scholarly journals PCSK9 Induces Rat Smooth Muscle Cell Proliferation and Counteracts the Pleiotropic Effects of Simvastatin

2021 ◽  
Vol 22 (8) ◽  
pp. 4114
Author(s):  
Maria Giovanna Lupo ◽  
Silvia Marchianò ◽  
Maria Pia Adorni ◽  
Francesca Zimetti ◽  
Massimiliano Ruscica ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Smooth Muscle ◽  
Atherosclerotic Plaque ◽  
Regulatory Element ◽  
Smooth Muscle Actin ◽  
Cholesterol Biosynthesis ◽  
Lipid Lowering ◽  
Direct Role ◽  
Contractile Phenotype

Human atherosclerotic plaque contains smooth muscle cells (SMCs) negative for the contractile phenotype (α-smooth muscle actin) but positive for proprotein convertase subtilisin/kexin type 9 (PCSK9). Thus, we generated rat SMCs which overexpressed human PCSK9 (SMCsPCSK9) with the aim of investigating the role of PCSK9 in the phenotype of SMCs. PCSK9 overexpression in SMCsPCSK9 led to a significant downregulation of the low-density lipoprotein receptor (Ldlr) as well as transgelin (Sm22α), a marker of the contractile phenotype. The cell proliferation rate of SMCsPCSK9 was significantly faster than that of the control SMCs (SMCspuro). Interestingly, overexpression of PCSK9 did not impact the migratory capacity of SMCs in response to 10% FCS, as determined by Boyden’s chamber assay. Expression and activity of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (Hmgcr) was significantly increased in the presence of PCSK9, both in SMCPCSK9 and after treatment with recombinant PCSK9. The transcriptional activity of sterol regulatory element-binding protein (SREBP) was also increased in the presence of PSCK9, suggesting a direct role of PCSK9 in the control of SRE-responsive genes, like HMGCR. We also observed that cholesterol biosynthesis is elevated in SMCPCSK9, potentially explaining the increased proliferation observed in these cells. Finally, concentration-dependent experiments with simvastatin demonstrated that SMCsPCSK9 were partially resistant to the antiproliferative and antimigratory effect of this drug. Taken together, these data further support a direct role of PCSK9 in proliferation, migration, and phenotypic changes in SMCs—pivotal features of atherosclerotic plaque development. We also provide new evidence on the role of PCSK9 in the pharmacological response to statins—gold standard lipid-lowering drugs with pleiotropic action.

scholarly journals Jagged1/Notch3 Signaling Modulates Hemangioma-Derived Pericyte Proliferation and Maturation

2016 ◽  
Vol 40 (5) ◽  
pp. 895-907 ◽  
Author(s):  
Yi Ji ◽  
Siyuan Chen ◽  
Bo Xiang ◽  
Yuan Li ◽  
Li Li ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Smooth Muscle ◽  
Smooth Muscle Actin ◽  
Growth Arrest ◽  
Notch Signaling Pathway ◽  
Cell Growth Arrest ◽  
Muscle Myosin ◽  
Dependent Mechanism

Background: The Notch signaling pathway has been implicated in the pericyte phenotype, but its exact roles in hemangioma-derived pericytes (Hem-pericytes) remain ill defined. Methods: Hem-pericytes were stimulated by immobilized recombinant Jagged1. The potential mechanisms of Notch-induced Hem-pericytes growth arrest were investigated by cell cycle assay, and the role of the Notch in promoting Hem-pericyte maturation was also analyzed by real-time PCR and western blot. Results: Activation of Notch3 in Hem-pericytes significantly reduced cell proliferation and inhibited cell cycle transition. This event was associated with an increase in the levels of p21Cip1. Knockdown of p21Cip1 resulted in a significant rescue of Notch-induced cell growth arrest and an entry into the cell cycle. We showed that Jagged1 activation of Notch3 signaling upregulated the expression of the pericyte contractile markers smooth muscle myosin heavy chain (smMHC) and α-smooth muscle actin (αSMA), concomitant with an increase in the expression of myocardin in Hem-pericytes. We further revealed that the endothelial-derived Jagged1 modulated the Hem-pericyte phenotype via a contact-dependent mechanism. Conclusions: Our results demonstrated that Jagged1 activation of Notch3 resulted in a significant decrease in cell proliferation while concomitantly promoting Hem-pericyte maturation. These data provide initial evidence that Notch induces a quiescent phenotype in Hem-pericytes.

scholarly journals SAT-311 Role of SREBP in Regulation of Corticotroph Tumor ACTH Secretion and Cell Proliferation

2020 ◽  
Vol 4 (Supplement_1) ◽  
Author(s):  
Dongyun Zhang ◽  
Robert Damoiseaux ◽  
Marvin Bergsneider ◽  
Marilene B Wang ◽  
William H Yong ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Kinase Inhibitor ◽  
Regulatory Element ◽  
Cholesterol Biosynthesis ◽  
Transcriptional Level ◽  
Type I ◽  
Acth Secretion ◽  
Cushing Disease ◽  
Pomc Mrna

Abstract Cushing Disease (CD) is a life-threatening condition with suboptimal medical treatment. To identify drugs that not only inhibit ACTH secretion to attain eucortisolemia but also inhibit tumor growth, we conducted a high throughput screen employing a novel “gain of signal” ACTH AlphaLISA assay. From a kinase inhibitor library containing 430 compounds, we identified the dual PI3K/HDAC inhibitor, CUDC-907, as a potent inhibitor of both in vitro and in vivo corticotroph tumor ACTH secretion and growth. By stepwise comparison of CUDC-907 with mono-functional PI3K and HDAC inhibitors, we demonstrated that CUDC-907 exerts its inhibitory effect on ACTH secretion primarily through its inhibition of HDAC activity at the POMC transcriptional level; while PI3K-mediated inhibition of corticotroph cell viability further contributes to reduced ACTH secretion. We also used RNA-seq to characterize the global transcriptiome changes associated with CUDC-907 treatment. Hierarchical clustering showed that 1432 differentially expressed genes (DEGs, p≤0.05 and fold-change≥1.5) were altered by CUDC-907 treatment in comparison to the vehicle-treated control cells. Gene ontology (GO) analysis of 456 downregulated and 976 upregulated DEGs revealed that the most enriched biological processes were cholesterol biosynthesis (GO:0006695, p=1.977e-17) and the type I interferon signaling pathway (GO: 0060337, p=4.928e-7) respectively. Further analysis demonstrated downregulation of the membrane-bound transcription factor sterol regulatory element binding proteins (SREBPs). Downregulation of SREBP-2 by CUDC-907 as well as the several other target enzymes in the cholesterol biosynthesis and uptake pathway including IDI2, NSDHL, MVD, and HMGCR, was confirmed by real-time PCR. To further characterize a role for SREBP-2 in regulation of corticotroph tumor ACTH secretion and proliferation, we employed siRNA targeting endogenous SREBP-2 (SREBP-2 mRNA, Control vs. siRNA 1±0.03 vs. 0.6±0.08, p<0.05), and demonstrated that knockdown of SREBP-2 not only inhibited POMC mRNA expression (POMC mRNA, 1±0.03 vs. 0.7±0.01, p<0.01), and ACTH secretion (ACTH (ng/mL) 29±0.4 vs. 23±0.3, p<0.005), but also suppressed cell proliferation (Relative Proliferation Rate, 1±0.01 vs. 0.7±0.01, p<0.005). This was further confirmed by overexpression of cleaved mature SREBP-2, which led to increased POMC expression and cell proliferation. We demonstrate for the first time the role of the SREBP-mediated cholesterol biosynthesis pathway in regulation of corticotroph tumor POMC regulation and growth. Our studies identify SREBP and cholesterol biosynthesis as a therapeutic target in CD.

Abstract 102: Cd98 Modulates Atherosclerotic Plaque Morphology by Regulating Smooth Muscle Cell Proliferation via the P-38 Map Kinase Pathway

2015 ◽  
Vol 35 (suppl_1) ◽  
Author(s):  
Sara McCurdy ◽  
Yvonne Baumer ◽  
Franz Hess ◽  
William A Boisvert
Keyword(s):  
Cell Proliferation ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Smooth Muscle Cell ◽  
Atherosclerotic Plaque ◽  
Necrotic Core ◽  
Plaque Morphology ◽  
Fibrous Cap ◽  
Plaque Area

Smooth muscle cells (SMC) are known to migrate and proliferate to form a stabilizing fibrous cap that encapsulates atherosclerotic plaques. It has been shown that CD98hc, a transmembrane protein with a known role in amino acid transport and integrin signaling, is involved in proliferation and survival of various cell types including SMC. Based on these data, we hypothesized that CD98hc deficiency selectively in SMC would have pathogenic effects on atherosclerosis development and plaque composition. To test this, we utilized mice with SMC-specific deletion of the CD98hc ( CD98hc fl/fl SM22Cre + ) to determine the effects of CD98hc deficiency on SMC function in the context of atherosclerosis. We performed in vitro proliferation and survival/apoptosis assays to investigate the role of CD98hc in the proliferation and survival of primary mouse aortic vascular smooth muscle cells. We found that VSMC isolated from whole aortas of CD98hc -/- animals displayed approximately 60% reduced cell counts compared to control (41 ± 8.2% of control) after 5 days in culture. EdU assays in vivo showed a defect in the ability of CD98hc -/- SMC to proliferate, with 25% reduction in EdU-positive VSMC compared to controls (2.3 ± 0.2% vs 3 ± 0.2%). In addition, caspase-3 staining of SMC in vitro displayed a 41% increase in propensity of CD98hc -/- SMC to undergo apoptosis compared to controls (7.9 ± 0.6% vs 5.6 ± 0.5%). Furthermore, the absence of CD98hc in SMC caused a sharp increase in phosphorylated p-38, which was partially abrogated towards control levels when the cells were treated with PDGF-BB to induce proliferation. Long-term atherosclerosis study using SMC-CD98hc -/- /LDLR -/- mice showed that atherosclerotic plaque morphology was altered with increased necrotic core area (25.8 ± 1.9% vs 10.9 ± 1.6% necrotic core area per plaque area) due to a reduction in infiltration of SMC within the plaque (2.1 ± 0.4% vs 4.3 ± 0.4% SM22α positive area per plaque area) compared to control LDLR -/- mice. These data support an important role for CD98hc and its regulation of p-38 MAP kinase signaling in aortic vascular smooth muscle cell proliferation and survival. We conclude that CD98hc is critical for the formation of fibrous cap that is important in maintaining the stability of atherosclerotic plaque.

IL-4 and IL-13 upregulate arginase I expression by cAMP and JAK/STAT6 pathways in vascular smooth muscle cells

2000 ◽  
Vol 279 (1) ◽  
pp. C248-C256 ◽  
Author(s):  
Liu Hua Wei ◽  
Aaron T. Jacobs ◽  
Sidney M. Morris ◽  
Louis J. Ignarro
Keyword(s):  
Cell Proliferation ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Muscle Growth ◽  
Muscle Cells ◽  
Protein Levels ◽  
Ifn Γ ◽  
Arginase Ii ◽  
Arginase I

The objectives of this study were to determine whether rat aortic smooth muscle cells (RASMC) express arginase and to elucidate the possible mechanisms involved in the regulation of arginase expression. The results show that RASMC contain basal arginase I (AI) activity, which is significantly enhanced by stimulating the cells with either interleukin (IL)-4 or IL-13, but arginase II (AII) expression was not detected under any condition studied here. We further investigated the signal transduction pathways responsible for AI induction. AI mRNA and protein levels were enhanced by addition of forskolin (1 μM) and inhibited by H-89 (30 μM), suggesting positive regulation of AI by a protein kinase A pathway. Genistein (10 μg/ml) and sodium orthovanadate (Na3VO4; 10 μM) were used to investigate the role of tyrosine phosphorylation in the control of AI expression. Genistein inhibited, whereas Na3VO4enhanced the induction of AI by IL-4 or IL-13. Along with immunoprecipitation and immunoblot analyses, these data implicate the JAK/STAT6 pathway in AI regulation. Dexamethasone (Dex) and interferon (IFN)-γ were investigated for their effects on AI induction. Dex (1 μM) and IFN-γ (100 U/ml) alone had no effect on basal AI expression in RASMC, but both reduced AI induction by IL-4 and IL-13. In combination, Dex and IFN-γ abolished AI induction by IL-4 and IL-13. Finally, both IL-4 and IL-13 significantly increased RASMC DNA synthesis as monitored by [3H]thymidine incorporation, demonstrating that upregulation of AI is correlated with an increase in cell proliferation. Blockade of AI induction by IFN-γ, H-89, or genistein also blocked the increase in cell proliferation. These observations are consistent with the possibility that upregulation of AI might play an important role in the pathophysiology of vascular disorders characterized by excessive smooth muscle growth.

Abstract 454: Myeloid CD98hc Deficiency Reduces Atherosclerotic Plaque Development via Impaired Proliferation of Macrophages

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Sara McCurdy ◽  
William A Boisvert
Keyword(s):  
Bone Marrow ◽  
Cell Proliferation ◽  
Atherosclerotic Plaque ◽  
Bone Marrow Cells ◽  
Transmembrane Protein ◽  
Mouse Bone Marrow ◽  
Plaque Development ◽  
Primary Mouse

Macrophage accumulation is a key process affecting all stages of atherosclerosis. Whether these cells accumulate in plaque solely by recruitment of monocytes from circulation or by proliferation within the plaque is an important question that has garnered much interest in recent years. Originally identified as a lymphocyte activation marker, CD98hc (SLC3A2) is a transmembrane protein involved in cell proliferation and survival via integrin signaling and MAP kinase activation. We hypothesized that CD98hc deficiency in myeloid cells would have a protective effect on atherosclerosis development and plaque composition by limiting macrophage proliferation. For the studies described, we utilized mice with myeloid-specific deletion of the CD98hc ( CD98hc fl/fl LysMCre + ) to determine the effects of CD98hc deficiency on macrophage function in the context of atherosclerosis . We performed in vitro assays to investigate the role of CD98hc in the proliferation and survival of primary mouse bone marrow derived macrophages. Although we found no differences in the number of bone marrow cells isolated from control or CD98hc -/- animals, after differentiation with MCS-F for 7 days, the number of macrophages obtained from CD98hc -/- mice was approximately 80% lower (7.2 ± 2.2 x 10 6 vs. 42.4 ± 4.6 x 10 6 per mouse) compared to control mice. Proliferation assays in vitro using EdU revealed approximately 50% (15.4 ± 2.5% vs. 7.5±1.8%) reduced cell proliferation in CD98hc -/- macrophages compared to control cells that could not be rescued with the addition M-CSF. In a 6-week atherosclerosis study using Ldlr -/- CD98hc fl/fl LysMCre + mice, Oil-Red O staining of whole aortae as well as aortic sinus sections showed that atherosclerotic plaque development was reduced compared to Ldlr -/- CD98hc fl/fl LysMCre - control mice. Additionally, immunohistochemical staining of atherosclerotic tissues revealed a reduction in macrophage abundance and proliferation within the plaque of Ldlr -/- CD98hc fl/fl LysMCre + mice compared to control mice. These findings support an important role of CD98hc in macrophage proliferation within the plaque environment, and provide a novel target for reducing atherosclerosis.

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