Lipoproteins in Cardiovascular Calcification: Potential Targets and Challenges

Previously considered a degenerative process, cardiovascular calcification is now established as an active process that is regulated in several ways by lipids, phospholipids, and lipoproteins. These compounds serve many of the same functions in vascular and valvular calcification as they do in skeletal bone calcification. Hyperlipidemia leads to accumulation of lipoproteins in the subendothelial space of cardiovascular tissues, which leads to formation of mildly oxidized phospholipids, which are known bioactive factors in vascular cell calcification. One lipoprotein of particular interest is Lp(a), which showed genome-wide significance for the presence of aortic valve calcification and stenosis. It carries an important enzyme, autotaxin, which produces lysophosphatidic acid (LPA), and thus has a key role in inflammation among other functions. Matrix vesicles, extruded from the plasma membrane of cells, are the sites of initiation of mineral formation. Phosphatidylserine, a phospholipid in the membranes of matrix vesicles, is believed to complex with calcium and phosphate ions, creating a nidus for hydroxyapatite crystal formation in cardiovascular as well as in skeletal bone mineralization. This review focuses on the contributions of lipids, phospholipids, lipoproteins, and autotaxin in cardiovascular calcification, and discusses possible therapeutic targets

Hyperphosphatemia-induced nanocrystals upregulate the expression of bone morphogenetic protein-2 and osteopontin genes in mouse smooth muscle cells in vitro

Vascular calcification, which contributes to cardiovascular disease in patients with uremic hyperphosphatemia, is associated with vascular cell expression of osteogenic genes, including bone morphogenetic protein (BMP)-2 and osteopontin (OPN). High inorganic phosphate levels in vitro stimulate the osteogenic conversion of smooth muscle cells; however, the mechanism governing this is not clear. We found that high-phosphate medium increased the expression of BMP-2 and OPN in mouse smooth muscle cells in culture. However, this effect was lost in the presence of the mineralization inhibitor, pyrophosphate, suggesting a contribution of calcium phosphate crystals. Addition of 1–2 mmol/l phosphate alone to growth medium was sufficient to induce nanosized crystals after 1 day at 37 °C. Isolated crystals were about 160 nm in diameter and had a calcium to phosphate ratio of 1.35, consistent with the hydroxyapatite precursor octacalcium phosphate. Nanocrystal formation increased fourfold in the absence of serum, was blocked by fetuin-A, and was dependent on time and on the concentrations of phosphate and calcium. Purified synthetic hydroxyapatite nanocrystals and isolated high-phosphate-induced nanocrystals, but not nanocrystal-free high-phosphate medium, also induced BMP-2 and OPN. Thus, our results suggest that BMP-2 and OPN are induced by calcium phosphate nanocrystals, rather than soluble phosphate. This mechanism may contribute, in part, to hyperphosphatemia-related vascular cell differentiation and calcification

Vitamin D3 supplementation of a high fat high sugar diet ameliorates prediabetic phenotype in female LDLR–/–and LDLR+/+mice

© 2017 The Authors. Immunity, Inflammation and Disease Published by John Wiley & Sons Ltd. INTRODUCTION: Fatty liver disease is prevalent in populations with high caloric intake. Nutritherapeutic approaches are being considered, such as supplementary Vitamin D 3 , to improve aspects of metabolic syndrome, namely fatty liver disease, hyperlipidemia, and insulin resistance associated with obesity. METHODS: We analyzed female LDLR -/- and LDLR +/+ mice on a 10-week diabetogenic diet for markers of fatty liver disease, metabolic strain, and inflammation. RESULTS: The groups on a high fat high sugar diet with supplementary Vitamin D 3 , in comparison with the groups on a high fat high sugar diet alone, showed improved transaminase levels, significantly less hypertriglyceridemia and hyperinsulinemia, and histologically, there was less pericentral hepatic steatosis. Levels of non-esterified fatty acids and lipid peroxidation products were significantly lower in the group supplemented with additional Vitamin D 3 , as were systemic markers of inflammation (serum endotoxin and IL-6). M2 macrophage phenotype predominated in the group supplemented with additional Vitamin D 3 . Beneficial changes were observed as early as five weeks’ supplementation with Vitamin D 3 and extended to restoration of high fat high sugar diet induced decrease of bone mineral density. CONCLUSION: In summary, Vitamin D 3 was a significantly beneficial dietary additive to blunt a prediabetic phenotype in diet-induced obesity of female LDLR -/- and LDLR +/+ mice

Patterns of periodic holes created by increased cell motility

The reaction and diffusion of morphogens is a mechanism widely used to explain many spatial patterns in physics, chemistry and developmental biology. However, because experimental control is limited in most biological systems, it is often unclear what mechanisms account for the biological patterns that arise. Here, we study a biological model of cultured vascular mesenchymal cells (VMCs), which normally self-organize into aggregates that form into labyrinthine configurations. We use an experimental control and a mathematical model that includes reacting and diffusing morphogens and a third variable reflecting local cell density. With direct measurements showing that cell motility was increased ninefold and threefold by inhibiting either Rho kinase or non-muscle myosin-II, respectively, our experimental results and mathematical modelling demonstrate that increased motility alters the multicellular pattern of the VMC cultures, from labyrinthine to a pattern of periodic holes. These results suggest implications for the tissue engineering of functional replacements for trabecular or spongy tissue such as endocardium and bone

Activating transcription factor-4 promotes mineralization in vascular smooth muscle cells

Emerging evidence indicates that upregulation of the ER stress–induced pro-osteogenic transcription factor ATF4 plays an important role in vascular calcification, a common complication in patients with aging, diabetes, and chronic kidney disease (CKD). In this study, we demonstrated the pathophysiological role of ATF4 in vascular calcification using global Atf4 KO, smooth muscle cell–specific (SMC-specific) Atf4 KO, and transgenic (TG) mouse models. Reduced expression of ATF4 in global ATF4-haplodeficient and SMC-specific Atf4 KO mice reduced medial and atherosclerotic calcification under normal kidney and CKD conditions. In contrast, increased expression of ATF4 in SMC-specific Atf4 TG mice caused severe medial and atherosclerotic calcification. We further demonstrated that ATF4 transcriptionally upregulates the expression of type III sodium-dependent phosphate cotransporters (PiT1 and PiT2) by interacting with C/EBPβ. These results demonstrate that the ER stress effector ATF4 plays a critical role in the pathogenesis of vascular calcification through increased phosphate uptake in vascular SMCs

Peroxisome proliferator-activated receptor activators modulate the osteoblastic maturation of MC3T3-E1 preosteoblasts

AbstractThe reduced bone mineral density (BMD) observed in osteoporosis results, in part, from reduced activity of bone-forming osteoblasts. We examined the effect of peroxisome proliferator-activated receptor (PPAR) activators on MC3T3-E1 preosteoblast maturation. Activators of PPARα, δ and γ induced alkaline phosphatase activity, matrix calcification and the expression of osteoblast genes as determined by reverse transcriptase-polymerase chain reaction. However, at relatively high concentrations of the specific PPARγ ligands, ciglitazone and troglitazone, maturation was inhibited. PPARα, δ and γ1 were expressed in MC3T3-E1 cells. PPARγ1 mRNA and protein levels were induced early during osteoblastic maturation. We speculate that endogenous and pharmacological PPAR activators may affect BMD by modulating osteoblastic maturation

Bone Density and Hyperlipidemia: The T-lymphocyte Connection

Osteoporosis, which contributes to morbidity and mortality, often coexists with cardiovascular disease, especially atherosclerosis. We have reported recently that in vitro exposure of human T-lymphocytes to oxidized lipids induced expression of a key osteoclastogenic cytokine, receptor activator of NF-κB ligand (RANKL). Our previous studies have shown that mice fed an atherogenic high-fat diet developed osteopenia and that bone marrow preosteoclasts from these hyperlipidemic mice have increased osteoclastic potential. To investigate the role of T-lymphocytes in the diet-induced bone loss, C57BL/6 mice were fed either chow or a high-fat diet, and bone parameters and T-lymphocyte activation were assessed at 6 and 11 months. Consistent with our previous findings, peripheral quantitative computed tomographic (pQCT) analysis showed that mice in the high-fat group had lower bone mineral content than mice in the chow group. Furthermore, histomorphometric analysis showed decreased structural parameters in the high-fat group. Coculture studies showed that bone marrow cells isolated from the high-fat group, which contained increased levels of activated memory T-lymphocytes compared with bone marrow cells from the chow mice, supported osteoclastic differentiation of RAW 264.7 cells. Additionally, RANKL expression was upregulated significantly in the T-lymphocytes isolated from the bone marrow of the high-fat group. Splenic T-lymphocytes isolated from the high-fat group also had increased expression of transcripts for the receptor for oxidized lipids (LOX-1) as well as for inflammatory and osteoclastogenic cytokines, including RANKL, interleukin 6 (IL-6), tumor necrosis factor α (TNF-α), IL-1β, and interferon γ (IFN-γ). Together these findings suggest that T-lymphocytes play a key role in the osteoclastogenesis induced by a high-fat diet and may contribute to the bone loss associated with diet-induced osteopenia. © 2010 American Society for Bone and Mineral Research