Browsing by Subject "Hyperplasia"
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Item Open Access Beta-arrestins regulate atherosclerosis and neointimal hyperplasia by controlling smooth muscle cell proliferation and migration.(Circ Res, 2008-07-03) Kim, Jihee; Zhang, Lisheng; Peppel, Karsten; Wu, Jiao-Hui; Zidar, David A; Brian, Leigh; DeWire, Scott M; Exum, Sabrina T; Lefkowitz, Robert J; Freedman, Neil JAtherosclerosis and arterial injury-induced neointimal hyperplasia involve medial smooth muscle cell (SMC) proliferation and migration into the arterial intima. Because many 7-transmembrane and growth factor receptors promote atherosclerosis, we hypothesized that the multifunctional adaptor proteins beta-arrestin1 and -2 might regulate this pathological process. Deficiency of beta-arrestin2 in ldlr(-/-) mice reduced aortic atherosclerosis by 40% and decreased the prevalence of atheroma SMCs by 35%, suggesting that beta-arrestin2 promotes atherosclerosis through effects on SMCs. To test this potential atherogenic mechanism more specifically, we performed carotid endothelial denudation in congenic wild-type, beta-arrestin1(-/-), and beta-arrestin2(-/-) mice. Neointimal hyperplasia was enhanced in beta-arrestin1(-/-) mice, and diminished in beta-arrestin2(-/-) mice. Neointimal cells expressed SMC markers and did not derive from bone marrow progenitors, as demonstrated by bone marrow transplantation with green fluorescent protein-transgenic cells. Moreover, the reduction in neointimal hyperplasia seen in beta-arrestin2(-/-) mice was not altered by transplantation with either wild-type or beta-arrestin2(-/-) bone marrow cells. After carotid injury, medial SMC extracellular signal-regulated kinase activation and proliferation were increased in beta-arrestin1(-/-) and decreased in beta-arrestin2(-/-) mice. Concordantly, thymidine incorporation and extracellular signal-regulated kinase activation and migration evoked by 7-transmembrane receptors were greater than wild type in beta-arrestin1(-/-) SMCs and less in beta-arrestin2(-/-) SMCs. Proliferation was less than wild type in beta-arrestin2(-/-) SMCs but not in beta-arrestin2(-/-) endothelial cells. We conclude that beta-arrestin2 aggravates atherosclerosis through mechanisms involving SMC proliferation and migration and that these SMC activities are regulated reciprocally by beta-arrestin2 and beta-arrestin1. These findings identify inhibition of beta-arrestin2 as a novel therapeutic strategy for combating atherosclerosis and arterial restenosis after angioplasty.Item Open Access G protein signaling and vein graft intimal hyperplasia: reduction of intimal hyperplasia in vein grafts by a Gbetagamma inhibitor suggests a major role of G protein signaling in lesion development.(Arterioscler Thromb Vasc Biol, 1998-08) Davies, MG; Fulton, Gregory J; Hagen, Per-Otto Frode; Huynh, Tam; Koch, Walter J; Lefkowitz, Robert J; Svendsen, EVein grafting results in the development of intimal hyperplasia with accompanying changes in guanine nucleotide-binding (G) protein expression and function. Several serum mitogens that act through G protein-coupled receptors, such as lysophosphatidic acid, stimulate proliferative pathways that are dependent on the G protein betagamma subunit (Gbetagamma)-mediated activation of p21ras. This study examines the role of Gbetagamma signaling in intimal hyperplasia by targeting a gene encoding a specific Gbetagamma inhibitor in an experimental rabbit vein graft model. This inhibitor, the carboxyl terminus of the beta-adrenergic receptor kinase (betaARK(CT)), contains a Gbetagamma-binding domain. Vein graft intimal hyperplasia was significantly reduced by 37% (P<0.01), and physiological studies demonstrated that the normal alterations in G protein coupling phenotypically seen in this model were blocked by betaARK(CT) treatment. Thus, it appears that Gbetagamma-mediated pathways play a major role in intimal hyperplasia and that targeting inhibitors of Gbetagamma signaling offers novel intraoperative therapeutic modalities to inhibit the development of vein graft intimal hyperplasia and subsequent vein graft failure.Item Open Access Kalirin promotes neointimal hyperplasia by activating Rac in smooth muscle cells.(Arteriosclerosis, thrombosis, and vascular biology, 2013-04) Wu, Jiao-Hui; Fanaroff, Alexander C; Sharma, Krishn C; Smith, Liisa S; Brian, Leigh; Eipper, Betty A; Mains, Richard E; Freedman, Neil J; Zhang, LishengObjective
Kalirin is a multifunctional protein that contains 2 guanine nucleotide exchange factor domains for the GTPases Rac1 and RhoA. Variants of KALRN have been associated with atherosclerosis in humans, but Kalirin's activity has been characterized almost exclusively in the central nervous system. We therefore tested the hypothesis that Kalirin functions as a Rho-guanine nucleotide exchange factor in arterial smooth muscle cells (SMCs).Approach and results
Kalirin-9 protein is expressed abundantly in aorta and bone marrow, as well as in cultured SMCs, endothelial cells, and macrophages. Moreover, arterial Kalirin was upregulated during early atherogenesis in apolipoprotein E-deficient mice. In cultured SMCs, signaling was affected similarly in 3 models of Kalirin loss-of-function: heterozygous Kalrn deletion, Kalirin RNAi, and treatment with the Kalirin Rho-guanine nucleotide exchange factor -1 inhibitor 1-(3-nitrophenyl)-1H-pyrrole-2,5-dione. With reduced Kalirin function, SMCs showed normal RhoA activation but diminished Rac1 activation, assessed as reduced Rac-GTP levels, p21-activated kinase autophosphorylation, and SMC migration. Kalrn(-/+) SMCs proliferated 30% less rapidly than wild-type SMCs. Neointimal hyperplasia engendered by carotid endothelial denudation was ≈60% less in Kalrn(-/+) and SMC-specific Kalrn(-/+) mice than in control mice.Conclusions
Kalirin functions as a guanine nucleotide exchange factor for Rac1 in SMCs, and promotes SMC migration and proliferation both in vitro and in vivo.Item Open Access Nrg1 is an injury-induced cardiomyocyte mitogen for the endogenous heart regeneration program in zebrafish.(Elife, 2015-04-01) Gemberling, Matthew; Karra, Ravi; Dickson, Amy L; Poss, Kenneth DHeart regeneration is limited in adult mammals but occurs naturally in adult zebrafish through the activation of cardiomyocyte division. Several components of the cardiac injury microenvironment have been identified, yet no factor on its own is known to stimulate overt myocardial hyperplasia in a mature, uninjured animal. In this study, we find evidence that Neuregulin1 (Nrg1), previously shown to have mitogenic effects on mammalian cardiomyocytes, is sharply induced in perivascular cells after injury to the adult zebrafish heart. Inhibition of Erbb2, an Nrg1 co-receptor, disrupts cardiomyocyte proliferation in response to injury, whereas myocardial Nrg1 overexpression enhances this proliferation. In uninjured zebrafish, the reactivation of Nrg1 expression induces cardiomyocyte dedifferentiation, overt muscle hyperplasia, epicardial activation, increased vascularization, and causes cardiomegaly through persistent addition of wall myocardium. Our findings identify Nrg1 as a potent, induced mitogen for the endogenous adult heart regeneration program.Item Open Access Patient-derived endothelial progenitor cells improve vascular graft patency in a rodent model.(Acta Biomater, 2012-01) Stroncek, JD; Ren, LC; Klitzman, B; Reichert, WMLate outgrowth endothelial progenitor cells (EPCs) derived from the peripheral blood of patients with significant coronary artery disease were sodded into the lumens of small diameter expanded polytetrafluoroethylene (ePTFE) vascular grafts. Grafts (1mm inner diameter) were denucleated and sodded either with native EPCs or with EPCs transfected with an adenoviral vector containing the gene for human thrombomodulin (EPC+AdTM). EPC+AdTM was shown to increase the in vitro rate of graft activated protein C (APC) production 4-fold over grafts sodded with untransfected EPCs (p<0.05). Unsodded control and EPC-sodded and EPC+AdTM-sodded grafts were implanted bilaterally into the femoral arteries of athymic rats for 7 or 28 days. Unsodded control grafts, both with and without denucleation treatment, each exhibited 7 day patency rates of 25%. Unsodded grafts showed extensive thrombosis and were not tested for patency over 28 days. In contrast, grafts sodded with untransfected EPCs or EPC+AdTM both had 7 day patency rates of 88-89% and 28 day patency rates of 75-88%. Intimal hyperplasia was observed near both the proximal and distal anastomoses in all sodded graft conditions but did not appear to be the primary occlusive failure event. This in vivo study suggests autologous EPCs derived from the peripheral blood of patients with coronary artery disease may improve the performance of synthetic vascular grafts, although no differences were observed between untransfected EPCs and TM transfected EPCs.Item Open Access Phosphorylation of USP20 on Ser334 by IRAK1 promotes IL-1β-evoked signaling in vascular smooth muscle cells and vascular inflammation.(The Journal of biological chemistry, 2023-07) Zhang, Lisheng; Wu, Jiao-Hui; Jean-Charles, Pierre-Yves; Murali, Pavitra; Zhang, Wenli; Jazic, Aeva; Kaur, Suneet; Nepliouev, Igor; Stiber, Jonathan A; Snow, Kamie; Freedman, Neil J; Shenoy, Sudha KReversible lysine-63 (K63) polyubiquitination regulates proinflammatory signaling in vascular smooth muscle cells (SMCs) and plays an integral role in atherosclerosis. Ubiquitin-specific peptidase 20 (USP20) reduces NFκB activation triggered by proinflammatory stimuli, and USP20 activity attenuates atherosclerosis in mice. The association of USP20 with its substrates triggers deubiquitinase activity; this association is regulated by phosphorylation of USP20 on Ser334 (mouse) or Ser333 (human). USP20 Ser333 phosphorylation was greater in SMCs of atherosclerotic segments of human arteries as compared with nonatherosclerotic segments. To determine whether USP20 Ser334 phosphorylation regulates proinflammatory signaling, we created USP20-S334A mice using CRISPR/Cas9-mediated gene editing. USP20-S334A mice developed ∼50% less neointimal hyperplasia than congenic WT mice after carotid endothelial denudation. WT carotid SMCs showed substantial phosphorylation of USP20 Ser334, and WT carotids demonstrated greater NFκB activation, VCAM-1 expression, and SMC proliferation than USP20-S334A carotids. Concordantly, USP20-S334A primary SMCs in vitro proliferated and migrated less than WT SMCs in response to IL-1β. An active site ubiquitin probe bound to USP20-S334A and USP20-WT equivalently, but USP20-S334A associated more avidly with TRAF6 than USP20-WT. IL-1β induced less K63-linked polyubiquitination of TRAF6 and less downstream NFκB activity in USP20-S334A than in WT SMCs. Using in vitro phosphorylation with purified IRAK1 and siRNA-mediated gene silencing of IRAK1 in SMCs, we identified IRAK1 as a novel kinase for IL-1β-induced USP20 Ser334 phosphorylation. Our findings reveal novel mechanisms regulating IL-1β-induced proinflammatory signaling: by phosphorylating USP20 Ser334, IRAK1 diminishes the association of USP20 with TRAF6 and thus augments NFκB activation, SMC inflammation, and neointimal hyperplasia.Item Open Access Targeting Gbeta gamma signaling in arterial vascular smooth muscle proliferation: a novel strategy to limit restenosis.(Proc Natl Acad Sci U S A, 1999-03-30) Iaccarino, G; Smithwick, LA; Lefkowitz, RJ; Koch, WJRestenosis continues to be a major problem limiting the effectiveness of revascularization procedures. To date, the roles of heterotrimeric G proteins in the triggering of pathological vascular smooth muscle (VSM) cell proliferation have not been elucidated. betagamma subunits of heterotrimeric G proteins (Gbetagamma) are known to activate mitogen-activated protein (MAP) kinases after stimulation of certain G protein-coupled receptors; however, their relevance in VSM mitogenesis in vitro or in vivo is not known. Using adenoviral-mediated transfer of a transgene encoding a peptide inhibitor of Gbetagamma signaling (betaARKct), we evaluated the role of Gbetagamma in MAP kinase activation and proliferation in response to several mitogens, including serum, in cultured rat VSM cells. Our results include the striking finding that serum-induced proliferation of VSM cells in vitro is mediated largely via Gbetagamma. Furthermore, we studied the effects of in vivo adenoviral-mediated betaARKct gene transfer on VSM intimal hyperplasia in a rat carotid artery restenosis model. Our in vivo results demonstrated that the presence of the betaARKct in injured rat carotid arteries significantly reduced VSM intimal hyperplasia by 70%. Thus, Gbetagamma plays a critical role in physiological VSM proliferation, and targeted Gbetagamma inhibition represents a novel approach for the treatment of pathological conditions such as restenosis.Item Open Access The Actin-Binding Protein Drebrin Inhibits Neointimal Hyperplasia.(Arteriosclerosis, thrombosis, and vascular biology, 2016-05) Stiber, Jonathan A; Wu, Jiao-Hui; Zhang, Lisheng; Nepliouev, Igor; Zhang, Zhu-Shan; Bryson, Victoria G; Brian, Leigh; Bentley, Rex C; Gordon-Weeks, Phillip R; Rosenberg, Paul B; Freedman, Neil JObjective
Vascular smooth muscle cell (SMC) migration is regulated by cytoskeletal remodeling as well as by certain transient receptor potential (TRP) channels, nonselective cation channels that modulate calcium influx. Proper function of multiple subfamily C TRP (TRPC) channels requires the scaffolding protein Homer 1, which associates with the actin-binding protein Drebrin. We found that SMC Drebrin expression is upregulated in atherosclerosis and in response to injury and investigated whether Drebrin inhibits SMC activation, either through regulation of TRP channel function via Homer or through a direct effect on the actin cytoskeleton.Approach and results
Wild-type (WT) and congenic Dbn(-/+) mice were subjected to wire-mediated carotid endothelial denudation. Subsequent neointimal hyperplasia was 2.4±0.3-fold greater in Dbn(-/+) than in WT mice. Levels of globular actin were equivalent in Dbn(-/+) and WT SMCs, but there was a 2.4±0.5-fold decrease in filamentous actin in Dbn(-/+) SMCs compared with WT. Filamentous actin was restored to WT levels in Dbn(-/+) SMCs by adenoviral-mediated rescue expression of Drebrin. Compared with WT SMCs, Dbn(-/+) SMCs exhibited increased TRP channel activity in response to platelet-derived growth factor, increased migration assessed in Boyden chambers, and increased proliferation. Enhanced TRP channel activity and migration in Dbn(-/+) SMCs were normalized to WT levels by rescue expression of not only WT Drebrin but also a mutant Drebrin isoform that binds actin but fails to bind Homer.Conclusions
Drebrin reduces SMC activation through its interaction with the actin cytoskeleton but independently of its interaction with Homer scaffolds.Item Open Access TNF is a potential therapeutic target to suppress prostatic inflammation and hyperplasia in autoimmune disease.(Nature communications, 2022-04) Vickman, Renee E; Aaron-Brooks, LaTayia; Zhang, Renyuan; Lanman, Nadia A; Lapin, Brittany; Gil, Victoria; Greenberg, Max; Sasaki, Takeshi; Cresswell, Gregory M; Broman, Meaghan M; Paez, J Sebastian; Petkewicz, Jacqueline; Talaty, Pooja; Helfand, Brian T; Glaser, Alexander P; Wang, Chi-Hsiung; Franco, Omar E; Ratliff, Timothy L; Nastiuk, Kent L; Crawford, Susan E; Hayward, Simon WAutoimmune (AI) diseases can affect many organs; however, the prostate has not been considered to be a primary target of these systemic inflammatory processes. Here, we utilize medical record data, patient samples, and in vivo models to evaluate the impact of inflammation, as seen in AI diseases, on prostate tissue. Human and mouse tissues are used to examine whether systemic targeting of inflammation limits prostatic inflammation and hyperplasia. Evaluation of 112,152 medical records indicates that benign prostatic hyperplasia (BPH) prevalence is significantly higher among patients with AI diseases. Furthermore, treating these patients with tumor necrosis factor (TNF)-antagonists significantly decreases BPH incidence. Single-cell RNA-seq and in vitro assays suggest that macrophage-derived TNF stimulates BPH-derived fibroblast proliferation. TNF blockade significantly reduces epithelial hyperplasia, NFκB activation, and macrophage-mediated inflammation within prostate tissues. Together, these studies show that patients with AI diseases have a heightened susceptibility to BPH and that reducing inflammation with a therapeutic agent can suppress BPH.Item Open Access Vein graft neointimal hyperplasia is exacerbated by CXCR4 signaling in vein graft-extrinsic cells.(Journal of vascular surgery, 2012-11) Zhang, Lisheng; Brian, Leigh; Freedman, Neil JObjective
Because vein graft neointimal hyperplasia engenders vein graft failure, and because most vein graft neointimal cells derive from outside the vein graft, we sought to determine whether vein graft neointimal hyperplasia is affected by activity of the CXC chemokine receptor-4 (CXCR4), which is important for bone marrow-derived cell migration.Methods
In congenic Cxcr4(-/+) and wild-type (WT) recipient mice, we performed interposition grafting of the common carotid artery with the inferior vena cava (IVC) of either Cxcr4(-/+) or WT mice to create four surgically chimeric groups of mice (n ≥ 5 each), characterized by vein graft donor/recipient: WT/WT; Cxcr4(-/+)/WT; WT/Cxcr4(-/+); and Cxcr4(-/+)/Cxcr4(-/+); vein grafts were harvested 6 weeks postoperatively.Results
The agonist for CXCR4 is expressed by cells in the arterializing vein graft. Vein graft neointimal hyperplasia was reduced by reducing CXCR4 activity in vein graft-extrinsic cells, but not in vein graft-intrinsic cells: the rank order of neointimal hyperplasia was WT/WT ≈ Cxcr4(-/+)/WT > WT/Cxcr4(-/+) ≈ Cxcr4(-/+)/Cxcr4(-/+); CXCR4 deficiency in graft-extrinsic cells reduced neointimal hyperplasia by 39% to 47% (P < .05). Vein graft medial area was equivalent in all grafts except Cxcr4(-/+)/Cxcr4(-/+), in which the medial area was 60% ± 20% greater (P < .05). Vein graft re-endothelialization was indistinguishable among all three vein graft groups. However, the prevalence of medial leukocytes was 40% ± 10% lower in Cxcr4(-/+)/Cxcr4(-/+) than in WT/WT vein grafts (P < .05), and the prevalence of smooth muscle actin-positive cells was 45% ± 20% higher (P < .05).Conclusions
We conclude that CXCR4 contributes to vein graft neointimal hyperplasia through mechanisms that alter homing to the vein graft of graft-extrinsic cells, particularly leukocytes.Clinical relevance
The utility of autologous vein grafts is severely reduced by neointimal hyperplasia, which accelerates subsequent graft atherosclerosis. Our study demonstrates that vein graft neointimal hyperplasia is aggravated by activity of the cell-surface “CXC” chemokine receptor-4 (CXCR4), which is critical for recruitment of bone marrow-derived cells to sites of inflammation. Our model for CXCR4 deficiency used mice with heterozygous deficiency of Cxcr4. Consequently, our results suggest the possibility that a CXCR4 antagonist--like plerixafor, currently in clinical use--could be applied to vein grafts periadventitially, and perhaps achieve beneficial effects on vein graft neointimal hyperplasia.