that uremia-induced medial calcification was increased in the abdominal aorta and aortic arch of PKCα−/− mice compared to wild-types. Blood urea nitrogen was also increased in PKCα−/− mice compared to wild-types; there was no correlation between blood urea nitrogen and calcification in PKCα−/− mice. Phosphorylated SMAD2 immunostaining was detected in calcified aortic arches from uremic PKCα−/− mice; the osteogenic marker Runx2 was also detected in these areas. No phosphorylated SMAD2 staining were detected in calcified arches from uremic wild-types. PKCα knock-down increased TGF-β1-induced SMAD2 phosphorylation in vascular smooth muscle cells in vitro, whereas the PKCα activator prostratin decreased SMAD2 phosphorylation. In conclusion, loss of PKCα increases uremia-induced medial calcification. The PKCα/TGF-β signaling axis could therefore represent a new therapeutic target for arterial medial calcification in chronic kidney disease.

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

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.