RESUMO
AIM: The study was to clarify the mechanism of miR-1258 targeting Prep1 (pKnox1) to control Transforming Growth Factor ß1 (TGF-ß1)/SMAD3 pathway in septic Acute Lung Injury (ALI)-induced oxidative stress and inflammation. METHODS: BEAS-2B cells and C57BL/6 mice were used to make in vitro and in vivo septic ALI models, respectively. miR-1258 expression was checked by RT-qPCR. After transfection in the in vitro experimental model, inflammation, oxidative stress, viability, and apoptosis were observed through ELISA, MTT, and flow cytometry. RESULTS: In the in vivo model after miR-1258 overexpression treatment, inflammation, oxidative stress, and lung injury were further investigated. The targeting relationship between miR-1258 and Pknox1 was tested. Low miR-1258 was expressed in septic ALI patients, LPS-treated BEAS-2B cells, and mice. Upregulated miR-1258 prevented inflammation, oxidative stress, and apoptosis but enhanced the viability of LPS-treated BEAS-2B cells. The impact of upregulated miR-1258 on LPS-treated BEAS-2B cells was mitigated by inhibiting Pknox1 expression. MiR-1258 overexpression had the alleviating effects on inflammation, oxidative stress, and lung injury of LPS-injured mice through suppressing Pknox1 expression and TGF-ß1/SMAD3 cascade activation. CONCLUSIONS: The study concludes that miR-1258 suppresses oxidative stress and inflammation in septic ALI through the Pknox1-regulated TGF-ß1/SMAD3 cascade.
Assuntos
Lesão Pulmonar Aguda , Apoptose , Camundongos Endogâmicos C57BL , MicroRNAs , Estresse Oxidativo , Sepse , Proteína Smad3 , Fator de Crescimento Transformador beta1 , Animais , Humanos , Masculino , Camundongos , Lesão Pulmonar Aguda/genética , Lesão Pulmonar Aguda/metabolismo , Modelos Animais de Doenças , Inflamação/metabolismo , MicroRNAs/metabolismo , MicroRNAs/genética , Sepse/complicações , Sepse/metabolismo , Sepse/genética , Transdução de Sinais , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Regulação para CimaRESUMO
Maternal malnutrition can alter developmental biology, programming health and disease in offspring. The increase in sugar consumption during the peripubertal period, a worldwide concern, also affects health through adulthood. Studies have shown that maternal exposure to a low protein diet (LPD) is associated with an increase in prostate disease with aging. However, the combined effects of maternal LPD and early postnatal sugar consumption on offspring prostate disorders were not investigated. The effects on aging were evaluated using a maternal gestational model with lactational LPD (6% protein) and sugar consumption (10%) from postnatal day (PND) 21-90, associating the consequences on ventral prostate (VP) rats morphophysiology on PND540. An increase was shown in mast cells and in the VP of the CTR + SUG and Gestational and Lactational Low Protein (GLLP) groups. In GLLP + SUG, a significant increase was shown in TGF-ß1 expression in both the systemic and intra-prostatic forms, and SMAD2/3p had increased. The study identified maternal LPD and sugar consumption as risk factors for prostatic homeostasis in senility, activating the TGFß1-SMAD2/3 pathway, a signaling pathway with potential markers for prostatic disorders.
Assuntos
Desnutrição , Fenômenos Fisiológicos da Nutrição Materna , Efeitos Tardios da Exposição Pré-Natal , Próstata , Doenças Prostáticas , Animais , Masculino , Feminino , Gravidez , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Doenças Prostáticas/patologia , Doenças Prostáticas/etiologia , Doenças Prostáticas/metabolismo , Desnutrição/complicações , Próstata/metabolismo , Próstata/patologia , Ratos , Inflamação/patologia , Inflamação/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta1/genética , Dieta com Restrição de Proteínas/efeitos adversos , Proteína Smad2/metabolismo , Ratos Wistar , Proteína Smad3/metabolismo , Proteína Smad3/genética , Transdução de Sinais , Animais Recém-Nascidos , Mastócitos/metabolismoRESUMO
INTRODUCTION AND OBJECTIVES: Caffeine consumption is associated with beneficial effects on hepatic disorders. The objectives of this study were to evaluate the antifibrotic effects of caffeine on experimental nonalcoholic steatohepatitis (NASH) induced with a high-fat, high-sucrose, high-cholesterol diet (HFSCD), as well as to evaluate the ability of caffeine to prevent the progression of experimental liver fibrosis induced by the administration of thioacetamide (TAA) in rats and explore the mechanisms of action. METHODS: NASH and fibrosis were induced in rats by the administration of an HFSCD for 15 weeks, and liver fibrosis was induced by intraperitoneal administration of 200 mg/kg TAA 3 times per week, for 6 weeks. Caffeine was administered at a dose of 50 mg/kg body weight. The effects of diet, TAA, and caffeine on fibrosis were evaluated by biochemical and histological examinations. The profibrotic pathways were analyzed by western blotting and immunohistochemistry. RESULTS: Rats exhibited liver fibrosis after HFSCD feeding and the administration of TAA. Caffeine could reduce the hepatic level of collagen and the fibrotic area in the liver. Caffeine prevented the progression of liver fibrosis by decreasing transforming growth factor-beta (TGF-ß), connective tissue growth factor (CTGF), and alpha-smooth muscle actin (α-SMA) expression and by inhibiting the activation of mitogen-activated protein kinases (MAPKs) and Smad3 phosphorylation. CONCLUSIONS: Caffeine attenuates NASH and the progression of liver fibrosis due to its antifibrotic effects and modulating the MAPK and TGF-ß pathways. Therefore, caffeine could be a suitable candidate for treating liver diseases associated with fibrosis.
Assuntos
Hepatopatia Gordurosa não Alcoólica , Tioacetamida , Animais , Cafeína/efeitos adversos , Cafeína/metabolismo , Fibrose , Fígado/patologia , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/tratamento farmacológico , Cirrose Hepática/prevenção & controle , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/etiologia , Hepatopatia Gordurosa não Alcoólica/prevenção & controle , Ratos , Transdução de Sinais , Proteína Smad3/metabolismo , Tioacetamida/efeitos adversos , Tioacetamida/metabolismo , Fator de Crescimento Transformador beta/metabolismoRESUMO
OBJECTIVES: To investigate the efficacy and potential molecular mechanism of Huangkui capsule in combination with leflunomide (HKL) for the treatment of immunoglobulin A nephropathy (IgAN). METHODS: IgAN rat models were constructed by treating rats with bovine serum albumin, lipopolysaccharide, and tetrachloromethane. Th22 cells were isolated from the blood samples of patients with IgAN using a CD4+ T cell isolation kit. The expression levels of the components of the TGF-ß1/Smad3 signaling pathway, namely, TGF-ß1, Smad2, Smad3, Smad4, and Smad7, were detected using quantitative reverse transcription polymerase chain reaction. Cell proliferation was determined using the MTT assay, cell viability was determined using the WST 1 method, and the chemotaxis of Th22 cells was observed using the wound healing assay. Changes in the histology of the kidney tissues were analyzed using hematoxylin and eosin staining. RESULTS: Compared with IgAN rats, the rats subjected to HKL treatment showed good improvement in kidney injuries, and the combined drug treatment performed much better than the single-drug treatment. In addition, following HKL treatment, the viability, proliferation, and chemotaxis of Th22 cells dramatically decreased (*p<0.05, **p<0.01, and ***p<0.001). In addition, CCL20, CCL22, and CCL27 levels decreased and the expression of the key components of the TGF-ß1/Smad3 signaling pathway was downregulated in IgAN rats and Th22 cells (*p<0.05, ***p<0.001). CONCLUSIONS: By targeting the TGF-ß1/Smad3 signaling pathway, HKL treatment can improve kidney injury in IgAN rats as well as the excessive proliferation and metastasis of Th22 cells.
Assuntos
Medicamentos de Ervas Chinesas , Glomerulonefrite por IGA , Leflunomida , Proteína Smad3 , Fator de Crescimento Transformador beta1 , Animais , Medicamentos de Ervas Chinesas/farmacologia , Glomerulonefrite por IGA/tratamento farmacológico , Glomerulonefrite por IGA/metabolismo , Humanos , Rim/metabolismo , Leflunomida/farmacologia , Ratos , Transdução de Sinais , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/metabolismoRESUMO
Octreotide (OCT) is used to inhibit hormone secretion and growth in somatotroph tumors, although a significant percentage of patients are resistant. It has also been tested in nonfunctioning (NF) tumors but with poor results, with these outcomes having been associated with SSTR2 levels and impaired signaling. We investigated whether OCT inhibitory effects can be improved by TGF-ß1 in functioning and nonfunctioning somatotroph tumor cells. OCT effects on hormone secretion and proliferation were analyzed in the presence of TGF-ß1 in WT and SSTR2-overexpressing secreting GH3 and silent somatotroph tumor cells. The mechanism underlying these effects was assessed by studying SSTR and TGFßR signaling pathways mediators. In addition, we analyzed the effects of OCT/TGF-ß1 treatment on tumor growth and cell proliferation in vivo. The inhibitory effects of OCT on GH- and PRL-secretion and proliferation were improved in the presence of TGF-ß1, as well as by SSTR2 overexpression. The OCT/TGF-ß1 treatment induced downregulation of pERK1/2 and pAkt, upregulation of pSmad3, and inhibition of cyclin D1. In vivo experiments showed that OCT in the presence of TGF-ß1 blocked tumor volume growth, decreased cell proliferation, and increased tumor necrosis. These results indicate that SSTR2 levels and the stimulation of TGF-ß1/TGFßR/Smad2/3 pathway are important for strengthening the antiproliferative and antisecretory effects of OCT.
Assuntos
Antineoplásicos Hormonais/farmacologia , Proliferação de Células/efeitos dos fármacos , Octreotida/farmacologia , Neoplasias Hipofisárias/tratamento farmacológico , Proteína Smad2/metabolismo , Proteína Smad3/metabolismo , Somatotrofos/efeitos dos fármacos , Fator de Crescimento Transformador beta1/farmacologia , Animais , Linhagem Celular , Feminino , Humanos , Camundongos Nus , Fosforilação , Neoplasias Hipofisárias/genética , Neoplasias Hipofisárias/metabolismo , Neoplasias Hipofisárias/patologia , Ratos , Receptores de Somatostatina/genética , Receptores de Somatostatina/metabolismo , Transdução de Sinais , Somatotrofos/metabolismo , Somatotrofos/patologia , Carga Tumoral/efeitos dos fármacosRESUMO
OBJECTIVES: To investigate the efficacy and potential molecular mechanism of Huangkui capsule in combination with leflunomide (HKL) for the treatment of immunoglobulin A nephropathy (IgAN) METHODS: IgAN rat models were constructed by treating rats with bovine serum albumin, lipopolysaccharide, and tetrachloromethane. Th22 cells were isolated from the blood samples of patients with IgAN using a CD4+ T cell isolation kit. The expression levels of the components of the TGF-β1/Smad3 signaling pathway, namely, TGF-β1, Smad2, Smad3, Smad4, and Smad7, were detected using quantitative reverse transcription polymerase chain reaction. Cell proliferation was determined using the MTT assay, cell viability was determined using the WST 1 method, and the chemotaxis of Th22 cells was observed using the wound healing assay. Changes in the histology of the kidney tissues were analyzed using hematoxylin and eosin staining. RESULTS: Compared with IgAN rats, the rats subjected to HKL treatment showed good improvement in kidney injuries, and the combined drug treatment performed much better than the single-drug treatment. In addition, following HKL treatment, the viability, proliferation, and chemotaxis of Th22 cells dramatically decreased (*p<0.05, **p<0.01, and ***p<0.001). In addition, CCL20, CCL22, and CCL27 levels decreased and the expression of the key components of the TGF-β1/Smad3 signaling pathway was downregulated in IgAN rats and Th22 cells (*p<0.05, ***p<0.001). CONCLUSIONS: By targeting the TGF-β1/Smad3 signaling pathway, HKL treatment can improve kidney injury in IgAN rats as well as the excessive proliferation and metastasis of Th22 cells.
Assuntos
Humanos , Animais , Ratos , Medicamentos de Ervas Chinesas/farmacologia , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Leflunomida/farmacologia , Glomerulonefrite por IGA/metabolismo , Glomerulonefrite por IGA/tratamento farmacológico , Transdução de Sinais , Rim/metabolismoRESUMO
Transforming growth factor beta (TGFß) signalling is involved in several aspects of regeneration in many organs and tissues of primitive vertebrates. It has been difficult to recognize the role of this signal in mammal regeneration due to the low ability of this animal class to reconstitute tissues. Nevertheless, ear-holes in middle-age female mice represent a model to study the limited epimorphic-like regeneration in mammals. Using this model, in this study we explored the possible participation of TGFß signalling in mammal regeneration. Positive pSmad3 cells, as well as TGFß1 and TGFß3 isoforms, were detected during the redifferentiation phase in the blastema-like structure. Daily administration of the inhibitor of the TGFß intracellular pathway, SB431542, during 7 days from the re-differentiation phase, resulted in a decreased level of pSmad3 accompanied by a transitory higher growth of the new tissue, larger cartilage nodules, and new muscle formation. These phenotypes were associated with a decrease in the number of α-SMA-positive cells and loose packing of collagen I. These results indicate that the modulation of the fibrosis mediated by TGFß signalling contributes to enhancing the differentiation of cartilage and muscle during limited ear-hole regeneration.
Assuntos
Diferenciação Celular/fisiologia , Orelha/fisiopatologia , Regeneração/fisiologia , Transdução de Sinais/fisiologia , Fator de Crescimento Transformador beta/metabolismo , Animais , Benzamidas/farmacologia , Diferenciação Celular/efeitos dos fármacos , Dioxóis/farmacologia , Orelha/patologia , Proteínas da Matriz Extracelular/metabolismo , Feminino , Fibrose , Camundongos Endogâmicos BALB C , Microscopia de Fluorescência/métodos , Regeneração/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta3/metabolismoRESUMO
INTRODUCTION AND OBJECTIVES: Curcumin, a polyphenol, is a natural compound that has been widely studied as a hepatoprotector; however, only a few studies have examined its ability to reduce fibrosis in previously established cirrhosis. The objective of this study was to investigate whether curcumin could reduce carbon tetrachloride (CCl4)-induced fibrosis and if so, to determine the action mechanisms involved in the reduction process. MATERIALS AND METHODS: CCl4 was administered to male Wistar rats (400â¯mg/kg, three times a week, i. p.) for 12 weeks; curcumin (100â¯mg/kg body weight twice per day, p. o.) was administered from week 9-12 of CCl4 treatment. Biochemical markers of hepatic injury and oxidative stress were evaluated. Hematoxylin and eosin, Masson's trichrome stains, transmission electron microscopy; immunohistochemistry, and zymography assays were carried out. Moreover, Smad3 and α-SMA mRNA and protein levels were studied. Western blotting by TGF-ß, CTGF, Col-I, MMP-13, NF-κB, IL-1, IL-10, Smad7, pSmad3, and pJNK proteins was developed. RESULTS AND CONCLUSIONS: Curcumin reduced liver damage, oxidative stress, fibrosis, and restored normal activity of MMP-9 and MMP-2. Besides, curcumin restored NF-κB, IL-1, IL-10, TGF-ß, CTGF, Col-I, MMP-13, and Smad7 protein levels. On the other hand, curcumin decreased JNK and Smad3 phosphorylation. Furthermore, curcumin treatment decreased α-SMA and Smad3 protein and mRNA levels. Curcumin normalized GSH, and NF-κB, JNK-Smad3, and TGF-ß-Smad3 pathways, leading to a decrement in activated hepatic stellate cells, thereby producing its antifibrotic effects.
Assuntos
Transdiferenciação Celular/efeitos dos fármacos , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Curcumina/farmacologia , Células Estreladas do Fígado/efeitos dos fármacos , Cirrose Hepática Experimental/prevenção & controle , Fígado/efeitos dos fármacos , Substâncias Protetoras/farmacologia , Proteína Smad3/metabolismo , Proteína Smad7/metabolismo , Animais , Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Tetracloreto de Carbono , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/patologia , Citocinas/metabolismo , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/ultraestrutura , Fígado/metabolismo , Fígado/ultraestrutura , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/metabolismo , Cirrose Hepática Experimental/patologia , Masculino , Estresse Oxidativo/efeitos dos fármacos , Fosforilação , Ratos Wistar , Transdução de SinaisRESUMO
Poor prognosis associated with the dysregulated expression of activin A in a number of malignancies has been related to with numerous aspects of tumorigenesis, including angiogenesis. The present study investigated the prognostic significance of activin A immunoexpression in blood vessels and cancer cells in a number of oral squamous cell carcinoma (OSCC) cases and applied in vitro strategies to determine the impact of activin A on angiogenesis. In a cohort of 95 patients with OSCC, immunoexpression of activin A in both blood vessels and tumor cells was quantified and the association with clinicopathological parameters and survival was analyzed. Effects of activin A on the tube formation, proliferation and migration of human umbilical vein endothelial cells (HUVECs) were evaluated in gainoffunction (treatment with recombinant activin A) or lossoffunction [treatment with activin Aantagonist follistatin or by stable transfection with short hairpin RNA (shRNA) targeting activin A] conditions. Conditioned medium from an OSCC cell line with shRNAmediated depletion of activin A was also tested. The profile of pro and antiangiogenic factors regulated by activin A was assessed with a human angiogenesis quantitative PCR (qPCR) array. Vascular endothelial growth factor A (VEGFA) and its major isoforms were evaluated by reverse transcriptionqPCR and ELISA. Activin A expression in blood vessels demonstrated an independent prognostic value in the multivariate analysis with a hazard ratio of 2.47 [95% confidence interval (CI), 1.304.71; P=0.006) for diseasespecific survival and 2.09 (95% CI, 1.074.08l: P=0.03) for diseasefree survival. Activin A significantly increased tubular formation of HUVECs concomitantly with an increase in proliferation. This effect was validated by reduced proliferation and tubular formation of HUVECs following inhibition of activin A by follistatin or shRNA, as well as by treatment of HUVECs with conditioned medium from activin Adepleted OSCC cells. Activin Aknockdown increased the migration of HUVECs. In addition, activin A stimulated the phosphorylation of SMAD2/3 and the expression and production of total VEGFA, significantly enhancing the expression of its proangiogenic isoform 121. The present findings suggest that activin A is a predictor of the prognosis of patients with OSCC, and provide evidence that activin A, in an autocrine and paracrine manner, may contribute to OSCC angiogenesis through differential expression of the isoform 121 of VEGFA.
Assuntos
Ativinas/metabolismo , Neoplasias Bucais/patologia , Neovascularização Patológica/patologia , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Ativinas/análise , Ativinas/antagonistas & inibidores , Ativinas/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Comunicação Autócrina/efeitos dos fármacos , Comunicação Autócrina/genética , Movimento Celular , Proliferação de Células , Feminino , Folistatina/farmacologia , Folistatina/uso terapêutico , Técnicas de Silenciamento de Genes , Células Endoteliais da Veia Umbilical Humana , Humanos , Masculino , Pessoa de Meia-Idade , Mucosa Bucal/patologia , Neoplasias Bucais/irrigação sanguínea , Neoplasias Bucais/tratamento farmacológico , Neoplasias Bucais/mortalidade , Comunicação Parácrina/efeitos dos fármacos , Comunicação Parácrina/genética , Fosforilação/efeitos dos fármacos , Fosforilação/genética , Prognóstico , Isoformas de Proteínas/metabolismo , Proteína Smad2/metabolismo , Proteína Smad3/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/irrigação sanguínea , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Carcinoma de Células Escamosas de Cabeça e Pescoço/mortalidadeRESUMO
Colorectal Cancer (CRC) therapy confronts challenges as chemoresistance and side effects. Therefore, drugs with antitumor properties that downmodulate aggressiveness mediators are required. Studies have shown the relevance of Low Molecular Weight Protein Tyrosine Phosphatase (LMWPTP), Protein Tyrosine Phosphatase 1B (PTP1B), and Transforming Growth Factor ß (TGFß) in mediating proliferation, chemoresistance, and metastasis. In this study, we aimed to investigate the responsiveness of colorectal cancer lines (HT29 and HCT116) towards Vemurafenib and whether this treatment could modulate these aggressiveness mediators. Cytotoxicity Assays (MTT and Trypan Exclusion Test) were performed to evaluate the viability of HT29 and HCT116 cells treated with Vemurafenib. Western blotting was performed to analyze the amount and/or the activity of mediators (LMWPTP, PTP1B, TGFß, SMAD3), and the immunoprecipitation was performed to evaluate LMWPTP activity. This study brought up novel aspects of Vemurafenib action in colorectal cancer, which can decrease the activity of protein tyrosine phosphatases (LMWPTP and PTP1B) and the TGFß pathway, making them important in the CRC aggressiveness. By downmodulating colorectal cancer hallmarks, Vemurafenib appears as an interesting candidate for CRC therapeutic protocols.