transforming-growth-factor-beta and goralatide

Topics: Animals; Depression, Chemical; Diabetic Nephropathies; DNA-Binding Proteins; Extracellular Matrix; Humans; Oligopeptides; Phosphorylation; Signal Transduction; Smad Proteins; Trans-Activators; Transforming Growth Factor beta

Topics: Amino Acid Sequence; Animals; Cell Division; Chemokine CCL4; Colony-Forming Units Assay; Cytokines; DNA Replication; Growth Inhibitors; Hematopoietic Cell Growth Factors; Hematopoietic Stem Cells; Humans; Macrophage Inflammatory Proteins; Mice; Molecular Sequence Data; Monokines; Oligopeptides; Pyrrolidonecarboxylic Acid; Rats; Transforming Growth Factor beta

Topics: Amino Acid Sequence; Animals; Cell Division; Cells, Cultured; Chemokine CCL4; Colony-Forming Units Assay; Cytokines; Depression, Chemical; Hematopoietic Cell Growth Factors; Hematopoietic Stem Cells; Macrophage Inflammatory Proteins; Macrophages; Mice; Molecular Sequence Data; Monokines; Oligopeptides; Pyrrolidonecarboxylic Acid; Transforming Growth Factor beta

The overall control of the haemopoietic system is ultimately articulated at the level of stem cell proliferative regulation. An understanding of the control processes involved is central to a full understanding of the regulation of haemopoiesis in health and disease. We describe here the recent advances in understanding of the negative regulation of primitive haemopoietic cells. The possible involvement of inhibitory factors in the development of haemopoietic malignancy is discussed. The known biological functions of many of these inhibitory molecules suggests a therapeutic potential for negative regulators.

Topics: Amino Acid Sequence; Animals; Chemokine CCL4; Cytokines; Growth Inhibitors; Hematopoiesis; Humans; Macrophage Inflammatory Proteins; Molecular Sequence Data; Monokines; Oligopeptides; Peptides; Pyrrolidonecarboxylic Acid; Transforming Growth Factor beta

Two class of drugs 1) angiotensin-converting enzyme inhibitors (ACEis) and 2) angiotensin II receptor blockers (ARBs) are well-known conventional drugs that can retard the progression of chronic nephropathies to end-stage renal disease. However, there is a lack of comparative studies on the effects of ACEi versus ARB on renal fibrosis. Here, we observed that ACEi ameliorated renal fibrosis by mitigating DPP-4 and TGFβ signaling, whereas, ARB did not show. Moreover, the combination of N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP), one of the substrates of ACE, with ACEi slightly enhanced the inhibitory effects of ACEi on DPP-4 and associated-TGFβ signaling. Further, the comprehensive miRome analysis in kidneys of ACEi+AcSDKP (combination) treatment revealed the emergence of miR-29s and miR-let-7s as key antifibrotic players. Treatment of cultured cells with ACEi alone or in combination with AcSDKP prevented the downregulated expression of miR-29s and miR-let-7s induced by TGFβ stimulation. Interestingly, ACEi also restored miR-29 and miR-let-7 family cross-talk in endothelial cells, an effect that is shared by AcSDKP suggesting that AcSDKP may be partially involved in the anti-mesenchymal action of ACEi. The results of the present study promise to advance our understanding of how ACEi regulates antifibrotic microRNAs crosstalk and DPP-4 associated-fibrogenic processes which is a critical event in the development of diabetic kidney disease.

Topics: Angiotensin Receptor Antagonists; Angiotensin-Converting Enzyme Inhibitors; Animals; Cell Line; Diabetes Mellitus, Experimental; Diabetic Nephropathies; Dipeptidyl Peptidase 4; Disease Models, Animal; Drug Synergism; Gene Expression Regulation; Humans; Mice; MicroRNAs; Oligopeptides; Signal Transduction; Transforming Growth Factor beta

In this study, the bleomycin model of pulmonary fibrosis was utilized to investigate putative anti-fibrotic activity of Ac-SDKP in vivo. Male CD-1 mice received intra-tracheal bleomycin (BLEO, 1 mg/kg) instillation in the absence or presence of Ac-SDKP (a dose of 0.6 mg/kg delivered intra-peritoneally on the day of BLEO treatment, d0, followed by bi-weekly additional doses). To evaluate therapeutic effects in a subset of mice, Ac-SDKP was administered one week after BLEO instillation (d7). Animals were sacrificed at one, two, or three weeks later. Measurement of fluid and collagen content in the lung, Broncho Alveolar Lavage Fluid (BALF) analysis, lung histology, immunohistochemistry (IHC), and molecular analysis were performed. Compared to BLEO-treated mice, animals that received also Ac-SDKP (at both d0 and d7) had significantly decreased mortality, weight loss, inflammation (edema, and leukocyte lung infiltration), lung damage (histological evidence of lung injury), and fibrosis (collagen histological staining and soluble collagen content in the lung) at up to 21 days. Moreover, IHC and quantitative RT-PCR results demonstrated a significant decrease in BLEO-induced IL-17 and TGF-β expression in lung tissue. Importantly, α-SMA expression, the hallmark of myofibroblast differentiation, was also decreased. This is the first report showing not only a preventive protective role of Ac-SDKP but also its significant therapeutic effects in the bleomycin model of pulmonary fibrosis, thus supporting further preclinical and clinical studies.

Topics: Actins; Animals; Bleomycin; Chemotaxis, Leukocyte; Collagen; Cytoprotection; Disease Models, Animal; Interleukin-17; Lung; Male; Mice; Myofibroblasts; Oligopeptides; Protective Agents; Pulmonary Edema; Pulmonary Fibrosis; Time Factors; Transforming Growth Factor beta

N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) is an endogenous antifibrotic peptide. We found that suppression of AcSDKP and induction of dipeptidyl peptidase-4 (DPP-4), which is associated with insufficient levels of antifibrotic microRNA (miR)s in kidneys, were imperative to understand the mechanisms of fibrosis in the diabetic kidneys. Analyzing streptozotocin (STZ)-induced diabetic mouse strains, diabetic CD-1 mice with fibrotic kidneys could be differentiated from less-fibrotic diabetic 129Sv mice by suppressing AcSDKP and antifibrotic miRs (miR-29s and miR-let-7s), as well as by the prominent induction of DPP-4 protein expression/activity and endothelial to mesenchymal transition. In diabetic CD-1 mice, these alterations were all reversed by AcSDKP treatment. Transfection studies in culture endothelial cells demonstrated crosstalk regulation of miR-29s and miR-let-7s against mesenchymal activation program; such bidirectional regulation could play an essential role in maintaining the antifibrotic program of AcSDKP. Finally, we observed that AcSDKP suppression in fibrotic mice was associated with induction of both interferon-γ and transforming growth factor-β signaling, crucial molecular pathways that disrupt antifibrotic miRs crosstalk. The present study provides insight into the physiologically relevant antifibrotic actions of AcSDKP via antifibrotic miRs; restoring such antifibrotic programs could demonstrate potential utility in combating kidney fibrosis in diabetes.

Topics: Animals; Diabetes Mellitus; Dipeptidyl Peptidase 4; Dipeptidyl-Peptidase IV Inhibitors; Endothelial Cells; Fibroblast Growth Factor 1; Fibrosis; Humans; Interferon-gamma; Kidney; Mice; MicroRNAs; Microvessels; Models, Biological; Oligopeptides; Phosphorylation; Signal Transduction; Transforming Growth Factor beta; Up-Regulation

Thymosin β4 (Tβ4) and its amino-terminal fragment comprising N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) have been reported to act as anti-inflammatory and anti-fibrotic agents in vitro and in vivo. In recent papers, we have shown that Tβ4 exerts a widely protective role in mice treated with bleomycin, and in particular, we have demonstrated its inhibitory effects on both inflammation and early fibrosis.. In this study, the putative anti-proliferative and anti-fibrogenic effects of Tβ4 and Ac-SDKP were evaluated in vitro. In addition, the effects of Tβ4 up to 21 days were evaluated in the bleomycin mouse model of lung fibrosis.. We utilized both control and TGF-β-stimulated primary human lung fibroblasts isolated from both idiopathic pulmonary fibrosis (IPF) and control tissues. The in vivo effects of Tβ4 were assessed in CD1 mice treated with bleomycin.. In the in vitro experiments, we observed significant anti-proliferative effects of Ac-SDKP in IPF fibroblasts. In those cells, Ac-SDKP significantly inhibited TGF-β-induced α-SMA and collagen expression, hallmarks of fibroblast differentiation into myofibroblasts triggered by TGF-β. In vivo, despite its previously described protective role in mice treated with bleomycin at 7 days, Tβ4 failed to prevent fibrosis induced by the drug at 14 and 21 days.. We conclude that, compared to Tβ4, Ac-SDKP may have greater potential as an anti-fibrotic agent in the lung. Further in vivo experiments are warranted.

Topics: Animals; Anti-Inflammatory Agents; Bleomycin; Cells, Cultured; Disease Models, Animal; Fibroblasts; Humans; Lung; Male; Mice; Oligopeptides; Pneumonia; Protein Structure, Tertiary; Pulmonary Fibrosis; Thymosin; Transforming Growth Factor beta

To investigate the distribution and expression of transforming growth factor beta (TGF-β) receptors I and II, p38 mitogen-activated protein kinase (p38 MAPK), and type I and type III collagen in the lungs of rats with silicosis and cultured pulmonary fibroblasts, and to investigate the relationship of the anti-fibrosis effect of N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) with its inhibition of TGF-β receptor-mediated p38 MAPK pathway activity.. Rats were randomly divided into control group, silicosis model group, and AcSDKP treatment group (n = 10 for each group). For the model group and AcSDKP treatment group, rats were intratracheally instilled with silica to establish a silicosis model. Cultured pulmonary fibroblasts from neonatal rats were divided into control group, TGF-β1 stimulation group, TGF-β receptor inhibition group, p38 MAPK pathway inhibition group, and AcSDKP treatment group. The protein expression of TGF-β receptors I and II, p38 MAPK, and type I and type III collagen were determined by immunohistochemistry and Western blot. The mRNA expression of TGF-β receptors I and II were determined by real-time PCR. The distribution and nuclear translocation of phospho-p38 MAPK in cultured fibroblasts were determined by laser scanner confocal microscopy.. In the AcSDKP treatment group, AcSDKP reduced the expression of TGF-β receptors I and II, phospho-p38 MAPK, and type I and type III collagen to 86.12%, 41.01%, 42.63%, 89.05%, and 52.71%, respectively, of those of the silicosis model group (P < 0.05). In cultured fibroblasts, AcSDKP reduced the mRNA expression of TGF-β receptors I and II to 42.26% and 54.33%, respectively, of those of the TGF-β1 stimulation group; the protein expression of TGF-β receptors I and II, phospho-p38 MAPK, and type 1 and type III collagen was reduced to 58.14%, 51.40%, 45.6%, 58.04%, and 44.74%, respectively, of those of the TGF-β1 stimulation group. The phospho-p38 MAPK translocation from plasma to the nucleus was also inhibited; the nucleus/plasma ratio of p38 MAPK and the protein expression of type I and type III collagen were reduced to 68.60%, 58.04%, and 44.74%, respectively, of those of the TGF-β stimulation group (P < 0.05).. AcSDKP can inhibit the expression of collagen through inhibition of TGF-β receptor-mediated p38 MAPK pathway activity, and is thus able to exert anti-fibrosis effect in rats with silicosis.

Topics: Animals; Cells, Cultured; Collagen; Disease Models, Animal; Fibroblasts; Male; MAP Kinase Signaling System; Oligopeptides; p38 Mitogen-Activated Protein Kinases; Protein Serine-Threonine Kinases; Rats; Rats, Wistar; Receptor, Transforming Growth Factor-beta Type I; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Silicosis; Transforming Growth Factor beta

To investigate whether N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) can inhibit the differentiation of pulmonary fibroblasts into myofibroblasts by regulating Rho-associated coiled-coil forming protein kinase (ROCK) pathway mediated by transforming growth factor-β1 (TGF-β1).. Primary culture of pulmonary fibroblasts was performed by trypsinization method. Four generations of pulmonary fibroblasts were divided into control group, TGF-β-induced differentiation group, Y-27632 treatment group, and Ac-SDKP treatment group. The intracellular distributions of ROCK, serum response factor (SRF), and α-smooth muscle actin (α-SMA) were observed by confocal laser scanning microscopy. The protein expression of ROCK, SFR, α-SMA, and type I and type III collagen in pulmonary fibroblasts was measured by Western blot. The mRNA expression of ROCK, SFR, and α-SMA was measured by real-time quantitative PCR.. Compared with the control group, the pulmonary fibroblasts stimulated by TGF-β1 had a lot of α-SMA antibody-labeled myofilaments in parallel or cross arrangement, as observed by confocal laser scanning microscopy, and the mRNA and protein expression of ROCK, SRF, and α-SMA and protein expression of type I and type III collagen increased significantly after 6, 12, and 24 h of stimulation (P < 0.05). Compared with the TGF-β1-induced differentiation group, the Y-27632 treatment group and Ac-SDKP treatment group had significantly decreased mRNA and protein expression of ROCK, SRF, and α-SMA and protein expression of type I and type III collagen at the same time point (P < 0.05).. Ac-SDKP can inhibit the differentiation of pulmonary fibroblasts into myofibroblasts and the synthesis of collagen in rats by regulating the ROCK pathway mediated by TGF-β1. That may be one of the mechanisms by which Ac-SDKP acts against (silicotic) pulmonary fibrosis.

Topics: Actins; Animals; Animals, Newborn; Cell Differentiation; Cells, Cultured; Collagen Type I; Collagen Type III; Fibroblasts; Lung; Myofibroblasts; Oligopeptides; Rats; Rats, Wistar; rho-Associated Kinases; Serum Response Factor; Transforming Growth Factor beta

N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) is an endogenous tetrapeptide which can inhibit the differentiation, migration and activation of macrophages and suppress the proliferation of fibroblast. This study examined the effects of Ac-SDKP on the progression of lupus nephritis (LN). MRL/lpr mice received subcutaneous infusion of Ac-SDKP (1.0 mg kg(-1) d(-1)) or vehicle through implanted osmotic mini-pumps from 12 to 20 weeks until being euthanized. MRL/MpJ mice served as normal controls. The data indicative of renal inflammation and fibrosis were evaluated before and after treatment. Ac-SDKP-treated MRL/lpr mice showed reduced proteinuria and improved renal function compared with vehicle-treated controls. Ac-SDKP-treated mice demonstrated decreased inflammatory infiltrates of T cells and macrophages in the kidneys as compared to vehicle-treated animals. The treatment also inhibited the activation of NF-κB and production of TNF-α. Despite this, immune complex deposition and plasma anti-dsDNA levels were not statistically different between the two groups. In addition, the treatment inhibited renal expression of TGF-β1, α-SMA and fibronectin as well as the phosphorylation of Smad2/3. Ac-SDKP treatment ameliorated LN through exerting anti-inflammatory and anti-fibrotic effects on MRL/lpr mice, providing therapeutic potential for halting the progression of LN.

Topics: Actins; Animals; Antibodies, Antinuclear; Body Weight; Chemokine CCL2; Drug Administration Schedule; Female; Fibronectins; Gene Expression Regulation; Inflammation; Kidney; Leukocytes; Lupus Nephritis; Mice; Mice, Inbred MRL lpr; NF-kappa B; Oligopeptides; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) is an endogenous tetrapeptide which has antifibrogenic effects at physiological concentrations in various tissues. AcSDKP is produced locally in the liver, however, little is known about its biological effect in this organ. We hypothesize that basal levels of endogenous AcSDKP decrease during the development of liver fibrosis and preservation of basal AcSDKP attenuates liver fibrosis.. Endogenous levels of AcSDKP in the liver were measured by enzyme immunoassay after 2, 6, and 10 weeks of carbon tetrachloride (CCl(4))-induced liver fibrosis in rats. Subcutaneous osmotic pump infusion of vehicle or AcSDKP (800 microg/kg/day) was administered to CCl(4)-treated rats for 8 weeks to study the effect of exogenous AcSDKP on liver fibrosis. The effect of AcSDKP on profibrogenic properties of hepatic stellate cells was studied in vitro.. Endogenous AcSDKP was significantly decreased in the liver of CCl(4)-treated rats. Chronic AcSDKP infusion preserved basal levels of AcSDKP and reduced liver injury, inflammation, fibrosis, and profibrogenic transforming growth factor-beta signaling. This was demonstrated by decreased aminotransferase serum levels, CD45 positive cells, collagen accumulation, alpha-smooth muscle actin positivity, transforming growth factor-beta1, phosphorylated Smad2/3 protein, increased bone morphogenetic protein-7, and phosphorylated Smad1/5/8. Further, AcSDKP exerts antifibrogenic effects on hepatic stellate cells (HSCs) by downregulation of HSC activation in vitro.. Maintaining physiological levels of AcSDKP is critical in negatively regulating the development of fibrosis in chronic liver injury. Preservation of AcSDKP may be a useful therapeutic approach in the management of liver fibrosis.

Topics: Actins; Animals; Carbon Tetrachloride Poisoning; Collagen; Gene Expression; Hepatic Stellate Cells; In Vitro Techniques; Liver Cirrhosis, Experimental; Male; Oligopeptides; Rats; Rats, Sprague-Dawley; Signal Transduction; Transforming Growth Factor beta

to investigate the effect of N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) on the expressions of c-Raf, ERK1/2 and TGF-β1 in the lung of rats with silicosis, thus to investigate the regulating of AcSDKP on the Ras-Raf-ERK1/2 signal transduction pathway.. rats were instilled with silica through trachea as silicotic models and administered AcSDKP in the experiment. Rats were divided into 6 groups randomly, 10 rats in each group: Control 1 and 2 of silicotic model: each rat was intratracheally instilled with 1.0 ml normal sodium and was killed after 4 or 8 weeks; Silicotic model 1 and Silicotic model 2: each rat was intratracheally instilled with 1ml silica suspension and was killed after 4 or 8 weeks; Anti-fibrosis treatment of AcSDKP: after each rat was intratracheally instilled with 1ml silica suspension for 4 weeks, AcSDKP 800 microg × kg(-1) × d(-1) was administered into every rat and rats were killed at the eighth week; Preventing fibrosis treatment of AcSDKP: after AcSDKP 800 microg × kg(-1) × d(-1) was administered into every rat for 48 hours, each rat was intratracheally instilled with 1.0 ml silica suspension and rats were killed at the eighth week. The expression of c-Raf, phospho-c-Raf, ERK1/2, phospho-ERK1/2 and TGF-β1 was measured by immunohistochemistry and western blot assay.. compared with the corresponding control groups, the expressions of phospho-c-Raf, phospho-ERK1/2 and TGF-β1 increased in the lung tissue of the silicotic models. Compared with the corresponding model groups, after administration AcSDKP, the expressions of phospho-c-Raf, phospho-ERK1/2 and TGF-β1 in the lung tissue reduced obviously. In anti-fibrosis treatment of AcSDKP group, expressions of phospho-c-Raf, phospho-ERK1/2 and TGF-β1 decreased to 52.25%, 51.72% and 67.74% compared with those of the silicotic model 1, and expressions of phospho-c-Raf, phospho-ERK1/2 and TGF-β1 decreased to 49.37%, 55.76%, 65.63% compared with those of the silicotic model 2; In preventing fibrosis treatment of AcSDKP group, expressions of phospho-c-Raf, phospho-ERK1/2 and TGF-β1 decreased to 54.64%, 55.76% and 78.91% compared with those of the silicotic model 2 (P < 0.05) while the expressions of c-Raf and ERK1/2 were not different significantly among each groups.. AcSDKP possibly plays an important role in anti-silicotic fibrosis by blocking the TGF-β-induced Ras-Raf-ERK1/2 signal transduction pathway.

Topics: Animals; Lung; Male; Mitogen-Activated Protein Kinase 3; Oligopeptides; Proto-Oncogene Proteins c-raf; Rats; Rats, Wistar; Signal Transduction; Silicosis; Transforming Growth Factor beta

Galectin-3 (Gal-3) is secreted by activated macrophages. In hypertension, Gal-3 is a marker for hypertrophic hearts prone to develop heart failure. Gal-3 infused in pericardial sac leads to cardiac inflammation, remodeling, and dysfunction. N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP), a naturally occurring tetrapeptide, prevents and reverses inflammation and collagen deposition in the heart in hypertension and heart failure postmyocardial infarction. In the present study, we hypothesize that Ac-SDKP prevents Gal-3-induced cardiac inflammation, remodeling, and dysfunction, and these effects are mediated by the transforming growth factor (TGF)-beta/Smad3 signaling pathway. Adult male rats were divided into four groups and received the following intrapericardial infusion for 4 wk: 1) vehicle (saline, n = 8); 2) Ac-SDKP (800 microg x kg(-1) x day(-1), n = 8); 3) Gal-3 (12 microg/day, n = 7); and 4) Ac-SDKP + Gal-3 (n = 7). Left ventricular ejection fraction, cardiac output, and transmitral velocity were measured by echocardiography; inflammatory cell infiltration, cardiomyocyte hypertrophy, and collagen deposition in the heart by histological and immunohistochemical staining; and TGF-beta expression and Smad3 phosphorylation by Western blot. We found that, in the left ventricle, Gal-3 1) enhanced macrophage and mast cell infiltration, increased cardiac interstitial and perivascular fibrosis, and causes cardiac hypertrophy; 2) increased TGF-beta expression and Smad3 phosphorylation; and 3) decreased negative change in pressure over time response to isoproterenol challenge, ratio of early left ventricular filling phase to atrial contraction phase, and left ventricular ejection fraction. Ac-SDKP partially or completely prevented these effects. We conclude that Ac-SDKP prevents Gal-3-induced cardiac inflammation, fibrosis, hypertrophy, and dysfunction, possibly via inhibition of the TGF-beta/Smad3 signaling pathway.

Topics: Animals; Anti-Inflammatory Agents; Blood Pressure; Body Weight; Cardiac Output; Cardiomegaly; Cardiotonic Agents; Collagen; Disease Models, Animal; Echocardiography, Doppler; Fibrosis; Galectin 3; Heart Rate; Hemodynamics; Inflammation; Infusions, Parenteral; Isoproterenol; Macrophages; Male; Mast Cells; Myocardial Contraction; Myocardium; Oligopeptides; Phosphorylation; Rats; Rats, Sprague-Dawley; Signal Transduction; Smad3 Protein; Stroke Volume; Time Factors; Transforming Growth Factor beta; Ventricular Function, Left; Ventricular Remodeling

To investigate whether the effect of N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) on transforming growth factor beta (TGF-beta1) and connective tissues growth factor (CTGF) was involved in AcSDKP's antifibrotic effect on the rats with silicosis.. Rats were divided into 6 groups randomly, 10 rats in each group: Control of silicotic model: 1.0 ml normal sodium and was killed after 4 or 8 weeks; Silicotic model 1: 50 mg/ml silica suspension and was killed after 4 weeks; Silicotic model 2: 50 mg/ml silica suspension and was killed after 8 weeks; Anti-fibrosis treatment of AcSDKP: after each rat was intratracheally instilled with 50 mg/ml silica suspension for 4 weeks, AcSDKP 800 microg/(kg x d) was administered into every rat and rats were killed at the 8 weeks; Preventing fibrosis treatment of AcSDKP: after AcSDKP [800 microg/(kg x d)] was administered into every rat for 48 hours, each rat was intratracheally instilled with 50 mg/ml silica suspension and rats were killed at the 8 weeks. Lung fibrosis in morphology was observed by HE staining. The expressions of TGF-beta1 and CTGF in lung were observed by immunohistochemistry. The mRNA expressions of TGF-beta1 and CTGF in lung were observed by real-time PCR.. In anti-fibrosis treatment of AcSDKP group, protein expression of TGF-beta1 and CTGF were (0.244 +/- 0.016) and (0.241 +/- 0.017) respectively, and significantly lower that those in the silicotic model 1 and 2 groups; mRNA expressions of TGF-beta1 and CTGF decreased, mRNA expressions of CTGF were significantly lower that those in the silicotic model 1 and 2 groups (P < 0.05); In preventing fibrosis treatment of AcSDKP group, protein expression and mRNA expression of TGF-beta1 were significantly lower that those in the silicotic model 2 group (P < 0.05).. AcSDKP can decrease the expressions of TGF-beta1 and CTGF in lung tissues of the rats with experimentally induced pulmonary fibrosis.

Topics: Animals; Connective Tissue Growth Factor; Disease Models, Animal; Lung; Male; Oligopeptides; Pulmonary Fibrosis; Rats; Rats, Sprague-Dawley; Silicosis; Transforming Growth Factor beta

Topics: Animals; Animals, Newborn; Cells, Cultured; Fibroblasts; Matrix Metalloproteinase 1; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Myocardium; Oligopeptides; Rats; Rats, Wistar; Transforming Growth Factor beta

Topics: Carpal Tunnel Syndrome; Gene Expression Regulation; Immunosuppressive Agents; NF-kappa B; Nucleic Acid Synthesis Inhibitors; Oligopeptides; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Rifampin; Smad2 Protein; Smad3 Protein; Transcription Factor AP-1; Transforming Growth Factor beta

N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP), which is hydrolyzed by angiotensin-converting enzyme, is a natural regulator of hematopoiesis. Here it is shown that Ac-SDKP inhibits TGF-beta action in mesangial cells. Because TGF-beta is thought to play a pivotal role in the development and progression of glomerulonephritis, the therapeutic effects of Ac-SDKP on an established model of renal dysfunction and histologic alteration in Wistar-Kyoto rats with anti-glomerular basement membrane nephritis was examined. Fourteen days after the induction of anti-glomerular basement membrane nephritis, the rats were treated subcutaneously with Ac-SDKP at a dose of 1 mg/kg per d for 4 wk. Treatment with Ac-SDKP significantly improved proteinuria and renal dysfunction, including increased plasma blood urea nitrogen and creatinine levels and decreased creatinine clearance. Histologic examination showed severe glomerulosclerosis and interstitial fibrosis in the vehicle-treated rats, whereas these histologic injuries were significantly ameliorated in rats that were treated with Ac-SDKP. The histologic improvements were accompanied by the suppression of gene and protein expression of fibronectin, interstitial collagen, and TGF-beta1 in the nephritic kidney. Furthermore, treatment with Ac-SDKP resulted in the inhibition of Smad2 phosphorylation, an increase in Smad7 expression in the kidney, and reduction of macrophage accumulation into the glomeruli and tubulointerstitium in nephritic rats. In conclusion, Ac-SDKP significantly ameliorated the progression of renal dysfunction and fibrosis even after the establishment of nephritis. The inhibitory effect of Ac-SDKP was mediated in part by the inhibition of TGF-beta/Smad signal transduction and the inflammatory response. These findings suggest that Ac-SDKP treatment may be a novel and useful therapeutic strategy for the treatment of progressive renal diseases.

Topics: Analysis of Variance; Animals; Anti-Glomerular Basement Membrane Disease; Biopsy, Needle; Blood Chemical Analysis; Blotting, Western; Disease Models, Animal; Disease Progression; Dose-Response Relationship, Drug; Drug Administration Schedule; Fibrosis; Immunohistochemistry; Injections, Subcutaneous; Kidney Function Tests; Male; Oligopeptides; Probability; Random Allocation; Rats; Rats, Inbred WKY; Reverse Transcriptase Polymerase Chain Reaction; Sensitivity and Specificity; Statistics, Nonparametric; Transforming Growth Factor beta; Urinalysis

We have previously reported that N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP), which is a tetrapeptide hydrolyzed by ACE, inhibits the transforming growth factor-beta (TGF-beta)-induced expression of extracellular matrix proteins via inhibition of the Smad signaling in human mesangial cells. To test in vivo the antifibrotic efficacy of Ac-SDKP, we examined whether long-term Ac-SDKP treatment can prevent renal insufficiency and glomerulosclerosis in diabetic db/db mice. Diabetic db/db mice or nondiabetic db/m mice were treated with Ac-SDKP for 8 weeks using osmotic minipumps. The treatment with Ac-SDKP increased plasma Ac-SDKP concentrations by approximately threefold in both groups but did not affect the blood glucose levels. Histologically, the increased glomerular surface area, mesangial matrix expansion, and overproduction of extracellular matrix proteins in db/db mice were significantly inhibited by Ac-SDKP. Furthermore, Ac-SDKP treatment normalized the increased plasma creatinine value in db/db mice, whereas the albuminuria in Ac-SDKP-treated db/db mice was somewhat decreased as compared with nontreated db/db mice, although the difference was not statistically significant. In addition, the nuclear translocation of Smad3 was inhibited by Ac-SDKP. These results demonstrate that long-term Ac-SDKP treatment ameliorates renal insufficiency and glomerulosclerosis in db/db mice via inhibition of TGF-beta/Smad pathway, suggesting that Ac-SDKP could be useful in the treatment of diabetic nephropathy.

Topics: Animals; Collagen Type IV; Diabetic Nephropathies; DNA-Binding Proteins; Extracellular Matrix Proteins; Fibronectins; Gene Expression; Glomerular Mesangium; Male; Mice; Mice, Knockout; Oligopeptides; Renal Insufficiency; Smad3 Protein; Trans-Activators; Transforming Growth Factor beta

Inflammation may play an important role in the pathogenesis of cardiac fibrosis in heart failure (HF) after myocardial infarction (MI). N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) is a naturally occurring antifibrotic peptide whose plasma concentration is increased 4- to 5-fold by angiotensin-converting enzyme inhibitors. We tested the hypothesis that in rats with HF after MI, Ac-SDKP acts as an anti-inflammatory cytokine, preventing and also reversing cardiac fibrosis in the noninfarcted area (reactive fibrosis), and thus affording functional improvement. We found that Ac-SDKP significantly decreased total collagen content in the prevention group from 23.7+/-0.9 to 15.0+/-0.7 microg/mg and in the reversal group from 22.6+/-2.2 to 14.4+/-1.6 (P<0.01). Interstitial collagen volume fraction and perivascular collagen were likewise significantly reduced. We also found that infiltrating macrophages were reduced from 264.7+/-8.1 to 170.2+/-9.2/mm2, P<0.001 (prevention), and from 257.5+/-9.1 to 153.1+/-8.5 mm2, P<0.001 (reversal), while transforming growth factor (TGF)-beta-positive cells were decreased from 195.6+/-8.4 to 129.6+/-5.7/mm2, P<0.01 (prevention), and from 195.6+/-8.4 to 130.7+/-10.8/mm2, P<0.01 (reversal). Ac-SDKP did not alter either blood pressure or left ventricular hypertrophy (LVH); however, it depressed systolic cardiac function in the prevention study while having no significant effect in the reversal group. We concluded that Ac-SDKP has an anti-inflammatory effect in HF that may contribute to its antifibrotic effect; however, this decrease in fibrosis without changes in LVH was not accompanied by an improvement in cardiac function.

Topics: Animals; Anti-Inflammatory Agents; Blood Pressure; Cardiac Output, Low; Cell Movement; Collagen; Fibrosis; Heart; Heart Rate; Inflammation; Macrophages; Male; Myocardial Infarction; Myocardium; Oligopeptides; Rats; Rats, Inbred Lew; Transforming Growth Factor beta

N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) is a potent natural inhibitor of hematopoietic stem cell proliferation which is degraded mainly by angiotensin-converting enzyme (ACE). In vitro, Ac-SDKP inhibits collagen production by cardiac fibroblasts; while in vivo it blocks collagen deposition in the left ventricle (LV) of rats with hypertension or myocardial infarction (MI). In addition, it reportedly prevents and reverses macrophage infiltration in the LV of rats with MI. We tested the hypothesis that when Ac-SDKP is infused at doses that cause plasma concentrations similar to those observed after ACE inhibition, it mimics the anti-inflammatory and antifibrotic effects of ACE inhibitors (ACEi) in the heart, and, further, that these effects are independent of changes in blood pressure.. Rats were divided into five groups: (1) controls, (2) Ang II (750 microg/kg per day, s.c.), (3) Ang II + captopril (100 mg/kg per day in drinking water), (4) Ang II + Ac-SDKP (400 microg/kg per day, s.c.), and (5) Ang II + Ac-SDKP (800 microg/kg per day, s.c.). We measured LV cell proliferation, inflammatory cell infiltration, cytokine expression, hypertrophy and fibrosis.. Plasma Ac-SDKP was five-fold higher in rats given ACEi and four- and ten-fold higher in rats given 400 and 800 microg/kg per day Ac-SDKP, respectively. ACEi significantly decreased Ang II-induced cell proliferation (Ki-67), LV macrophage/mast cell infiltration, transforming growth factor-beta, connective tissue growth factor and collagen deposition without affecting hypertension, LV hypertrophy or myocyte cross-sectional area, and these effects were mimicked by exogenous Ac-SDKP (400 microg/kg per day) which raised plasma Ac-SDKP to levels similar to ACEi. BP was not decreased by either ACEi or Ac-SDKP.. We concluded that Ac-SDKP may be an important mediator of the anti-inflammatory and antifibrotic effects of ACEi in hypertension independent of its hemodynamic effects.

Topics: Angiotensin-Converting Enzyme Inhibitors; Animals; Blood Pressure; Captopril; Cell Division; Collagen; Connective Tissue Growth Factor; Drug Therapy, Combination; Fibrosis; Growth Inhibitors; Heart Rate; Hypertension; Hypertrophy, Left Ventricular; Immediate-Early Proteins; Intercellular Signaling Peptides and Proteins; Kidney; Macrophages; Male; Mast Cells; Monocytes; Myocardium; Myocytes, Cardiac; Oligopeptides; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta

Although increased activity of angiotensin-converting enzyme (ACE) has been associated with increased cardiac collagen, no studies to date have established a direct cause-and-effect relation between the two.. We used transgenic rats that overexpress human ACE selectively in the myocardium. Two independent heterozygous transgenic rat lines were studied, one expressing 2 to 3 copies (L1172) and the other expressing 5 to 10 copies (L1173) of the ACE transgene. These rats were normotensive but developed a proportionate increase in myocardial collagen depending on the ACE gene dose (up to 2.5-fold, P<0.01), but cardiac angiotensin II levels remained normal, whereas collagen content reversed to control levels on ACE inhibition. To explain these changes, we investigated N-acetyl-Ser-Asp-Lys-Pro (AcSDKP), an alternative substrate that is catabolized exclusively by ACE. Increased cardiac expression of ACE was paralleled by a reciprocal decrease in cardiac AcSDKP and a proportionate increase in phosphorylated Smad2 and Smad3, all of which normalized after both ACE inhibition and AcSDKP infusion. Furthermore, a functional link of this signaling cascade was demonstrated, because AcSDKP inhibited Smad3 phosphorylation in a dose-dependent manner in cultured cardiac fibroblasts and in vivo.. Our findings suggest that increased cardiac ACE activity can increase cardiac collagen content by degradation of AcSDKP, an inhibitor of the phosphorylation of transforming growth factor-beta signaling molecules Smad2 and Smad3. This implies that the antifibrotic effects of ACE inhibitors are mediated in part by increasing cardiac AcSDKP, with subsequent inhibition of Smad 2/3 phosphorylation.

Topics: Activin Receptors, Type I; Angiotensin II; Animals; Animals, Genetically Modified; Binding, Competitive; Collagen; DNA-Binding Proteins; DNA, Complementary; Fibrosis; Humans; Hypertrophy, Left Ventricular; Myocardium; Oligopeptides; Peptidyl-Dipeptidase A; Phosphorylation; Protein Processing, Post-Translational; Protein Serine-Threonine Kinases; Rats; Rats, Sprague-Dawley; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Smad2 Protein; Smad3 Protein; Substrate Specificity; Trans-Activators; Transforming Growth Factor beta; Transforming Growth Factor beta1

Recent large clinical trials indicate that angiotensin-converting enzyme inhibitors (ACE-I) attenuate the detrimental outcome of progressive renal disease. The hemoregulatory tetrapeptide N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP, AcSDKP) is hydrolyzed by ACE, and plasma Ac-SDKP level is increased by fivefold after treatment with ACE-I. Ac-SDKP was found to ameliorate cardiac and renal fibrosis in hypertensive animal models. However, the molecular mechanisms by which Ac-SDKP mediates anti-fibrotic effects remain unclear. This study is an examination of the interaction between Ac-SDKP and transforming growth factor-beta (TGF-beta), one of the key cytokines in the progression of renal disease, in human mesangial cells. Ac-SDKP inhibited TGF-beta1-induced plasminogen activator inhibitor-1 (PAI-1) and alpha2 (I) collagen mRNA. Ac-SDKP suppressed not only TGF-beta1-induced Smad2 phosphorylation at Ser-465/467 in a dose-dependent manner, but also the nuclear accumulation of receptor-regulated Smads (R-Smad), Smad2 and Smad3. As expected, Ac-SDKP inhibited TGF-beta-responsive Smad-dependent luciferase reporters, 3TP-luc and 4xSBE-luc. Immunofluorescence analysis revealed that the inhibitory Smad, Smad7, was exported to the cytoplasm from the nucleus by the treatment with Ac-SDKP. These findings provide novel evidence that Ac-SDKP inhibits TGF-beta signal transduction through the suppression of R-Smad activation via nuclear export of Smad7, highlighting an alternative mechanism involved in the reno-protective efficacy of ACE-I.

Topics: Cell Culture Techniques; DNA-Binding Proteins; Fibrosis; Glomerular Mesangium; Growth Inhibitors; Humans; Oligopeptides; Plasminogen Activator Inhibitor 1; RNA, Messenger; Signal Transduction; Smad Proteins; Smad7 Protein; Trans-Activators; Transforming Growth Factor beta

N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) is a natural substrate for the N-terminal active site of angiotensin-converting enzyme (ACE). We previously reported that Ac-SDKP prevented cardiac fibrosis in rats with renovascular or aldosterone-salt hypertension. However, it is not clear whether Ac-SDKP reverses cardiac fibrosis in hypertension, nor the mechanism(s) involved. In the present study, we tested the hypothesis that Ac-SDKP reversal of hypertension-induced cardiac fibrosis involves a decrease in transforming growth factor-beta (TGF-beta) and/or connective tissue growth factor (CTGF). In 2-kidney, 1-clip (2K-1C) hypertensive rats, Ac-SDKP at 400 or 800 microg/kg per day SC was started 8 weeks after hypertension and cardiac fibrosis were established and was continued for 8 weeks. Left ventricular (LV) collagen in rats with 2K-1C plus vehicle at 8 and 16 weeks after clipping was similar but higher than in the sham group (P<0.05). Ac-SDKP at 400 and 800 microg/kg per day, which increased plasma Ac-SDKP 2- and 5-fold, respectively, reversed the increase in LV collagen in a dose-dependent manner. The mechanism by which Ac-SDKP reverses LV fibrosis does not appear to depend on ACE inhibition by Ac-SDKP, since we found that Ac-SDKP at various doses did not affect blood pressure responses to exogenous angiotensin I or bradykinin. However, Ac-SDKP reversed the increase in LV TGF-beta and CTGF compared with rats with 2K-1C plus vehicle (P<0.005). We concluded that in hypertension, Ac-SDKP reverses cardiac fibrosis, perhaps due in part to a decrease in TGF-beta and CTGF in the heart.

Topics: Angiotensins; Animals; Blood Pressure; Body Weight; Bradykinin; Collagen; Connective Tissue Growth Factor; Fibrosis; Heart Ventricles; Hypertension, Renovascular; Immediate-Early Proteins; Intercellular Signaling Peptides and Proteins; Male; Myocardium; Oligopeptides; Organ Size; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta

N-Acetyl-Ser-Asp-Lys-Pro (AcSDKP) is a specific substrate for the N-terminal site of ACE and increases 5-fold during ACE inhibitor therapy. It is known to inhibit the proliferation of hematopoietic stem cells and has also recently been reported to inhibit the growth of cardiac fibroblasts. We investigated its mode of action in cardiac fibroblasts by assessing its influence on transforming growth factor beta(1) (TGFbeta1)-mediated Smad signaling. AcSDKP inhibited the proliferation of isolated cardiac fibroblasts (P<0.05) but significantly stimulated the proliferation of vascular smooth muscle cells. Flow cytometry of rat cardiac fibroblasts treated with AcSDKP showed significant inhibition of the progression of cells from G0/G1 phase to S phase of the cell cycle. In cardiac fibroblasts transfected with a Smad-sensitive luciferase reporter construct, AcSDKP decreased luciferase activity by 55+/-9.7% (P=0.01). Moreover, phosphorylation and nuclear translocation of Smad2 was decreased in cardiac fibroblasts treated with AcSDKP. To conclude, AcSDKP inhibits the growth of cardiac fibroblasts and also inhibits TGFbeta1-stimulated phosphorylation of Smad2. Because AcSDKP increases substantially during ACE inhibitor therapy, this suggests a novel pathway independent of angiotensin II, by which ACE inhibitors can inhibit cardiac fibrosis.

Topics: Animals; Animals, Newborn; Biological Transport; Cattle; Cell Division; Cell Line; Cell Nucleus; DNA-Binding Proteins; Dose-Response Relationship, Drug; Fetal Blood; Fibroblasts; G1 Phase; Luciferases; Muscle, Smooth, Vascular; Myocardium; Oligopeptides; Phosphorylation; Promoter Regions, Genetic; Rats; Recombinant Fusion Proteins; Response Elements; S Phase; Smad2 Protein; Trans-Activators; Transforming Growth Factor beta; Transforming Growth Factor beta1

The long-term culture (LTC) system has been useful for analyzing mechanisms by which stromal cells regulate the proliferative activity of primitive normal, but not chronic myeloid leukemia (CML), hematopoietic progenitor cells. In previous studies, we identified two endogenous inhibitors in this system. One is transforming growth factor-beta (TGF-beta), which is equally active on primitive normal and CML progenitors. The other we now show to be monocyte chemoattractant protein-1 (MCP-1). Thus, MCP-1, when added to LTC, blocked the activation of primitive normal progenitors but did not arrest the cycling of primitive CML progenitors. Moreover, the endogenous inhibitory activity of LTC stromal layers could be overcome by the addition of neutralizing antibodies to MCP-1, but not to macrophage inflammatory protein-1alpha (MIP-1alpha). However, neither of these antibodies antagonized the inhibitory activity of NAc-Ser-Asp-Lys-Pro (AcSDKP) on primitive normal but not CML progenitor cycling in this system. Moreover, none of six other -C-C- or -C-X-C- chemokines, previously shown to inhibit primitive normal human CFC proliferation in semisolid assays, were found to act as negative regulators when added to normal LTC. These results provide further support for the concept that primitive CML progenitor cell proliferation is deregulated when these cells are exposed to limiting concentrations of multiple inhibitors, only some of which have differential actions on normal and Ph+/BCR-ABL+ cells.

Topics: Bone Marrow; Cell Cycle; Cell Division; Cells, Cultured; Chemokine CCL2; Chemokine CCL3; Chemokine CCL4; Cytokines; Fusion Proteins, bcr-abl; Hematopoiesis; Hematopoietic Stem Cells; Humans; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Macrophage Inflammatory Proteins; Neoplastic Stem Cells; Oligopeptides; Recombinant Proteins; Signal Transduction; Stromal Cells; Transforming Growth Factor beta; Tumor Cells, Cultured

We have compared the effects of AcSDKP, Thymosin beta4 (Tbeta4), MIP1alpha and TGFbeta on acute myeloid leukemia (AML) and B-lineage acute lymphoid leukemia (B-ALL) cells using liquid cultures in the presence of GM-CSF, IL-3 and SCF for AML cells and IL-3 and IL-7 for ALL cells. Each molecule was added daily and cell proliferation was evaluated on day 3 by thymidine incorporation. Whereas TGFbeta was found inhibitory in all the AML and B-ALL cases studied, MIP1alpha was inhibitory in 6/12 AML cases and had no effect on B-ALL cells. AcSDKP and Tbeta4 showed an inhibitory effect in a few cases but only at high doses which were inactive on normal cells. Thus, our study not only confirms the effect of TGFbeta, MIP1alpha and AcSDKP on AML cells but also provides new data concerning their effect on B-ALL and the possible inhibitory effect of AcSDKP at high doses. Furthermore, we show for the first time the effect of Tbeta4 on leukemic cells. Altogether, our data indicate differences of sensitivity of leukemic cells to negative regulators, some leukemias being inhibited by one or several of these molecules whereas others were unresponsive to all used. The clinical relevance of these observations still remains to be determined.

Topics: Adolescent; Adult; Aged; Burkitt Lymphoma; Cell Division; Chemokine CCL4; Child; Child, Preschool; Female; Humans; Infant; Leukemia, Myeloid, Acute; Macrophage Inflammatory Proteins; Male; Middle Aged; Oligopeptides; Thymosin; Transforming Growth Factor beta; Tumor Cells, Cultured

The aim of this study was to compare the inhibitory effect of the tetrapeptide AcSDKP, tumor necrosis factor-alpha (TNF-alpha), which contains the sequence of the peptide, transforming growth factor-beta (TGF-beta), and macrophage inflammatory protein-1 alpha (MIP-1 alpha) on sorted CD34+ cells using both proliferation and clonogenic assays. Although a short treatment with any of the molecules decreased the growth of colony-forming unit granulocyte/macrophage (CFU-GM) and burst-forming unit-erythroid (BFU-E) progenitors (except for TNF-alpha as it is a greater inhibitor for CFU-GM), further experiments using a 6-day liquid culture in the presence of a combination of growth factors (recombinant human interleukin-3 [rhIL-3], IL-6, IL-1 beta, GM colony-stimulating factor [GM-CSF], G-CSF, erythropoeitin [Epo], and stem cell factor [SCF]) allowed us to determine a number of differences between their effects: 1) TGF-beta and TNF-alpha induced a stronger decrease in the proliferation and clonogenicity of CD34+ subsets than MIP-1 alpha and AcSDKP, 2) the dose-response curves appeared different, and 3) contrary to TGF-beta and TNF-alpha, AcSDKP and MIP-1 alpha required repeated addition to induce inhibition. Therefore, our data clearly show that while the inhibitory effect of TNF-alpha and AcSDKP appeared to be different, there is a close similarity in the effect of AcSDKP and MIP-1 alpha on normal human progenitor response to the combination of growth factors used.

Topics: Antigens, CD; Antigens, CD34; Bone Marrow; Cell Division; Cell Separation; Cells, Cultured; Chemokine CCL4; Cytokines; Hematopoiesis; Hematopoietic Stem Cells; Humans; Macrophage Inflammatory Proteins; Monokines; Oligopeptides; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha