Increased ADAMTS1 mediates SPARC-dependent collagen deposition in the aging myocardium

Secreted protein acidic and rich in cysteine (SPARC) is a collagen-binding matricellular protein highly expressed during fibrosis. Fibrosis is a prominent component of cardiac aging that reduces myocardial elasticity. Previously, we reported that SPARC deletion attenuated myocardial stiffness and collagen deposition in aged mice. To investigate the mechanisms by which SPARC promotes age-related cardiac fibrosis, we evaluated six groups of mice ( n = 5–6/group): young (3–5 mo old), middle-aged (10–12 mo old), and old (18–29 mo old) C57BL/6 wild type (WT) and SPARC-null (Null) mice. Collagen content, determined by picrosirius red staining, increased in an age-dependent manner in WT but not in Null mice. A disintegrin and metalloproteinase with thrombospondin-like motifs 1 (ADAMTS1) increased in middle-aged and old WT compared with young, whereas in Null mice only old animals showed increased ADAMTS1 expression. Versican, a substrate of ADAMTS1, decreased with age only in WT. To assess the mechanisms of SPARC-induced collagen deposition, we stimulated cardiac fibroblasts with SPARC. SPARC treatment increased secretion of collagen I and ADAMTS1 (both the 110-kDa latent and 87-kDa active forms) into the conditioned media as well as the cellular expression of transforming growth factor-β1-induced protein (Tgfbi) and phosphorylated Smad2. An ADAMTS1 blocking antibody suppressed the SPARC-induced collagen I secretion, indicating that SPARC promoted collagen production directly through ADAMTS1 interaction. In conclusion, ADAMTS1 is an important mediator of SPARC-regulated cardiac aging.

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MicroRNA-210-5p Alleviates Cardiac Fibrosis via Targeting Transforming Growth Factor-beta Type I Receptor in Rats on High Salt Diet

10.21203/rs.3.rs-243221/v1 ◽

2021 ◽

Author(s):

Kun Zhao ◽

Yukang Mao ◽

Xiaoman Ye ◽

Jiazheng Ma ◽

Litao Sun ◽

...

Keyword(s):

Blood Pressure ◽

Growth Factor ◽

Transforming Growth Factor Beta ◽

Cardiac Fibrosis ◽

Transforming Growth Factor ◽

Collagen I ◽

Type I ◽

High Salt Diet ◽

Salt Diet ◽

Growth Factor Beta

Abstract Background: The aim of the present study was to explore whether high salt diet (HSD) caused cardiac fibrosis regardless of blood pressure in rats, and to determine the effects of microRNA (miR)-210-5p on sodium chloride (NaCl)-induced fibrosis in neonatal rat cardiac fibroblasts (NRCFs) and its target. Methods: The rats received 8% HSD in vivo, and NRCFs were treated with NaCl in vitro. Results: The levels of collagen I, alpha-smooth muscle actin (α-SMA) and transforming growth factor-beta (TGF-β) were increased in the heart of hypertension (HTN), hypertension-prone (HP) and hypertension-resistant (HR) rats on HSD. Middle and high doses (50 mM and 100 mM) of NaCl increased the levels of collagen I, α-SMA and TGF-β in NRCFs. The expression level of miR-210-5p was reduced in NaCl-treated NRCFs by miR high-throughput sequencing. The NaCl-induced increases of collagen I, α-SMA and TGF-β were inhibited by miR-210-5p agomiR, and further enhanced by miR-210-5p antagomiR. Bioinformatics analysis and luciferase reporter assays demonstrated that TGF-β type I receptor (TGFβRI) was a direct target gene of miR-210-5p. These results indicated that HSD resulted in cardiac fibrosis regardless of blood pressure. Conclusion: The upregulation of miR-210-5p could attenuate NRCF fibrosis via targeting TGFβRI. Thus, upregulating miR-210-5p to inhibit TGF-β signaling pathway might be a strategy for the treatment of cardiac fibrosis.

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Neutrophil PAD4 negatively influences cardiac function and remodeling with increasing age

European Heart Journal ◽

10.1093/eurheartj/ehab724.3340 ◽

2021 ◽

Vol 42 (Supplement_1) ◽

Author(s):

S Van Bruggen ◽

J Van Wauwe ◽

P Carai ◽

L Frederix ◽

L Vangilbergen ◽

...

Keyword(s):

Heart Failure ◽

Cardiac Function ◽

Cardiac Fibrosis ◽

Essential Gene ◽

Heart Function ◽

Systolic Function ◽

Natural Aging ◽

Collagen Deposition ◽

National Budget ◽

Age Related

Abstract Background Aging can be viewed as a status of chronic inflammation, in which neutrophils have a lower threshold for activation. The enzyme peptidylarginine deiminase 4 (PAD4), which catalyzes the conversion of arginine to citrulline, will be activated in a certain population of neutrophils. When this conversion takes place on the histones, neutrophils can form neutrophil extracellular traps (NETs), which are both prothrombotic and proinflammatory. Mice lacking this enzyme systemically were previously reported to be protected from age-related fibrosis. Purpose We aimed to study the long-term effect of neutrophils on cardiac health during the process of natural aging. We hypothesized that neutrophil PAD4, and in consequence NETs, are involved in cardiac fibrosis development, which in turn will result in impaired cardiac function. Methods We generated a mouse model of impaired NET release capability via deletion of PAD4, a NET-essential gene, under the neutrophil-specific promoter (PAD4fl/flMRP8Cre+). In order to study heart failure (HF) development, these specific deletion mice and their littermate controls were aged for a period of two years (coinciding with approximately 70 years of age in the human population; the age at which HF is the number one cause of hospitalization), after which cardiac function and remodeling were evaluated by echocardiography and histology, respectively. A separate set of young mice (12 weeks) were evaluated in parallel. Results We performed a comprehensive echocardiography analysis including both structural and functional parameters. As for systolic function, we could see that in old wild type (WT) mice, ejection fraction (EF) significantly decreased as compared to EF in young and healthy (YH) mice (YH - 67±6%, WT - 53±10%; p<0.0001) (Figure 1B). However, this decrease in systolic function was absent in the old PAD4fl/flMRP8Cre+ mice, with EF being comparable to the YH group (PAD4fl/flMRP8Cre+ - 67±7%; p=0.9169) (figure 1 A,B). As for diastolic function, again we could see a marked decrease in E/A ratio in the WT as compared to the YH population (YH- 1.50±0.23, WT – 1.21±0.17; p<0.0001), while this functional deterioration was absent in aged PAD4fl/flMRP8Cre+ animals (PAD4fl/flMRP8Cre+ - 1.38±0.21; p=0.0837) (Figure 1 C,D). To link this decline in heart function to tissue remodeling, we quantified collagen deposition in the heart. We saw that natural aging resulted in an increase in cardiac collagen deposition in the WT population as compared to YH mice (YH – 0.86±0.63%, WT – 4.02±1.71%). This increased collagen deposition was absent in the neutrophil deletion mice (PAD4fl/flMRP8Cre+ - 1.7±0.76%). Additionally, when comparing WT to PAD4 deletion-mice, we saw that the increase in collagen deposition is significantly higher in the WT mice (p<0.0001). Conclusion Our data confirms neutrophil PAD4 involvement in heart failure progression by promoting cardiac fibrosis, resulting in cardiac dysfunction. FUNDunding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Fonds Wetenschappelijk Onderzoek (FWO) - Vlaanderen

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Peroxynitrite augments fibroblast-mediated tissue remodeling via myofibroblast differentiation

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.90264.2008 ◽

2008 ◽

Vol 295 (5) ◽

pp. L800-L808 ◽

Cited By ~ 23

Author(s):

Tomohiro Ichikawa ◽

Hisatoshi Sugiura ◽

Akira Koarai ◽

Satoru Yanagisawa ◽

Masae Kanda ◽

...

Keyword(s):

Transforming Growth Factor ◽

Airway Remodeling ◽

Smooth Muscle Actin ◽

Fetal Lung ◽

Collagen I ◽

Airflow Limitation ◽

Lung Fibroblasts ◽

Myofibroblast Differentiation ◽

Dependent Manner ◽

Concentration Dependent Manner

Irreversible airflow limitation in asthma is associated with airway remodeling in which the differentiation of fibroblasts to myofibroblasts plays a pivotal role. In asthmatic airways, excessive production of reactive nitrogen species (RNS) has been observed. The aim of this study is to evaluate whether peroxynitrite, one of the RNS, can affect the differentiation of fibroblasts to myofibroblasts. Human fetal lung fibroblasts were treated with various concentrations of authentic peroxynitrite or a peroxynitrite donor 3-morpholinosydnonimine hydrochloride (SIN-1), and the expressions of α-smooth muscle actin (α-SMA) and desmin, markers of myofibroblast differentiation, were evaluated. The releases of transforming growth factor-β1 (TGF-β1) and ECM proteins including fibronectin and collagen I were assessed. To clarify the mechanism in this differentiation, the effect of anti-TGF-β antibody or NF-κB inhibitors on the α-SMA expression and ECM production was assessed. Peroxynitrite and SIN-1 significantly augmented the α-SMA expression compared with control in a concentration-dependent manner ( P < 0.01 and P < 0.05, respectively). Peroxynitrite significantly increased desmin and TGF-β1 production ( P < 0.01). Peroxynitrite enhanced the translocation of NF-κB into the nucleus confirmed by immunocytostaining and immunoblotting. Peroxynitrite-augmented α-SMA expression was blocked by NF-κB inhibitors, MG132 and caffeic acid phenethyl ester (CAPE), and anti-TGF-β antibody. CAPE completely inhibited the peroxynitrite-augmented TGF-β1 release. The production of fibronectin and collagen I was significantly increased by peroxynitrite ( P < 0.01) and inhibited by anti-TGF-β antibody. These results suggest that RNS can affect the differentiation to myofibroblasts and excessive ECM production via a NF-κB-TGF-β1-dependent pathway.

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Activated Cardiac Fibroblasts Control Contraction of Human Fibrotic Cardiac Microtissues by a β-Adrenoreceptor-Dependent Mechanism

Cells ◽

10.3390/cells9051270 ◽

2020 ◽

Vol 9 (5) ◽

pp. 1270 ◽

Cited By ~ 1

Author(s):

Przemysław Błyszczuk ◽

Christian Zuppinger ◽

Ana Costa ◽

Daria Nurzynska ◽

Franca Di Meglio ◽

...

Keyword(s):

Molecular Mechanisms ◽

Cardiac Fibrosis ◽

Transforming Growth Factor ◽

Cardiac Fibroblasts ◽

Experimental Models ◽

Cardiac Contraction ◽

Dependent Manner ◽

Direct Stimulation ◽

Beating Rate ◽

Tgf Β1

Cardiac fibrosis represents a serious clinical problem. Development of novel treatment strategies is currently restricted by the lack of the relevant experimental models in a human genetic context. In this study, we fabricated self-aggregating, scaffold-free, 3D cardiac microtissues using human inducible pluripotent stem cell (iPSC)-derived cardiomyocytes and human cardiac fibroblasts. Fibrotic condition was obtained by treatment of cardiac microtissues with profibrotic cytokine transforming growth factor β1 (TGF-β1), preactivation of foetal cardiac fibroblasts with TGF-β1, or by the use of cardiac fibroblasts obtained from heart failure patients. In our model, TGF-β1 effectively induced profibrotic changes in cardiac fibroblasts and in cardiac microtissues. Fibrotic phenotype of cardiac microtissues was inhibited by treatment with TGF-β-receptor type 1 inhibitor SD208 in a dose-dependent manner. We observed that fibrotic cardiac microtissues substantially increased the spontaneous beating rate by shortening the relaxation phase and showed a lower contraction amplitude. Instead, no changes in action potential profile were detected. Furthermore, we demonstrated that contraction of human cardiac microtissues could be modulated by direct electrical stimulation or treatment with the β-adrenergic receptor agonist isoproterenol. However, in the absence of exogenous agonists, the β-adrenoreceptor blocker nadolol decreased beating rate of fibrotic cardiac microtissues by prolonging relaxation time. Thus, our data suggest that in fibrosis, activated cardiac fibroblasts could promote cardiac contraction rate by a direct stimulation of β-adrenoreceptor signalling. In conclusion, a model of fibrotic cardiac microtissues can be used as a high-throughput model for drug testing and to study cellular and molecular mechanisms of cardiac fibrosis.

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Abstract 16161: GSK-3β Activates Autophagy and Protects Against Cardiac Aging Through Ulk1

Circulation ◽

10.1161/circ.132.suppl_3.16161 ◽

2015 ◽

Vol 132 (suppl_3) ◽

Author(s):

Peiyong Zhai ◽

Akihiro Shirakabe ◽

Takanobu Yamamoto ◽

Yoshiyuki Ikeda ◽

Bonaventure Magrys ◽

...

Keyword(s):

Cardiac Fibrosis ◽

Glycogen Synthase ◽

Left Ventricular ◽

Protective Mechanism ◽

Cross Sectional ◽

Cardiac Aging ◽

Beneficial Effects ◽

Stiffness Constant ◽

Age Related ◽

Gsk 3Β

Accumulating lines of evidence suggest that glycogen synthase kinase-3β (GSK-3β) is involved in aging. However, the effects of GSK-3β on cardiac aging and the underlying mechanisms remain to be elucidated. Autophagy, a protective mechanism in aging, decreases with age. We hypothesized that GSK-3β attenuates cardiac aging via Ulk1, a regulator of autophagy, and studied constitutively active GSK-3βS9A knock-in mice (βKI), GSK-3βS9A/Ulk1+/- bigenic mice (Bigenic), and GSK-3β+/- mice (βKO) up to 24 months (M) of age. Left ventricular (LV) weight/body weight (LVW/BW, mg/g) was not significantly different among wild-type mice (WT), βKI and βKO at 6M. It was lower in βKI (2.4±0.1, p<0.005) and higher in βKO (4.8±0.8, p<0.05) than in WT (3.8±0.2) at 24M. Cardiomyocyte cross-sectional area (CSA, μm2) was smaller in βKI (360±9, p<0.001) but bigger in βKO (540±11, p<0.01) than in WT (502±5) at 24M. The LVW/BW was greater (3.5±0.2, p<0.001) and the CSA was bigger (527±4, p<0.001) in Bigenic than in βKI at 24M. These data demonstrate that GSK-3β inhibits age-dependent cardiac hypertrophy via Ulk1. Cardiac fibrosis (%) was more in βKO (5.4±0.1, p<0.001) and less in βKI (2.4±0.1, p<0.001) than in WT (4.0±0.3) at 24M. There was much more fibrosis in Bigenic (5.5±0.6, p<0.001) than in βKI at 24M. These data show that GSK-3β reduces age-related cardiac fibrosis via Ulk1. LV end-systolic elastance (Ees, mmHg/μl) and chamber stiffness constant (CSC, μl-1) were not significantly different among WT, βKI, and βKO at 6M. At 24M, the Ees was lower in βKO (4±1, p<0.05) and higher in βKI (12±3, p<0.05) than in WT (7±0), and the CSC was higher in βKO (0.19±0.01, p<0.001) and lower in βKI (0.06±0.01, p<0.001) than in WT (0.14±0.01). The beneficial effects of GSK-3β on cardiac function were abolished in the Bigenic, indicating that GSK-3β prevents age-specific cardiac dysfunction via Ulk1. The level of p62, a protein degraded by autophagy, was lower in βKI and higher in βKO than in WT. The numbers of autophagosomes and autolysosomes were significantly greater in βKI/tfLC3 (tandem fluorescent mRFP-GFP-LC3) mice than in tfLC3 or βKI/tfLC3/Ulk1+/- mice. These data suggest that GSK-3β activates autophagy via Ulk1. In conclusion, GSK-3β attenuates cardiac aging by activating Ulk1-dependent autophagy.

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Abstract 384: Nov/ccn3: A Potential Antagonist of Angiotensin Ii Effects in Kidney Disease

Hypertension ◽

10.1161/hyp.60.suppl_1.a384 ◽

2012 ◽

Vol 60 (suppl_1) ◽

Author(s):

Christos Chadjichristos ◽

Pierre-Olivier Marchal ◽

Aurelie Cuinet ◽

Jihane Pakradouni ◽

Julie Toubas ◽

...

Keyword(s):

Growth Factor ◽

Kidney Disease ◽

Renal Disease ◽

Transforming Growth Factor ◽

Tissue Growth ◽

Matricellular Protein ◽

Dependent Manner ◽

Angiotensin Receptor ◽

Protein Levels ◽

Fibrotic Process

Progression of chronic kidney disease (CKD) represents a constant challenge for the identification of new therapeutic targets. The NOV/CCN3 matricellular protein, belonging to the CCN (CYR61/CCN1, CTGF/CCN2, NOV/CCN3) family, could represent a new endogenous inhibitor of the fibrotic process that plays a crucial role in tissue damage and regression of renal function. However, the relation between NOV and renal disease remains unclear. Our objective was to study whether alterations in renal NOV expression are involved in the progression of CKD. NOV mRNA expression was significantly induced by 2.5-3 fold at 12-16 weeks in renin-transgenic mice (RenTg), a well established model of AngII-mediated nephropathy, through the Angiotensin receptor 1 (AT 1 R) compared to normotensive control animals (n=7 mice per group, p<0.05). NOV immunodetection revealed an up-regulation mainly in vascular smooth muscle cells (VSMC) in the renal cortex of RenTg mice. Interestingly, NOV expression was inversely correlated to known fibrotic markers such as Connective tissue growth factor, Transforming growth factor and collagen I, during the progression of renal disease in RenTg mice. NOV up-regulation at mRNA and protein levels (∼ 2.5 fold p<0.05) by AngII through AT 1 R was also observed in VSMC cultures. Furthermore, infection of these cells with an adenovirus recombinant for NOV strongly repressed in a dose-dependent manner the expression of the Angiotensin Receptor 1 reaching 72 and 78% (p<0.05) at mRNA and protein levels respectively. Treatment with exogenous NOV gave similar results. Conversely, knocking down NOV by small interfering RNA increased Angiotensin Receptor 1 by ∼3.5 to 4.5 fold and by ∼7.5 fold (p<0.05) at mRNA and protein levels respectively. These results indicate that NOV is involved in an AngII- AT 1 R regulatory loop. In subsequent studies the NOV-induced downregulation of AT 1 R was abrogated by integrin β1 specific siRNAs. Thus, NOV can attenuate the deleterious profibrotic effects of AngII during the progression of CKD and may provide a potential therapeutic option against this pathology.

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Abstract 1: Opposing Roles for Endoglin and Soluble Endoglin in Cardiac Remodeling and Heart Failure

Circulation Research ◽

10.1161/res.109.suppl_1.a1 ◽

2011 ◽

Vol 109 (suppl_1) ◽

Author(s):

Navin K Kapur ◽

Szuhuei Wilson ◽

Adil A Yunis ◽

Corey Baker ◽

Mark J Aronovitz ◽

...

Keyword(s):

Heart Failure ◽

Transforming Growth Factor Beta ◽

Neutralizing Antibodies ◽

Collagen Synthesis ◽

Cardiac Fibrosis ◽

Transforming Growth Factor ◽

Cardiomyocyte Hypertrophy ◽

Lv Function ◽

Dependent Manner ◽

Soluble Endoglin

Transforming growth factor beta-1 (TGFb1) promotes cardiac fibrosis. The transmembrane co-receptor Endoglin (Eng; CD105) facilitates TGFb1 signaling via SMAD effector proteins. In contrast, a circulating form of soluble endoglin (sEng) inhibits TGFb1 signaling in vascular endothelium. We recently reported that increased sEng levels in human serum correlate with clinical indices of heart failure severity. Therefore, we tested the hypothesis that Eng and sEng mediate opposing effects on cardiac fibrosis in heart failure. In male, wild-type mice (WT), Eng expression increased in the left ventricle (LV) after 2, 4, and 10 weeks of thoracic aortic constriction (TAC) accompanied by progressive LV fibrosis and hypertrophy. In contrast to WT mice, Eng haploinsufficient (Eng +/− ) mice had preserved LV function (FS%: 78±4 vs 22±16, Eng +/− vs WT, p<0.01) and improved survival [88%(7/8) vs 50%(4/8), Eng +/− vs WT, p<0.001) after 10 weeks of TAC. Reduced LV fibrosis was observed in Eng +/− mice, while LV mass, cardiomyocyte hypertrophy, and calcineurin, SerCA, and bMHC expression were comparable to WT after TAC. Capillary density was significantly higher in Eng +/− mice after TAC compared to WT. LV SMAD phosphorylation (pSMAD) after TAC was studied and a preferential increase was observed in pSMAD1/5/8 expression in Eng +/− mice as compared to WT mice, in which cardiac pSmad2/3 expression was increased. The dependence of TGFb1 induced collagen synthesis on Eng expression was tested in vitro using human cardiac fibroblasts (hCF). Neutralizing antibodies and siRNA against Eng each attenuated TGFb1 induced collagen synthesis. In contrast, conditioned media from cells transfected with an adenovirus over-expressing sEng attenuated pSMAD2/3 expression and TGFb1 induced collagen synthesis in hCF. Treatment of hCF with recombinant sEng blocked TGFb1 induced collagen synthesis in a dose-dependent manner, confirming an inhibitory role of sEng. These results indicate that Eng and sEng mediate opposite effects on TGFb1 induced collagen synthesis. Reduced Eng expression uncouples cardiac fibrosis from cardiomyocyte hypertrophy, promotes SMAD1/5/8-signaling, and enhances angiogenesis. Eng may represent a novel therapeutic target to improve survival in heart failure.

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Treadmill Training Increases SIRT-1 and PGC-1αProtein Levels and AMPK Phosphorylation in Quadriceps of Middle-Aged Rats in an Intensity-Dependent Manner

Mediators of Inflammation ◽

10.1155/2014/987017 ◽

2014 ◽

Vol 2014 ◽

pp. 1-11 ◽

Cited By ~ 18

Author(s):

Nara R. C. Oliveira ◽

Scherolin O. Marques ◽

Thais F. Luciano ◽

José R. Pauli ◽

Leandro P. Moura ◽

...

Keyword(s):

Training Session ◽

Treadmill Training ◽

Dependent Manner ◽

Aged Rats ◽

Middle Aged ◽

Protein Levels ◽

Ampk Phosphorylation ◽

Age Related ◽

Male Wistar Rats ◽

Inflammatory Proteins

The present study investigated the effects of running at 0.8 or 1.2 km/h on inflammatory proteins (i.e., protein levels of TNF-α, IL-1β, and NF-κB) and metabolic proteins (i.e., protein levels of SIRT-1 and PGC-1α, and AMPK phosphorylation) in quadriceps of rats. Male Wistar rats at 3 (young) and 18 months (middle-aged rats) of age were divided into nonexercised (NE) and exercised at 0.8 or 1.2 km/h. The rats were trained on treadmill, 50 min per day, 5 days per week, during 8 weeks. Forty-eight hours after the last training session, muscles were removed, homogenized, and analyzed using biochemical and western blot techniques. Our results showed that: (a) running at 0.8 km/h decreased the inflammatory proteins and increased the metabolic proteins compared with NE rats; (b) these responses were lower for the inflammatory proteins and higher for the metabolic proteins in young rats compared with middle-aged rats; (c) running at 1.2 km/h decreased the inflammatory proteins and increased the metabolic proteins compared with 0.8 km/h; (d) these responses were similar between young and middle-aged rats when trained at 1.2 km. In summary, the age-related increases in inflammatory proteins, and the age-related declines in metabolic proteins can be reversed and largely improved by treadmill training.

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Abstract 15308: SPARC Deletion Suppresses Age-related Cardiac Inflammation

Circulation ◽

10.1161/circ.130.suppl_2.15308 ◽

2014 ◽

Vol 130 (suppl_2) ◽

Author(s):

Hiroe Toba ◽

Lisandra E de Castro Brás ◽

Catalin F Baicu ◽

Michael R Zile ◽

Merry L Lindsey ◽

...

Keyword(s):

Structural Changes ◽

Matricellular Protein ◽

Secreted Protein ◽

Wild Type ◽

Middle Aged ◽

Collagen Binding ◽

Cardiac Inflammation ◽

Age Related ◽

Age Dependent ◽

Inflammatory Chemokines

Secreted protein acidic and rich in cysteine (SPARC) is a collagen-binding matricellular protein that facilitates the formation and assembly of mature cross-linked collagen fibrils. We previously reported that aged mice exhibit increased SPARC levels in the heart, and SPARC deletion inhibits age-related collagen deposition. The present study investigated whether SPARC deletion could suppress age-related cardiac inflammation. Six groups of mice were studied: young (3-5 month old), middle-aged (10-12 m.o.) and old (18-29 m.o.) C57BL/6J wild type (WT) and SPARC-null (Null) mice (n=5-10 per group). Echocardiography and histology revealed that LV wall thickness and myocyte size were 15% and 43% increased with age in WT, respectively (both p<0.05 old versus young). These age-dependent structural changes were blunted in the Null. The pro-inflammatory chemokines Ccl5 and Ccr2 were increased in middle-aged and old WT, and Cxcl1 was increased in old WT, compared to young WT (all p<0.05). Interestingly, the age-dependent increases in Ccl5, Ccr2, and Cxcl1 were delayed in Null. Migration inhibitory factor (Mif) and Cx3cl1 showed an age-dependent increase in both WT and Null. Cxcl5 expression levels increased age-dependently only in Null, suggesting a compensatory increase for the decline of other chemokines. In summary, these results suggest that SPARC deletion slowed age-related cardiac inflammation.

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Transforming growth factor-β inhibition of proteasomal activity: a potential mechanism of growth arrest

AJP Cell Physiology ◽

10.1152/ajpcell.00550.2002 ◽

2003 ◽

Vol 285 (2) ◽

pp. C277-C285 ◽

Cited By ~ 117

Author(s):

Laura Tadlock ◽

Yoko Yamagiwa ◽

James Hawker ◽

Carla Marienfeld ◽

Tushar Patel

Keyword(s):

Cell Cycle ◽

Growth Factor ◽

Cell Growth ◽

Transforming Growth Factor ◽

Potent Inhibitor ◽

Dependent Manner ◽

Proteasomal Activity ◽

Cellular Expression ◽

Hydrolysis Of

Although the proteasome plays a critical role in the controlled degradation of proteins involved in cell cycle control, the direct modulation of proteasomal function by growth regulatory signaling has not yet been demonstrated. We assessed the effect of transforming growth factor (TGF)-β, a potent inhibitor of cell growth, on proteasomal function. TGF-β selectively decreased hydrolysis of the proteasomal substrate Cbz-Leu-Leu-Leu-7-amido-4-methyl-coumarin (z-LLL-AMC) in a concentration-dependent manner but did not inhibit hydrolysis of other substrates Suc-Leu-Leu-Val-Tyr-AMC (suc-LLVY-AMC) or Cbz-Leu-Leu-Glu-AMC (z-LLE-AMC). An increase in intracellular oxidative injury occurred during incubation with TGF-β. Furthermore, in vitro hydrolysis of z-LLL-AMC, but not suc-LLVY-AMC, was decreased by hydrogen peroxide. TGF-β did not increase cellular expression of heat shock protein (HSP)90, a potent inhibitor of z-LLL-AMC hydrolysis in vitro. The physiological relevance of TGF-β inhibition of proteasomal activity was studied by assessing the role of z-LLL-AMC hydrolysis on cyclin-dependent kinase inhibitor expression and cell growth. TGF-β increased expression of p27KIP1 but did not alter expression of p21WAF1 or p16INK4A. The peptide aldehyde Cbz-Leu-Leu-leucinal (LLL-CHO or MG132) potently inhibited z-LLL-AMC hydrolysis in cell extracts as well as increasing p27KIP1 and decreasing cell proliferation. Thus growth inhibition by TGF-β decreases a specific proteasomal activity via an HSP90-independent mechanism that may involve oxidative inactivation or modulation of proteasomal subunit composition and results in altered cellular expression of key cell cycle regulatory proteins such as p27KIP1.

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