Amirala Bakhshian Nik, Katherine Kaiser, Patrick Sun, Bohdan B Khomtchouk, Joshua D Hutcheson
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We hypothesized that caveolin-1 may play divergent roles in calcifying EV formation from vascular smooth muscle cells (VSMCs) and osteoblasts (HOBs).</p><p><strong>Methods: </strong>Primary human coronary artery VSMCs and osteoblasts were cultured for up to 28 days in an osteogenic media. CAV1 expression was knocked down using siRNA. Methyl β-cyclodextrin (MβCD) and a calpain inhibitor were used, respectively, to disrupt and stabilize the caveolar domains in VSMCs and HOBs.</p><p><strong>Results: </strong>CAV1 genetic variation demonstrates significant inverse relationships between bone-mineral density (BMD) and coronary artery calcification (CAC) across two independent epidemiological cohorts. Culture in osteogenic (OS) media increased calcification in HOBs and VSMCs. siRNA knockdown of CAV1 abrogated VSMC calcification with no effect on osteoblast mineralization. MβCD-mediated caveolae disruption led to a 3-fold increase of calcification in VSMCs treated with osteogenic media (<i>p</i> < 0.05) but hindered osteoblast mineralization (<i>p</i> < 0.01). Conversely, stabilizing caveolae by calpain inhibition prevented VSMC calcification (<i>p</i> < 0.05) without affecting osteoblast mineralization. There was no significant difference in CAV1 content between lipid domains from HOBs cultured in OS and control media.</p><p><strong>Conclusion: </strong>Our data indicate fundamental cellular-level differences in physiological and pathophysiological mineralization mediated by CAV1 dynamics. This is the first study to suggest that divergent mechanisms in calcifying EV formation may play a role in the calcification paradox.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s12195-023-00779-7.</p>","PeriodicalId":9687,"journal":{"name":"Cellular and molecular bioengineering","volume":null,"pages":null},"PeriodicalIF":2.3000,"publicationDate":"2023-08-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10550882/pdf/","citationCount":"0","resultStr":"{\"title\":\"Altered Caveolin-1 Dynamics Result in Divergent Mineralization Responses in Bone and Vascular Calcification.\",\"authors\":\"Amirala Bakhshian Nik, Katherine Kaiser, Patrick Sun, Bohdan B Khomtchouk, Joshua D Hutcheson\",\"doi\":\"10.1007/s12195-023-00779-7\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Introduction: </strong>Though vascular smooth muscle cells adopt an osteogenic phenotype during pathological vascular calcification, clinical studies note an inverse correlation between bone mineral density and arterial mineral-also known as the calcification paradox. Both processes are mediated by extracellular vesicles (EVs) that sequester calcium and phosphate. Calcifying EV formation in the vasculature requires caveolin-1 (CAV1), a membrane scaffolding protein that resides in membrane invaginations (caveolae). Of note, caveolin-1-deficient mice, however, have increased bone mineral density. We hypothesized that caveolin-1 may play divergent roles in calcifying EV formation from vascular smooth muscle cells (VSMCs) and osteoblasts (HOBs).</p><p><strong>Methods: </strong>Primary human coronary artery VSMCs and osteoblasts were cultured for up to 28 days in an osteogenic media. CAV1 expression was knocked down using siRNA. Methyl β-cyclodextrin (MβCD) and a calpain inhibitor were used, respectively, to disrupt and stabilize the caveolar domains in VSMCs and HOBs.</p><p><strong>Results: </strong>CAV1 genetic variation demonstrates significant inverse relationships between bone-mineral density (BMD) and coronary artery calcification (CAC) across two independent epidemiological cohorts. Culture in osteogenic (OS) media increased calcification in HOBs and VSMCs. siRNA knockdown of CAV1 abrogated VSMC calcification with no effect on osteoblast mineralization. MβCD-mediated caveolae disruption led to a 3-fold increase of calcification in VSMCs treated with osteogenic media (<i>p</i> < 0.05) but hindered osteoblast mineralization (<i>p</i> < 0.01). Conversely, stabilizing caveolae by calpain inhibition prevented VSMC calcification (<i>p</i> < 0.05) without affecting osteoblast mineralization. There was no significant difference in CAV1 content between lipid domains from HOBs cultured in OS and control media.</p><p><strong>Conclusion: </strong>Our data indicate fundamental cellular-level differences in physiological and pathophysiological mineralization mediated by CAV1 dynamics. 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引用次数: 0
摘要
引言：尽管血管平滑肌细胞在病理性血管钙化过程中具有成骨表型，但临床研究注意到骨密度和动脉矿物质之间存在负相关，也称为钙化悖论。这两个过程都是由细胞外小泡（EV）介导的，这些小泡能螯合钙和磷酸盐。钙化血管系统中EV的形成需要caveolin-1（CAV1），这是一种存在于膜内陷（caveolae）中的膜支架蛋白。值得注意的是，小窝蛋白-1缺陷小鼠的骨密度增加。我们假设caveolin-1可能在血管平滑肌细胞（VSMCs）和成骨细胞（HOBs）钙化EV形成中发挥不同的作用。使用siRNA降低CAV1的表达。甲基β-环糊精（MβCD）和钙蛋白酶抑制剂分别用于破坏和稳定VSMCs和HOBs的小窝结构域。结果：在两个独立的流行病学队列中，CAV1基因变异表明骨密度（BMD）和冠状动脉钙化（CAC）之间存在显著的负相关。在成骨（OS）培养基中培养增加了HOBs和VSMCs的钙化。CAV1的siRNA敲除消除了VSMC钙化，而对成骨细胞矿化没有影响。MβCD介导的小窝破裂导致成骨介质处理的VSMCs钙化增加3倍（p p p 结论：我们的数据表明了CAV1动力学介导的生理和病理生理矿化的基本细胞水平差异。这是第一项表明钙化EV形成的不同机制可能在钙化悖论中发挥作用的研究。补充信息：在线版本包含补充材料，可访问10.1007/s12195-023-00779-7。
Altered Caveolin-1 Dynamics Result in Divergent Mineralization Responses in Bone and Vascular Calcification.
Introduction: Though vascular smooth muscle cells adopt an osteogenic phenotype during pathological vascular calcification, clinical studies note an inverse correlation between bone mineral density and arterial mineral-also known as the calcification paradox. Both processes are mediated by extracellular vesicles (EVs) that sequester calcium and phosphate. Calcifying EV formation in the vasculature requires caveolin-1 (CAV1), a membrane scaffolding protein that resides in membrane invaginations (caveolae). Of note, caveolin-1-deficient mice, however, have increased bone mineral density. We hypothesized that caveolin-1 may play divergent roles in calcifying EV formation from vascular smooth muscle cells (VSMCs) and osteoblasts (HOBs).
Methods: Primary human coronary artery VSMCs and osteoblasts were cultured for up to 28 days in an osteogenic media. CAV1 expression was knocked down using siRNA. Methyl β-cyclodextrin (MβCD) and a calpain inhibitor were used, respectively, to disrupt and stabilize the caveolar domains in VSMCs and HOBs.
Results: CAV1 genetic variation demonstrates significant inverse relationships between bone-mineral density (BMD) and coronary artery calcification (CAC) across two independent epidemiological cohorts. Culture in osteogenic (OS) media increased calcification in HOBs and VSMCs. siRNA knockdown of CAV1 abrogated VSMC calcification with no effect on osteoblast mineralization. MβCD-mediated caveolae disruption led to a 3-fold increase of calcification in VSMCs treated with osteogenic media (p < 0.05) but hindered osteoblast mineralization (p < 0.01). Conversely, stabilizing caveolae by calpain inhibition prevented VSMC calcification (p < 0.05) without affecting osteoblast mineralization. There was no significant difference in CAV1 content between lipid domains from HOBs cultured in OS and control media.
Conclusion: Our data indicate fundamental cellular-level differences in physiological and pathophysiological mineralization mediated by CAV1 dynamics. This is the first study to suggest that divergent mechanisms in calcifying EV formation may play a role in the calcification paradox.
Supplementary information: The online version contains supplementary material available at 10.1007/s12195-023-00779-7.
期刊介绍：
The field of cellular and molecular bioengineering seeks to understand, so that we may ultimately control, the mechanical, chemical, and electrical processes of the cell. A key challenge in improving human health is to understand how cellular behavior arises from molecular-level interactions. CMBE, an official journal of the Biomedical Engineering Society, publishes original research and review papers in the following seven general areas:
Molecular: DNA-protein/RNA-protein interactions, protein folding and function, protein-protein and receptor-ligand interactions, lipids, polysaccharides, molecular motors, and the biophysics of macromolecules that function as therapeutics or engineered matrices, for example.
Cellular: Studies of how cells sense physicochemical events surrounding and within cells, and how cells transduce these events into biological responses. Specific cell processes of interest include cell growth, differentiation, migration, signal transduction, protein secretion and transport, gene expression and regulation, and cell-matrix interactions.
Mechanobiology: The mechanical properties of cells and biomolecules, cellular/molecular force generation and adhesion, the response of cells to their mechanical microenvironment, and mechanotransduction in response to various physical forces such as fluid shear stress.
Nanomedicine: The engineering of nanoparticles for advanced drug delivery and molecular imaging applications, with particular focus on the interaction of such particles with living cells. Also, the application of nanostructured materials to control the behavior of cells and biomolecules.
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