Assuntos
Síndrome de Brugada/etiologia , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Canais de Cátion TRPM/genética , Arritmias Cardíacas/genética , Arritmias Cardíacas/fisiopatologia , Síndrome de Brugada/genética , Humanos , Fator de Crescimento Derivado de Plaquetas/farmacologia , Canais de Cátion TRPM/efeitos adversosRESUMO
Fibrotic diseases pose significant clinical challenges due to their broadness and complexity. Thus, a better understanding of fibrogenesis and the development of more effective treatments is imperative. Recent evidence suggests a significant antifibrotic potential of an endogenous glycoprotein, endostatin. While endostatin has been widely studied for its role as an anticancer adjuvant by inhibiting tumor angiogenesis, its possible implication in fibrosis remains largely unclear. Here, we review the role of endostatin in various cellular processes and highlight its antifibrotic activity. We hypothesize that endostatin conveys a homeostatic function in the process of fibrosis by regulating (a) TGF-ß1 and its downstream signaling; (b) RhoA/ROCK pathway; (c) NF-κB signaling pathway; (d) expression of EGR-1; (e) PDGF/PDGFR pathway; (f) autophagy-related pathways; (g) pathways associated with cell proliferation and apoptosis. Finally, we propose a schematic model of the antifibrotic roles and mechanisms of endostatin; also, we outline future research directions of endostatin and aim to present a potential therapeutic approach for fibrosis.
Assuntos
Endostatinas/farmacologia , Fibrose/tratamento farmacológico , Fator de Crescimento Transformador beta1/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , NF-kappa B/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Proteína rhoA de Ligação ao GTP/efeitos dos fármacosRESUMO
FGF-2 displays multifarious functions in regulation of angiogenesis and vascular remodeling. However, effective drugs for treating FGF-2+ tumors are unavailable. Here we show that FGF-2 modulates tumor vessels by recruiting NG2+ pricytes onto tumor microvessels through a PDGFRß-dependent mechanism. FGF-2+ tumors are intrinsically resistant to clinically available drugs targeting VEGF and PDGF. Surprisingly, dual targeting the VEGF and PDGF signaling produces a superior antitumor effect in FGF-2+ breast cancer and fibrosarcoma models. Mechanistically, inhibition of PDGFRß ablates FGF-2-recruited perivascular coverage, exposing anti-VEGF agents to inhibit vascular sprouting. These findings show that the off-target FGF-2 is a resistant biomarker for anti-VEGF and anti-PDGF monotherapy, but a highly beneficial marker for combination therapy. Our data shed light on mechanistic interactions between various angiogenic and remodeling factors in tumor neovascularization. Optimization of antiangiogenic drugs with different principles could produce therapeutic benefits for treating their resistant off-target cancers.
Assuntos
Inibidores da Angiogênese/farmacologia , Fator 2 de Crescimento de Fibroblastos/efeitos dos fármacos , Fator 2 de Crescimento de Fibroblastos/metabolismo , Neoplasias/tratamento farmacológico , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/efeitos dos fármacos , Animais , Biomarcadores Tumorais , Pressão Sanguínea , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Permeabilidade Capilar , Proliferação de Células , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Quimioterapia Combinada , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias/patologia , Neovascularização Patológica/tratamento farmacológico , Fator de Crescimento Derivado de Plaquetas/metabolismo , Receptor beta de Fator de Crescimento Derivado de Plaquetas , Transdução de Sinais/efeitos dos fármacos , Hipóxia Tumoral , Microambiente Tumoral/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
BACKGROUND: Intestinal fibrosis is a hallmark of Crohn's disease. Here, we investigated the impact of several putative antifibrotic compounds on the expression of fibrosis markers using murine precision-cut intestinal slices. METHODS: Murine precision-cut intestinal slices were cultured for 48 hours in the presence of profibrotic and/or antifibrotic compounds. The fibrotic process was studied on gene and protein level using procollagen 1a1 (Col1α1), heat shock protein 47 (Hsp47), fibronectin (Fn2), and plasminogen activator inhibitor-1 (Pai-1). The effects of potential antifibrotic drugs mainly inhibiting the transforming growth factor ß (TGF-ß) pathway (eg, valproic acid, tetrandrine, pirfenidone, SB203580, and LY2109761) and compounds mainly acting on the platelet-derived growth factor (PDGF) pathway (eg, imatinib, sorafenib, and sunitinib) were assessed in the model at nontoxic concentrations. RESULTS: Murine precision-cut intestinal slices remained viable for 48 hours, and an increased expression of fibrosis markers was observed during culture, including Hsp47, Fn2, and Pai-1. Furthermore, TGF-ß1 stimulated fibrogenesis, whereas PDGF did not have an effect. Regarding the tested antifibrotics, pirfenidone, LY2109761, and sunitinib had the most pronounced impact on the expression of fibrosis markers, both in the absence and presence of profibrotic factors, as illustrated by reduced levels of Col1α1, Hsp47, Fn2, and Pai-1 after treatment. Moreover, sunitinib significantly reduced Hsp47 and Fn2 protein expression and the excretion of procollagen 1. CONCLUSIONS: Precision-cut intestinal slices can successfully be used as a potential preclinical screening tool for antifibrotic drugs. We demonstrated that sunitinib reduced the expression of several fibrosis markers, warranting further evaluation of this compound for the treatment of intestinal fibrosis.
Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Inibidores Enzimáticos/farmacologia , Fármacos Gastrointestinais/farmacologia , Intestinos/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Colágeno Tipo I/efeitos dos fármacos , Cadeia alfa 1 do Colágeno Tipo I , Doença de Crohn/tratamento farmacológico , Doença de Crohn/patologia , Fibronectinas/efeitos dos fármacos , Fibrose/tratamento farmacológico , Fibrose/patologia , Proteínas de Choque Térmico HSP47/efeitos dos fármacos , Intestinos/patologia , Camundongos , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Serpina E2/efeitos dos fármacos , Sunitinibe/farmacologia , Fator de Crescimento Transformador beta/antagonistas & inibidoresRESUMO
OBJECTIVE: This study evaluated the angiogenesis-enhancing potential of a tricalcium silicate-based mineral trioxide aggregate (ProRoot MTA), Biodentine, and a novel bioceramic root canal sealer (Well-Root ST) in human dental pulp stem cells (hDPSCs), human periodontal ligament stem cells (hPLSCs), and human tooth germ stem cells (hTGSCs). METHODOLOGY: Dulbecco's modified Eagle's medium was conditioned for 24 h by exposure to ProRoot MTA, Biodentine, or Well-Root ST specimens (prepared according to the manufacturers' instructions). The cells were cultured in these conditioned media and their viability was assessed with 3-(4,5-dimethyl-thiazol-2-yl)-5-(3-carboxy-methoxy-phenyl)-2-(4-sulfo-phenyl)-2H tetrazolium (MTS) on days 1, 3, 7, 10, and 14. Angiogenic growth factors [platelet-derived growth factor (PDGF), basic ï¬broblast growth factor (FGF-2), and vascular endothelial growth factor (VEGF)] were assayed by sandwich enzyme-linked immunosorbent assay (ELISA) on days 1, 7, and 14. Human umbilical vein endothelial cell (HUVEC) migration assays were used to evaluate the vascular effects of the tested materials at 6-8 h. Statistical analyses included Kruskal-Wallis, Mann-Whitney U, and Friedman and Wilcoxon signed rank tests. RESULTS: None of tricalcium silicate-based materials were cytotoxic and all induced a similar release of angiogenic growth factors (PDGF, FGF-2, and VEGF) (p>0.05). The best cell viability was observed for hDPSCs (p<0.05) with all tricalcium silicate-based materials at day 14. Tube formation by HUVECs showed a significant increase with all tested materials (p<0.05). CONCLUSION: The tricalcium silicate-based materials showed potential for angiogenic stimulation of all stem cell types and significantly enhanced tube formation by HUVECs.
Assuntos
Indutores da Angiogênese/farmacologia , Compostos de Cálcio/farmacologia , Cerâmica/farmacologia , Materiais Restauradores do Canal Radicular/farmacologia , Silicatos/farmacologia , Células-Tronco/efeitos dos fármacos , Materiais Biocompatíveis/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Polpa Dentária/citologia , Polpa Dentária/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Fator 2 de Crescimento de Fibroblastos/análise , Fator 2 de Crescimento de Fibroblastos/efeitos dos fármacos , Citometria de Fluxo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Teste de Materiais , Neovascularização Fisiológica/efeitos dos fármacos , Ligamento Periodontal/citologia , Ligamento Periodontal/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/análise , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Reprodutibilidade dos Testes , Estatísticas não Paramétricas , Germe de Dente/citologia , Germe de Dente/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/análise , Fator A de Crescimento do Endotélio Vascular/efeitos dos fármacosRESUMO
Abstract Objective: This study evaluated the angiogenesis-enhancing potential of a tricalcium silicate-based mineral trioxide aggregate (ProRoot MTA), Biodentine, and a novel bioceramic root canal sealer (Well-Root ST) in human dental pulp stem cells (hDPSCs), human periodontal ligament stem cells (hPLSCs), and human tooth germ stem cells (hTGSCs). Methodology: Dulbecco's modified Eagle's medium was conditioned for 24 h by exposure to ProRoot MTA, Biodentine, or Well-Root ST specimens (prepared according to the manufacturers' instructions). The cells were cultured in these conditioned media and their viability was assessed with 3-(4,5-dimethyl-thiazol-2-yl)-5-(3-carboxy-methoxy-phenyl)-2-(4-sulfo-phenyl)-2H tetrazolium (MTS) on days 1, 3, 7, 10, and 14. Angiogenic growth factors [platelet-derived growth factor (PDGF), basic fibroblast growth factor (FGF-2), and vascular endothelial growth factor (VEGF)] were assayed by sandwich enzyme-linked immunosorbent assay (ELISA) on days 1, 7, and 14. Human umbilical vein endothelial cell (HUVEC) migration assays were used to evaluate the vascular effects of the tested materials at 6-8 h. Statistical analyses included Kruskal-Wallis, Mann-Whitney U, and Friedman and Wilcoxon signed rank tests. Results: None of tricalcium silicate-based materials were cytotoxic and all induced a similar release of angiogenic growth factors (PDGF, FGF-2, and VEGF) (p>0.05). The best cell viability was observed for hDPSCs (p<0.05) with all tricalcium silicate-based materials at day 14. Tube formation by HUVECs showed a significant increase with all tested materials (p<0.05). Conclusion: The tricalcium silicate-based materials showed potential for angiogenic stimulation of all stem cell types and significantly enhanced tube formation by HUVECs.
Assuntos
Humanos , Materiais Restauradores do Canal Radicular/farmacologia , Células-Tronco/efeitos dos fármacos , Cerâmica/farmacologia , Silicatos/farmacologia , Compostos de Cálcio/farmacologia , Indutores da Angiogênese/farmacologia , Ligamento Periodontal/citologia , Ligamento Periodontal/efeitos dos fármacos , Germe de Dente/citologia , Germe de Dente/efeitos dos fármacos , Materiais Biocompatíveis/farmacologia , Teste de Materiais , Fator de Crescimento Derivado de Plaquetas/análise , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Sobrevivência Celular/efeitos dos fármacos , Reprodutibilidade dos Testes , Fator 2 de Crescimento de Fibroblastos/análise , Fator 2 de Crescimento de Fibroblastos/efeitos dos fármacos , Estatísticas não Paramétricas , Neovascularização Fisiológica/efeitos dos fármacos , Polpa Dentária/citologia , Polpa Dentária/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/análise , Fator A de Crescimento do Endotélio Vascular/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Citometria de FluxoRESUMO
Nintedanib is an oral, small-molecule tyrosine kinase inhibitor approved for the treatment of idiopathic pulmonary fibrosis and patients with advanced non-small cell cancer of adenocarcinoma tumour histology. Nintedanib competitively binds to the kinase domains of vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF) and fibroblast growth factor (FGF). Studies in healthy volunteers and in patients with advanced cancer have shown that nintedanib has time-independent pharmacokinetic characteristics. Maximum plasma concentrations of nintedanib are reached approximately 2-4 h after oral administration and thereafter decline at least bi-exponentially. Over the investigated dose range of 50-450 mg once daily and 150-300 mg twice daily, nintedanib exposure increases are dose proportional. Nintedanib is metabolised via hydrolytic ester cleavage, resulting in the formation of the free acid moiety that is subsequently glucuronidated and excreted in the faeces. Less than 1% of drug-related radioactivity is eliminated in urine. The terminal elimination half-life of nintedanib is about 10-15 h. Accumulation after repeated twice-daily dosing is negligible. Sex and renal function have no influence on nintedanib pharmacokinetics, while effects of ethnicity, low body weight, older age and smoking are within the inter-patient variability range of nintedanib exposure and no dose adjustments are required. Administration of nintedanib in patients with moderate or severe hepatic impairment is not recommended, and patients with mild hepatic impairment should be monitored closely and the dose adjusted accordingly. Nintedanib has a low potential for drug-drug interactions, especially with drugs metabolised by cytochrome P450 enzymes. Concomitant treatment with potent inhibitors or inducers of the P-glycoprotein transporter can affect the pharmacokinetics of nintedanib. At an investigated dose of 200 mg twice daily, nintedanib does not have proarrhythmic potential.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/patologia , Fibrose Pulmonar Idiopática/tratamento farmacológico , Indóis/farmacocinética , Inibidores de Proteínas Quinases/farmacocinética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Administração Oral , Idoso , Animais , Carcinoma Pulmonar de Células não Pequenas/complicações , Estudos de Casos e Controles , Sistema Enzimático do Citocromo P-450/metabolismo , Interações Medicamentosas , Feminino , Fatores de Crescimento de Fibroblastos/efeitos dos fármacos , Humanos , Indóis/administração & dosagem , Indóis/sangue , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Modelos Animais , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/sangue , Ratos , Fator A de Crescimento do Endotélio Vascular/efeitos dos fármacosRESUMO
BACKGROUND The aim of this study was to evaluate the effects of 3-tetrazolyl methyl-3-hydroxy-oxindole hybrid (THOH) on cell proliferation, apoptosis, and the cell cycle in human lung cancer cell lines SK-LU-1, A549, and A-427, and the normal lung fibroblast cell line, MRC-5, in vitro. MATERIAL AND METHODS Human lung adenocarcinoma cells SK-LU-1, A549, and A-427, and the normal lung fibroblast cells, MRC-5 were cultured and treated with increasing concentrations of 10 mM of a stock solution of THOH in dimethyl sulfoxide (DMSO). An MTT cell proliferation assay was used. Cell apoptosis and the cell cycle were studied using fluorescence-activated cell sorting (FACs) with fluorescein isothiocyanate (FITC), Annexin-V, propidium iodide (PI), and nuclear staining with 4',6-diamidino-2-phenylindole (DAPI). DNA damage was measured using the comet (single-cell gel electrophoresis) assay. Cell migration was evaluated using a wound healing assay, and Western blotting was used to measure protein expression levels. RESULTS Treatment of SK-LU-1 cells with THOH inhibited cell migration. Treatment of lung cancer cells, SK-LU-1, A549, and A-427, with THOH inhibited cell proliferation, with the most marked inhibition found in the SK-LU-1 lung cancer cells (IC50, 12 µM). Treatment of lung cancer cells, SK-LU-1, A549, and A-427, with THOH increased cell apoptosis, resulted in G2/M cell cycle arrest, and inhibited both the platelet-derived growth factor D (PDGF-D) and MEK/ERK signaling pathways. CONCLUSIONS Treatment of adenocarcinoma cells, SK-LU-1, A549, and A-427, with THOH inhibited cell proliferation, apoptosis, and resulted in G2/M cell cycle arrest by targeting PDGF-D and the MEK/ERK signaling pathway.
Assuntos
Indóis/farmacologia , Linfocinas/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Tetrazóis/farmacologia , Células A549/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Humanos , Indóis/metabolismo , Neoplasias Pulmonares/patologia , Oxindóis , Transdução de Sinais , Tetrazóis/metabolismoRESUMO
BACKGROUND: A serious adverse effect of corticosteroid therapy is adrenal suppression. Our aim was to identify genetic variants affecting susceptibility to corticosteroid-induced adrenal suppression. METHODS: We enrolled children with asthma who used inhaled corticosteroids as part of their treatment from 25 sites across the UK (discovery cohort), as part of the Pharmacogenetics of Adrenal Suppression with Inhaled Steroids (PASS) study. We included two validation cohorts, one comprising children with asthma (PASS study) and the other consisting of adults with chronic obstructive pulmonary disorder (COPD) who were recruited from two UK centres for the Pharmacogenomics of Adrenal Suppression in COPD (PASIC) study. Participants underwent a low-dose short synacthen test. Adrenal suppression was defined as peak cortisol less than 350 nmol/L (in children) and less than 500 nmol/L (in adults). A case-control genome-wide association study was done with the control subset augmented by Wellcome Trust Case Control Consortium 2 (WTCCC2) participants. Single nucleotide polymorphisms (SNPs) that fulfilled criteria to be advanced to replication were tested by a random-effects inverse variance meta-analysis. This report presents the primary analysis. The PASS study is registered in the European Genome-phenome Archive (EGA). The PASS study is complete whereas the PASIC study is ongoing. FINDINGS: Between November, 2008, and September, 2011, 499 children were enrolled to the discovery cohort. Between October, 2011, and December, 2012, 81 children were enrolled to the paediatric validation cohort, and from February, 2010, to June, 2015, 78 adults were enrolled to the adult validation cohort. Adrenal suppression was present in 35 (7%) children in the discovery cohort and six (7%) children and 17 (22%) adults in the validation cohorts. In the discovery cohort, 40 SNPs were found to be associated with adrenal suppression (genome-wide significance p<1â×â10-6), including an intronic SNP within the PDGFD gene locus (rs591118; odds ratio [OR] 7·32, 95% CI 3·15-16·99; p=5·8â×â10-8). This finding for rs591118 was validated successfully in both the paediatric asthma (OR 3·86, 95% CI 1·19-12·50; p=0·02) and adult COPD (2·41, 1·10-5·28; p=0·03) cohorts. The proportions of patients with adrenal suppression by rs591118 genotype were six (3%) of 214 patients with the GG genotype, 15 (6%) of 244 with the AG genotype, and 22 (25%) of 87 with the AA genotype. Meta-analysis of the paediatric cohorts (discovery and validation) and all three cohorts showed genome-wide significance of rs591118 (respectively, OR 5·89, 95% CI 2·97-11·68; p=4·3â×â10-9; and 4·05, 2·00-8·21; p=3·5â×â10-10). INTERPRETATION: Our findings suggest that genetic variation in the PDGFD gene locus increases the risk of adrenal suppression in children and adults who use corticosteroids to treat asthma and COPD, respectively. FUNDING: Department of Health Chair in Pharmacogenetics.
Assuntos
Corticosteroides/efeitos adversos , Insuficiência Adrenal/genética , Antiasmáticos/efeitos adversos , Asma/tratamento farmacológico , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , Adolescente , Corticosteroides/administração & dosagem , Insuficiência Adrenal/induzido quimicamente , Adulto , Idoso , Antiasmáticos/administração & dosagem , Asma/genética , Estudos de Casos e Controles , Criança , Feminino , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Hidrocortisona/análise , Linfocinas/efeitos dos fármacos , Linfocinas/genética , Masculino , Pessoa de Meia-Idade , Variantes Farmacogenômicos , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/genética , Polimorfismo de Nucleotídeo Único , Doença Pulmonar Obstrutiva Crônica/genética , Adulto JovemRESUMO
Cholangiocyte senescence has been linked to primary sclerosing cholangitis (PSC). Persistent secretion of growth factors by senescent cholangiocytes leads to the activation of stromal fibroblasts (ASFs), which are drivers of fibrosis. The activated phenotype of ASFs is characterized by an increased sensitivity to apoptotic stimuli. Here, we examined the mechanisms of apoptotic priming in ASFs and explored a combined targeting strategy to deplete senescent cholangiocytes and ASFs from fibrotic tissue to ameliorate liver fibrosis. Using a coculture system, we determined that senescent cholangiocytes promoted quiescent mesenchymal cell activation in a platelet-derived growth factor (PDGF)-dependent manner. We also identified B-cell lymphoma-extra large (Bcl-xL) as a key survival factor in PDGF-activated human and mouse fibroblasts. Bcl-xL was also up-regulated in senescent cholangiocytes. In vitro, inhibition of Bcl-xL by the small molecule Bcl-2 homology domain 3 mimetic, A-1331852, or Bcl-xL-specific small interfering RNA induced apoptosis in PDGF-activated fibroblasts, but not in quiescent fibroblasts. Likewise, inhibition of Bcl-xL reduced the survival and increased apoptosis of senescent cholangiocytes, compared to nonsenescent cells. Treatment of multidrug resistance 2 gene knockout (Mdr2-/- ) mice with A-1331852 resulted in an 80% decrease in senescent cholangiocytes, a reduction of fibrosis-inducing growth factors and cytokines, decrease of α-smooth muscle actin-positive ASFs, and finally in a significant reduction of liver fibrosis. CONCLUSION: Bcl-xL is a key survival factor in ASFs as well as in senescent cholangiocytes. Treatment with the Bcl-xL-specific inhibitor, A-1331852, reduces liver fibrosis, possibly by a dual effect on activated fibroblasts and senescent cholangiocytes. This mechanism represents an attractive therapeutic strategy in biliary fibrosis. (Hepatology 2018;67:247-259).
Assuntos
Benzotiazóis/farmacologia , Ductos Biliares/citologia , Colangite Esclerosante/patologia , Fibroblastos/efeitos dos fármacos , Isoquinolinas/farmacologia , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Animais , Biópsia por Agulha , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Colangite Esclerosante/tratamento farmacológico , Modelos Animais de Doenças , Resistência a Múltiplos Medicamentos , Fibroblastos/metabolismo , Fibroblastos/patologia , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Terapia de Alvo Molecular , Fator de Crescimento Derivado de Plaquetas/metabolismo , Distribuição Aleatória , Valores de ReferênciaRESUMO
OBJECTIVE: To elucidate the role of gene candidates involved in pulmonary hypertension (PH) associated with systemic sclerosis (SSc). METHODS: Gene candidates were identified through microarray experiments performed on Affymetrix GeneChip Human Exon 1.0 ST arrays in endothelial progenitor cell (EPC)-derived endothelial cells (ECs) obtained from patients with SSc-associated PH, patients with SSc without PH, and healthy control subjects. Expression of identified gene candidates was assessed by quantitative sandwich enzyme-linked immunosorbent assay in the serum, and by immunohistochemistry in lesional lung tissue. The functional importance of the identified gene candidates was then evaluated in fos-related antigen 2-transgenic (Fra-2-Tg) mice that spontaneously develop SSc-like features associated with an intense pulmonary vascular remodeling. RESULTS: Microarray experiments revealed that the matrix metalloproteinase 10 (MMP-10) gene was the top up-regulated gene in SSc-associated PH EPC-derived ECs. Circulating serum proMMP10 concentrations were markedly increased in patients with SSc-associated PH compared to SSc patients without PH and healthy controls. Consistent with these observations, a strong MMP10 staining of the thickened wall of distal pulmonary arteries was found both in the lungs of patients with SSc-associated PH and in the lungs of Fra-2-Tg mice. Daily treatment of Fra-2-Tg mice with neutralizing anti-MMP10 antibodies did not significantly affect the development and severity of pulmonary fibrosis, but did reverse established PH and markedly reduced pulmonary vascular remodeling by reducing cell proliferation, cell survival, and the platelet-derived growth factor signaling axis. CONCLUSION: Gene expression profiling of EPC-derived ECs identified MMP10 as a novel candidate gene in SSc-associated PH. MMP10 is overexpressed in the serum and pulmonary arteries of patients with SSc-associated PH, and its blockade alleviates PH in the Fra-2-Tg mouse model. MMP10 appears to be a prospective treatment target for this devastating disorder.
Assuntos
Células Endoteliais/metabolismo , Hipertensão Pulmonar/genética , Metaloproteinase 10 da Matriz/genética , Artéria Pulmonar/metabolismo , Fibrose Pulmonar/genética , Escleroderma Sistêmico/genética , Adulto , Idoso , Animais , Anticorpos Neutralizantes/farmacologia , Estudos de Casos e Controles , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular , Modelos Animais de Doenças , Células Progenitoras Endoteliais , Ensaio de Imunoadsorção Enzimática , Feminino , Antígeno 2 Relacionado a Fos/genética , Humanos , Hipertensão Pulmonar/etiologia , Hipertensão Pulmonar/metabolismo , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Masculino , Metaloproteinase 10 da Matriz/imunologia , Metaloproteinase 10 da Matriz/metabolismo , Camundongos , Camundongos Transgênicos , Análise em Microsséries , Pessoa de Meia-Idade , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/metabolismo , Fibrose Pulmonar/etiologia , Fibrose Pulmonar/metabolismo , Escleroderma Sistêmico/complicações , Escleroderma Sistêmico/metabolismo , Transdução de Sinais , Remodelação Vascular/efeitos dos fármacosRESUMO
Xanthohumol (1) is a principal prenylated chalcone found in hops. The aim of this study was to examine its influence on platelet-derived growth factor (PDGF)-BB-triggered vascular smooth muscle cell (VSMC) proliferation and migration in vitro and on experimentally induced neointima formation in vivo. Quantification of resazurin conversion indicated that 1 can inhibit PDGF-BB-induced VSMC proliferation concentration-dependently (IC50 = 3.49 µM). Furthermore, in a wound-healing assay 1 potently suppresses PDGF-BB-induced VSMC migration at 15 µM. Tested in a mouse femoral artery cuff model, 1 significantly reduces neointima formation. Taken together, we show that 1 represses PDGF-BB-induced VSMC proliferation and migration in vitro as well as neointima formation in vivo. This novel activity suggests 1 as an interesting candidate for further studies addressing a possible therapeutic application to counteract vascular proliferative disease.
Assuntos
Flavonoides/farmacologia , Humulus/química , Neointima/metabolismo , Propiofenonas/farmacologia , Animais , Becaplermina , Movimento Celular , Proliferação de Células , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Flavonoides/química , Flavonoides/isolamento & purificação , Sistema de Sinalização das MAP Quinases , Camundongos , Estrutura Molecular , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Neointima/induzido quimicamente , Oxazinas/metabolismo , Fosforilação , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Propiofenonas/química , Propiofenonas/isolamento & purificação , Proteínas Proto-Oncogênicas c-sis , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Cicatrização/efeitos dos fármacos , Xantenos/metabolismoRESUMO
BACKGROUND: Hydrogen sulfide (H2S) is a gasotransmitter that regulates multiple cardiovascular functions. Krüppel-like factor 5 (KLF5) exerts diverse functions in the cardiovascular system. Whether and how H2S regulates KLF5 in myocardial hypertrophy is unknown. METHODS AND RESULTS: In our study, hypertrophic myocardial samples in the clinic were collected and underwent histological and molecular biological analysis. Spontaneously hypertensive rats and neonatal rat cardiomyocytes were studied for functional and signaling responses to GYY4137, an H2S-releasing compound. Expression of cystathionine γ-lyase, a principal enzyme for H2S generation in heart, decreased in human hypertrophic myocardium, whereas KLF5 expression increased. After GYY4137 administration for 4 weeks, myocardial hypertrophy was inhibited in spontaneously hypertensive rats, as demonstrated by improvement in cardiac structural parameters, heart mass, size of cardiac myocytes, and expression of atrial natriuretic peptide. H2S diminished expression of KLF5 in myocardium of spontaneously hypertensive rats and in hypertrophic cardiomyocytes. H2S also inhibits platelet-derived growth factor A promoter activity, decreased recruitment of KLF5 to the platelet-derived growth factor A promoter, and reduced atrial natriuretic peptide expression in angiotensin II-stimulated cardiomyocytes, and these effects are suppressed by KLF5 knockdown. KLF5 promoter activity and KLF5 expression was also reversed by H2S. H2S increased the S-sulfhydration on specificity protein 1 in cardiomyocytes. Moreover, H2S decreased KLF5 promoter activity; reduced KLF5 mRNA expression; attenuated specificity protein 1 binding activity with KLF5 promoter; and inhibited hypertrophy after specificity protein 1 mutated at Cys659, Cys689, and Cys692 but not Cys664 overexpression. CONCLUSIONS: These findings suggest that H2S regulates KLF5 transcription activity via specificity protein 1 S-sulfhydration at Cys664 to prevent myocardial hypertrophy.
Assuntos
Cardiomegalia/genética , Cistationina gama-Liase/metabolismo , Sulfeto de Hidrogênio/metabolismo , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Miocárdio/metabolismo , Miócitos Cardíacos/metabolismo , RNA Mensageiro/metabolismo , Angiotensina II/sangue , Animais , Animais Recém-Nascidos , Fator Natriurético Atrial/efeitos dos fármacos , Fator Natriurético Atrial/metabolismo , Cardiomegalia/metabolismo , Estudos de Casos e Controles , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Fatores de Transcrição Kruppel-Like/efeitos dos fármacos , Masculino , Morfolinas/farmacologia , Miócitos Cardíacos/efeitos dos fármacos , Compostos Organotiofosforados/farmacologia , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/metabolismo , RNA Mensageiro/efeitos dos fármacos , Ratos , Ratos Endogâmicos SHR , Ratos Sprague-Dawley , Fator de Transcrição Sp1/metabolismo , Ativação Transcricional/efeitos dos fármacosRESUMO
Abnormal high mobility group protein B1 (HMGB1) activation is involved in the pathogenesis of pulmonary fibrosis. Pulmonary rehabilitation mixture (PRM), which combines extracts from eight traditional Chinese medicines, has very good lung protection in clinical use. However, it is not known if PRM has anti-fibrotic activity. In this study, we investigated the effects of PRM on transforming growth factor-β1 (TGF-β1)-mediated and bleomycin (BLM)-induced pulmonary fibrosis in vitro and in vivo. The effects of PRM on TGF-β1-mediated epithelial-mesenchymal transition (EMT) in A549 cells, on the proliferation of human lung fibroblasts (HLF-1) in vitro, and on BLM-induced pulmonary fibrosis in vivo were investigated. PRM treatment resulted in a reduction of EMT in A549 cells that was associated with attenuating an increase of vimentin and a decrease of E-cadherin. PRM inhibited the proliferation of HLF-1 at an IC50 of 0.51 µg/mL. PRM ameliorated BLM-induced pulmonary fibrosis in rats, with reduction of histopathological scores and collagen deposition, and a decrease in α-smooth muscle actin (α-SMA) and HMGB1 expression. An increase in receptor for advanced glycation end-product (RAGE) expression was found in BLM-instilled lungs. PRM significantly decreased EMT and prevented pulmonary fibrosis through decreasing HMGB1 and regulating RAGE in vitro and in vivo. PRM inhibited TGF-β1-induced EMT via decreased HMGB1 and vimentin and increased RAGE and E-cadherin levels. In summary, PRM prevented experimental pulmonary fibrosis by modulating the HMGB1/RAGE pathway.
Assuntos
Animais , Humanos , Masculino , Medicamentos de Ervas Chinesas/farmacologia , Fibrose Pulmonar/tratamento farmacológico , Fibrose Pulmonar/prevenção & controle , Antibióticos Antineoplásicos , Receptor para Produtos Finais de Glicação Avançada/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Bleomicina , Western Blotting , Células Cultivadas , Colágeno/efeitos dos fármacos , Misturas Complexas/farmacologia , Medicamentos de Ervas Chinesas/uso terapêutico , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Proteína HMGB1/efeitos dos fármacos , Hidroxiprolina/análise , Imuno-Histoquímica , Pulmão/efeitos dos fármacos , Pulmão/patologia , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Fibrose Pulmonar/patologia , Distribuição Aleatória , Ratos Sprague-Dawley , Reprodutibilidade dos Testes , Fator de Crescimento Transformador beta1/efeitos dos fármacosRESUMO
Abnormal high mobility group protein B1 (HMGB1) activation is involved in the pathogenesis of pulmonary fibrosis. Pulmonary rehabilitation mixture (PRM), which combines extracts from eight traditional Chinese medicines, has very good lung protection in clinical use. However, it is not known if PRM has anti-fibrotic activity. In this study, we investigated the effects of PRM on transforming growth factor-ß1 (TGF-ß1)-mediated and bleomycin (BLM)-induced pulmonary fibrosis in vitro and in vivo. The effects of PRM on TGF-ß1-mediated epithelial-mesenchymal transition (EMT) in A549 cells, on the proliferation of human lung fibroblasts (HLF-1) in vitro, and on BLM-induced pulmonary fibrosis in vivo were investigated. PRM treatment resulted in a reduction of EMT in A549 cells that was associated with attenuating an increase of vimentin and a decrease of E-cadherin. PRM inhibited the proliferation of HLF-1 at an IC50 of 0.51 µg/mL. PRM ameliorated BLM-induced pulmonary fibrosis in rats, with reduction of histopathological scores and collagen deposition, and a decrease in α-smooth muscle actin (α-SMA) and HMGB1 expression. An increase in receptor for advanced glycation end-product (RAGE) expression was found in BLM-instilled lungs. PRM significantly decreased EMT and prevented pulmonary fibrosis through decreasing HMGB1 and regulating RAGE in vitro and in vivo. PRM inhibited TGF-ß1-induced EMT via decreased HMGB1 and vimentin and increased RAGE and E-cadherin levels. In summary, PRM prevented experimental pulmonary fibrosis by modulating the HMGB1/RAGE pathway.
Assuntos
Medicamentos de Ervas Chinesas/farmacologia , Fibrose Pulmonar/tratamento farmacológico , Fibrose Pulmonar/prevenção & controle , Animais , Antibióticos Antineoplásicos , Apoptose/efeitos dos fármacos , Bleomicina , Western Blotting , Células Cultivadas , Colágeno/efeitos dos fármacos , Misturas Complexas/farmacologia , Medicamentos de Ervas Chinesas/uso terapêutico , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Proteína HMGB1/efeitos dos fármacos , Humanos , Hidroxiprolina/análise , Imuno-Histoquímica , Pulmão/efeitos dos fármacos , Pulmão/patologia , Masculino , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Fibrose Pulmonar/patologia , Distribuição Aleatória , Ratos Sprague-Dawley , Receptor para Produtos Finais de Glicação Avançada/efeitos dos fármacos , Reprodutibilidade dos Testes , Fator de Crescimento Transformador beta1/efeitos dos fármacosRESUMO
AIM: Eukaryotic elongation factor 2 kinase (eEF2K), also known as calmodulin (CaM)-dependent protein kinase (CaMK) III, is a unique member of CaMK family protein. We have recently found that expression of eEF2K protein increased in mesenteric artery from spontaneously hypertensive rats. As pathogenesis of hypertension is in part regulated by vascular structural remodelling via proliferation and migration of vascular smooth muscle cells (SMCs), we tested the hypothesis that eEF2K controls SMCs proliferation and migration. METHODSAND RESULTS: In rat mesenteric arterial SMCs, an eEF2K inhibitor, A-484954 (10 µm), significantly inhibited platelet-derived growth factor (PDGF)-BB (10 ng mL(-1) )-induced SMCs proliferation as determined by a cell counting and bromodeoxyuridine incorporation assay. PDGF-BB (10 ng mL(-1) )-induced SMCs migration was significantly inhibited by A-484954 (10 µm) as determined by a Boyden chamber assay. A-484954 (10 µm) significantly inhibited PDGF-BB (10 ng mL(-1) )-induced phosphorylation of eEF2K, extracellular signal-regulated kinase (ERK), Akt, p38 and heat-shock protein (HSP) 27 as determined by Western blotting. It was confirmed that a CaM inhibitor, W-7 (50 µm), inhibited PDGF-BB (10 ng mL(-1) )-induced phosphorylation of eEF2K. In an ex vivo mesenteric arterial ring assay, 10% foetal bovine serum-induced SMCs outgrowth was significantly inhibited by A-484954 (10 µm). CONCLUSION: We for the first time revealed that eEF2K mediates PDGF-BB-induced SMCs proliferation and migration through activating ERK, Akt, p38 and HSP27 signals in a CaM-dependent manner. Our results suggest eEF2K as a novel pharmaceutical target for the prevention of hypertensive cardiovascular diseases.
Assuntos
Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Quinase do Fator 2 de Elongação/metabolismo , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Animais , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Células Cultivadas , Quinase do Fator 2 de Elongação/antagonistas & inibidores , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas de Choque Térmico HSP27/metabolismo , Proteínas de Choque Térmico , Hipertensão/tratamento farmacológico , Masculino , Proteínas Quinases Ativadas por Mitógeno/efeitos dos fármacos , Chaperonas Moleculares , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/metabolismo , Ratos Wistar , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
BACKGROUND: The pathogenesis of human chronic rhinosinusitis with nasal polyps (CRSwNP) comprising eosinophilic CRSwNP (ECRSwNP) and non-eosinophilic (nECRSwNP) is not completely understood. Recent evidence has suggested that platelet-derived growth factor receptor alpha (PDGFRα) is implicated in cell growth, transformation, proliferation, migration, and vascular permeability and platelet-derived growth factor-A (PDGF-A) is a specific ligand for PDGFRα. However, little is known about their roles in CRSwNP. Therefore, we aimed to investigate the expression and role of PDGFRα and PDGF-A in CRSwNP. METHODS: PDGFRα protein expression was investigated by immunohistochemistry method and messenger RNA (mRNA) expression of PDGFRα and PDGF-A were assessed by real-time polymerase chain reaction (PCR) in CRSwNP patients and control subjects. Moreover, the effects of various stimulators with different concentrations and time on PDGFRα were evaluated on nasal explant culture. RESULTS: Quantitative analysis of immunostaining for PDGFRα showed an obvious elevation in immunolabeling of PDGFRα in CRSwNP groups compared with controls. Furthermore, PDGFRα protein was significantly stronger expressed in ECRSwNP group than nECRSwNP group and atopic patients showed stronger expression of PDGFRα protein than nonatopic patients. The mRNA of PDGFRα and PDGF-A were overexpressed in CRSwNP, especially in ECRSwNP. PDGFRα mRNA expression was closely related to PDGF-A mRNA. In nasal explant culture and stimulation, PDGFRα mRNA was augmented by interleukin 4 (IL-4), IL-5, or IL-1ß respectively, but suppressed by IL-27. CONCLUSION: PDGFRα may play a pivotal role in the pathophysiology of ECRSwNP and nECRSwNP by combining with PDGF-A. IL-4, IL-5, or IL-1ß may be critical for PDGFRα gene expression.
Assuntos
Eosinofilia/etiologia , Pólipos Nasais/etiologia , Fator de Crescimento Derivado de Plaquetas/fisiologia , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/fisiologia , Rinite/etiologia , Sinusite/etiologia , Adulto , Doença Crônica , Feminino , Humanos , Imuno-Histoquímica , Interleucinas/farmacologia , Masculino , Seios Paranasais/metabolismo , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/metabolismo , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
PRP cryopreservation remains a controversial point. Our purpose was to investigate the effect of freezing/thawing on PRP molecule release, and its effects on the metabolism of chondrocytes and synoviocytes. PRP was prepared from 10 volunteers, and a half volume underwent one freezing/thawing cycle. IL-1ß, HGF, PDGF AB/BB, TGF-ß1, and VEGF were assayed 1 hour and 7 days after activation. Culture media of chondrocytes and synoviocytes were supplemented with fresh or frozen PRP, and, at 7 days, proliferation, gene expression, and secreted proteins levels were evaluated. Results showed that in the freeze-thawed PRP the immediate and delayed molecule releases were similar or slightly lower than those in fresh PRP. TGF-ß1 and PDGF AB/BB concentrations were significantly reduced after freezing both at 1 hour and at 7 days, whereas HGF concentration was significantly lower in frozen PRP at 7 days. In fresh PRP IL-1ß and HGF concentrations underwent a significant further increase after 7 days. Similar gene expression was found in chondrocytes cultured with both PRPs, whereas in synoviocytes HGF gene expression was higher in frozen PRP. PRP cryopreservation is a safe procedure, which sufficiently preserves PRP quality and its ability to induce proliferation and the production of ECM components in chondrocytes and synoviocytes.
Assuntos
Condrócitos/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/metabolismo , Plasma Rico em Plaquetas/química , Líquido Sinovial/efeitos dos fármacos , Adulto , Proliferação de Células/efeitos dos fármacos , Criopreservação , Meios de Cultura , Congelamento , Humanos , Masculino , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-sis/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-sis/metabolismo , Líquido Sinovial/citologiaRESUMO
BACKGROUND: Acute respiratory distress syndrome (ARDS) is characterized by overwhelming inflammatory responses and lung remodeling. We hypothesized that leukocyte infiltration during the inflammatory response modulates epithelial remodeling through a mechanism of epithelial-mesenchymal transition (EMT). METHODS: Human lung epithelial cells were treated for 30 min with hydrochloric acid (HCl). Human monocytes were then cocultured with the epithelial cells for up to 48 h, in the presence or absence of blocking peptides against lymphocyte function-associated antigen-1 (LFA-1), or tyrphostin A9, a specific inhibitor for platelet-derived growth factor (PDGF) receptor tyrosine kinase. RESULTS: Exposure of lung epithelial cells to HCl resulted in increased expression of intercellular adhesion molecule-1 (ICAM-1) and production of interleukin (IL)-8 at 24 h. The expression of the epithelial markers E-cadherin decreased while the mesenchymal markers vimentin and α-smooth muscle actin (α-SMA) increased at 24 h and remained high at 48 h. The addition of monocytes augmented the profiles of lower expression of epithelial markers and higher mesenchymal markers accompanied by increased collagen deposition. This EMT profile was associated with an enhanced production of IL-8 and PDGF. Treatment of the lung epithelial cells with the LAF-1 blocking peptides CD11a237-246 or/and CD18112-122 suppressed monocyte adhesion, production of IL-8, PDGF and hydroxyproline as well as EMT markers. Treatment with tyrphostin A9 prevented the EMT profile shift induced by HCl stimulation. CONCLUSIONS: The interaction between epithelial cells and monocytes enhanced epithelial remodelling after initial injury through EMT signalling that is associated with the release of soluble mediators, including IL-8 and PDGF.
Assuntos
Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/patologia , Transição Epitelial-Mesenquimal , Monócitos , Actinas/metabolismo , Células Epiteliais Alveolares/fisiologia , Antígeno CD11a , Antígenos CD18 , Caderinas/metabolismo , Adesão Celular/efeitos dos fármacos , Células Cultivadas , Técnicas de Cocultura , Humanos , Ácido Clorídrico/farmacologia , Molécula 1 de Adesão Intercelular/metabolismo , Interleucina-8/efeitos dos fármacos , Interleucina-8/metabolismo , Antígeno-1 Associado à Função Linfocitária/efeitos dos fármacos , Peptídeos/farmacologia , Fator de Crescimento Derivado de Plaquetas/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/metabolismo , Tirfostinas/farmacologia , Vimentina/metabolismoRESUMO
INTRODUCTION: Prostacyclin (PGI2) is a biomolecule capable of enhancing angiogenesis and cellular proliferation. METHODS: We investigated the influence of a PGI2 analogue (iloprost) on dental pulp revascularization in vitro and in vivo by using human dental pulp cells (HDPCs) and a rat tooth injury model, respectively. Iloprost stimulated the human dental pulp cell mRNA expression of vascular endothelial growth factor (VEGF), fibroblast growth factor-2 (FGF-2), and platelet-derived growth factor (PDGF) in a significant dose-dependent manner. This mRNA up-regulation was significantly inhibited by pretreatment with a PGI2 receptor antagonist and forskolin (a protein kinase A activator). In contrast, a protein kinase A inhibitor significantly enhanced the iloprost-induced mRNA expression of VEGF, FGF-2, and PDGF. Pretreatment with a fibroblast growth factor receptor inhibitor attenuated the VEGF, FGF-2, and PDGF mRNA expression, indicating opposing regulatory mechanisms. RESULTS: The effect of iloprost on the dental pulp was investigated in vivo by using a rat molar pulp injury model. The iloprost-treated group exhibited a significant increase in pulpal blood flow at 72 hours compared with control. CONCLUSIONS: The present study indicates that iloprost may be a candidate agent to promote neovascularization in dental pulp tissue, suggesting the potential clinical use of iloprost in vital pulp therapy.