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1.
Braz J Med Biol Res ; 52(10): e8324, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31596310

RESUMO

The aim of this study was to investigate the role of kinase-insert domain-containing receptor (KDR) in intrauterine adhesions (IUA) and its mechanism. The Case group consisted of 92 patients diagnosed with IUA, and the Control group included 86 patients with uterine septum who had normal endometrium verified with an uteroscope. In addition, 50 rats were randomly assigned into Control, Sham, Model, NC-siRNA, and KDR-siRNA groups. Rats in the Model, NC-siRNA, and KDR-siRNA groups were induced by uterine curettage and lipopolysaccharide (LPS) treatment to establish the IUA model. Then, immunohistochemistry was applied for detection of VEGF and KDR expression, HE staining was used for observation of the endometrial morphology and gland counting, Masson staining for measurement of the degree of endometrial fibrosis, and qRT-PCR and western blot for the expression of KDR, VEGF, MMP-9, as well as TGF-ß1/Smads pathway-related proteins. Compared with the Control group, the mRNA and protein expressions of KDR were significantly higher in IUA endometrial tissues, and the expression of KDR was positively correlated to the severity of IUA. In addition, the injection of si-KDR increased the number of endometrial glands, reduced the area of fibrosis, inhibited mRNA and protein expression of KDR and VEGF, up-regulated the expression of MMP-9 and Smad7, and decreased the expression level of TGF-ß1, p-Smad2, p-Smad3, and Smad4 in rats with IUA. Highly-expressed KDR was related to patients' severity of IUA, and silencing KDR may prevent the occurrence and development of IUA via TGF-ß1/Smads signaling pathway and up-regulating the expression of MMP-9.


Assuntos
Transdução de Sinais , Proteínas Smad/metabolismo , Aderências Teciduais/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Doenças Uterinas/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Adulto , Animais , Western Blotting , Estudos de Casos e Controles , Modelos Animais de Doenças , Feminino , Humanos , Imuno-Histoquímica , Pessoa de Meia-Idade , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Índice de Gravidade de Doença , Proteínas Smad/genética , Aderências Teciduais/patologia , Fator de Crescimento Transformador beta1/genética , Doenças Uterinas/patologia , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Adulto Jovem
2.
J Biol Regul Homeost Agents ; 33(5): 1337-1345, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31637903

RESUMO

The effects of miR-145 (microRNA 145) on M. pneumoniae (MP)-infected MRC-5 (Medical Research Council cell strain 5) cell TGF-ß/Smad (transforming growth factor beta/Smad) fibrosis pathway were explored through constructing MP-infected MRC-5 cell models. In addition, the qPCR (quantitative real-time polymerase chain reaction) and Western blot were applied to detect the mRNA and protein expressions of miR-145, TGF-ß1 (transforming growth factor beta 1), Smad3, Smad4, MMP2 (matrix metalloproteinase 2), FN1 (fibronectin 1), ELN (elastin) and COLI α1 (collagen type I alpha 1) signaling molecules in TGF-ß/Smad fibrosis pathway. The results showed that the expression of miR-145 in MRC-5 cells was significantly increased after MP infection. In addition, miR-145 inhibited the fibrosis promoting TGF-ß/Smad pathway by targeting Smad3, a key factor in the TGF-ß/Smad pathway. It can be concluded that, in the process of MP infection, the expression of miR-145 is stimulated to negatively regulate the fibrosis-promoting pathway of TGF-ß/Smad.


Assuntos
Fibroblastos/patologia , MicroRNAs/metabolismo , Mycoplasma pneumoniae , Proteínas Smad/metabolismo , Linhagem Celular , Fibroblastos/microbiologia , Fibrose , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Fator de Crescimento Transformador beta1/metabolismo
3.
Cell Prolif ; 52(5): e12633, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31264317

RESUMO

OBJECTIVES: Matrix metalloproteinase 9 (MMP-9) has been frequently noticed in the breast cancers. In this study, we aim to investigate the associations of MMP-9 with the activation of transforming growth factor beta (TGF-ß)/SMAD signalling and the malignancy of breast malignant tumour cells. MATERIALS AND METHODS: The distributions of MMP-9 and TGF-ß in the tissues of canine breast cancers were screened by immunohistochemical assays. A recombinant plasmid expressing mouse MMP-9 was generated and transiently transfected into three different breast cancer cell lines. Cell Counting Kit-8 and colony formation assay were used to study cell viability. Migration and invasion ability were analysed by wound assay and transwell filters. Western blot and quantitative real-time PCR were used to determine the protein and mRNA expression. RESULT: Remarkable strong MMP-9 and TGF-ß signals were observed in the malignant tissues of canine breast cancers. In the cultured three cell lines receiving recombinant plasmid expressing mouse MMP-9, the cell malignancy was markedly increased, including the cell colony formation, migration and epithelial-mesenchymal transition. The levels of activated TGF-ß, as well as SMAD4, SMAD2/3 and phosphorylation of SMAD2, were increased, reflecting an activation of TGF-ß/SMAD signalling. We also demonstrated that the inhibitors specific for MMP-9 and TGF-ß sufficiently blocked the overexpressing MMP-9 induced the activation of SMAD signalling and enhancement on invasion in the tested breast cancer cell lines. CONCLUSION: Overexpression of MMP-9 increases the malignancy of breast cancer cell lines, largely via activation of the TGF-ß/SMAD signalling.


Assuntos
Metaloproteinase 9 da Matriz/metabolismo , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Transição Epitelial-Mesenquimal , Feminino , Humanos , Metaloproteinase 9 da Matriz/química , Metaloproteinase 9 da Matriz/genética , Fosforilação , Transdução de Sinais , Fator de Crescimento Transformador beta/antagonistas & inibidores
4.
Life Sci ; 232: 116669, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31326566

RESUMO

AIMS: This study investigated the effects of hyaluronic acid (HA), a commonly used osteogenic medium referred to as DAG, and the combined administration of HA and DAG (CG) on the osteogenic differentiation of human amniotic mesenchymal stem cells (hAMSCs), and the underlying mechanism. MAIN METHODS: The phenotype of hAMSCs was detected by flow cytometry and immunocytochemical staining. Alkaline phosphatase (ALP) and calcium deposition assays were employed for evaluating the osteogenic differentiation of hAMSCs. The expression of osteogenesis-related genes and proteins was determined by quantitative reverse transcription PCR (qRT-PCR) and Western blotting, respectively. Meanwhile, the molecular mechanism of osteogenic differentiation of hAMSCs was detected by PCR array and qRT-PCR. KEY FINDINGS: The results showed that treatment with CG could significantly stimulate hAMSC ALP activity and calcium deposition compared to treatment with DAG, while HA had little effect. The expression of osteogenesis-related molecules and stemness-related molecules was up-regulated at the mRNA and protein levels in all three groups, and this up-regulation was most significant in the CG group. In addition, treatment with CG significantly increased the gene expressions involved in regulation of the TGF-ß/Smad signalling pathway compared to treatment with DAG. Furthermore, the pro-osteogenic differentiation effects as well as the up-regulated expression of genes observed in the CG treatment group were significantly inhibited when the cells were pre-treated with SB431542, an inhibitor of the TGF-ß/Smad pathway. SIGNIFICANCE: These results suggest that HA in combination with DAG could significantly enhance the osteogenic differentiation of hAMSCs, potentially via the TGF-ß/Smad signalling pathway.


Assuntos
Âmnio/citologia , Diferenciação Celular/efeitos dos fármacos , Ácido Hialurônico/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Transdução de Sinais , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Células Cultivadas , Humanos , Ácido Hialurônico/química , Células-Tronco Mesenquimais/citologia , Peso Molecular
5.
Life Sci ; 231: 116674, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31344427

RESUMO

Hypertrophic scar formation is a fibroproliferative disorder caused by abnormal wound healing. At present, there are limited treatment strategies for hypertrophic scars. In this study, we identified an endogenous peptide, LYENRL, through peptidomics screening that is downregulated in scar skin tissues. The peptide exhibited concentration dependent inhibitory effects on the proliferation, migration and extracellular matrix (ECM) production of scar fibroblasts. By eukaryotic transcriptome sequencing analysis, we noted that LYENRL downregulated gene sets in scar fibroblasts were associated with the transforming growth factor-ß (TGF-ß) signaling pathway. Further experiments revealed that LYENRL was able to inhibit the activation of TGF-ß1/Smad signaling and TGF-ß1-induced activation of scar fibroblasts at the source by blocking the binding of AP-1 to the corresponding region of the Tgfb1 promoter, which in turn inhibited gene expression of Tgfb1. Taken together, we concluded that the effects of LYENRL on scar fibroblasts make it a potential peptide drug for hypertrophic scar treatment.


Assuntos
Cicatriz Hipertrófica/metabolismo , Cicatriz Hipertrófica/patologia , Peptídeos/farmacologia , Actinas/metabolismo , Linhagem Celular , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Matriz Extracelular/metabolismo , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/fisiologia , Humanos , Peptídeos/metabolismo , Cultura Primária de Células , Transdução de Sinais/efeitos dos fármacos , Pele/metabolismo , Proteínas Smad/metabolismo , Proteínas Smad/fisiologia , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta1/fisiologia
6.
Cancer Sci ; 110(9): 2834-2845, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31278880

RESUMO

Recurrence and chemoresistance in colorectal cancer remain important issues for patients treated with conventional therapeutics. Metformin and phenformin, previously used in the treatment of diabetes, have been shown to have anticancer effects in various cancers, including breast, lung and prostate cancers. However, their molecular mechanisms are still unclear. In this study, we examined the effects of these drugs in chemoresistant rectal cancer cell lines. We found that SW837 and SW1463 rectal cancer cells were more resistant to ionizing radiation and 5-fluorouracil than HCT116 and LS513 colon cancer cells. In addition, metformin and phenformin increased the sensitivity of these cell lines by inhibiting cell proliferation, suppressing clonogenic ability and increasing apoptotic cell death in rectal cancer cells. Signal transducer and activator of transcription 3 and transforming growth factor-ß/Smad signaling pathways were more activated in rectal cancer cells, and inhibition of signal transducer and activator of transcription 3 expression using an inhibitor or siRNA sensitized rectal cancer cells to chemoresistant by inhibition of the expression of antiapoptotic proteins, such as X-linked inhibitor of apoptosis, survivin and cellular inhibitor of apoptosis protein 1. Moreover, metformin and phenformin inhibited cell migration and invasion by suppression of transforming growth factor ß receptor 2-mediated Snail and Twist expression in rectal cancer cells. Therefore, metformin and phenformin may represent a novel strategy for the treatment of chemoresistant rectal cancer by targeting signal transducer and activator of transcription 3 and transforming growth factor-ß/Smad signaling.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Metformina/farmacologia , Fenformin/farmacologia , Neoplasias Retais/terapia , Transdução de Sinais/efeitos dos fármacos , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/efeitos da radiação , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/efeitos da radiação , Quimiorradioterapia/métodos , Colo/patologia , Neoplasias do Colo/patologia , Neoplasias do Colo/terapia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos da radiação , Fluoruracila/farmacologia , Fluoruracila/uso terapêutico , Humanos , Masculino , Metformina/uso terapêutico , Camundongos , Camundongos Nus , Recidiva Local de Neoplasia , Fenformin/uso terapêutico , Neoplasias Retais/patologia , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos da radiação , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
7.
Artigo em Chinês | MEDLINE | ID: mdl-31177710

RESUMO

Objective: To investigate the possible mechanism of doxycycline inhibiting paraquat-induced pulmonary fibrosis and provide a theoretical basis for its clinical application. Methods: Human lung fibroblast HFL1 cells were selected as the research object in the cell group. Divided into blank group, paraquat group, paraquat+doxycycline group. The expression of TGF-ß1, a-SMA, Smad3 and Smad2 protein was detected by ELISA using 40 ml of paraquat 40 umol/L and 3 mg/L of oleic acid 10 mg/L. In the animal group, 120 healthy and clean SD rats were randomly divided into three groups: blank group, paraquat group, paraquat+doxycycline group. The expression of TGF-ß1, a-SMA, Smad3 and Smad2 protein in lung tissue of mice at 1 day, 3 days, 7 days, 14 days and 21 days was detected by Elisa method. The expression of TGF-ß1, a-SMA, Smad3 and Smad2 protein in lung tissue of 21-day mice was detected by Western Blotting. The pathological changes of lung tissue were observed by HE staining for 1 day, 3 days, 7 days, 14 days and 21 days. Results: In the cell group experiment, the expression of TGF-ß1, a-SMA, Smad3 and Smad2 protein increased gradually with paraquat in the paraquat group, and the expression of TGF-ß1, a-SMA, Smad3 and Smad2 protein was significantly higher than that in the blank group. The difference was statistically significant (P<0.05) . The expressions of TGF-ß1, a-SMA, Smad3 and Smad2 in the paraquat+doxycycline group were significantly lower than those in the paraquat group, but still higher than the blank group, the difference was statistically significant (P<0.05) . Conclusion: Doxycycline inhibits paraquat-induced pulmonary fibrosis by inhibiting the expression of TGF-ß1, a-SMA and Smad3, Smad2 proteins.


Assuntos
Antibacterianos , Doxiciclina , Paraquat , Fibrose Pulmonar , Proteínas Smad , Fator de Crescimento Transformador beta1 , Animais , Antibacterianos/farmacologia , Doxiciclina/farmacologia , Humanos , Camundongos , Paraquat/toxicidade , Fibrose Pulmonar/induzido quimicamente , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Proteínas Smad/efeitos dos fármacos , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta1/efeitos dos fármacos , Fator de Crescimento Transformador beta1/metabolismo
8.
Int J Nanomedicine ; 14: 3645-3667, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31190817

RESUMO

Background: Neo-adjuvant chemotherapy is an effective strategy for improving treatment of breast cancers. However, the efficacy of this treatment strategy is limited for treatment of triple negative breast cancer (TNBC). Gene therapy may be a more effective strategy for improving the prognosis of TNBC. Methods: A novel 25 nucleotide sense strand of miRNA was designed to treat TNBC by silencing the Slug gene, and encapsulated into DSPE-PEG2000-tLyp-1 peptide-modified functional liposomes. The efficacy of miRNA liposomes was evaluated on invasive TNBC cells and TNBC cancer-bearing nude mice. Furthermore, functional vinorelbine liposomes were constructed to investigate the anticancer effects of combined treatment. Results: The functional miRNA liposomes had a round shape and were nanosized (120 nm). Functional miRNA liposomes were effectively captured by TNBC cells in vitro and were target to mitochondria. Treatment with functional liposomes silenced the expression of Slug and Slug protein, inhibited the TGF-ß1/Smad pathway, and inhibited invasiveness and growth of TNBC cells. In TNBC cancer-bearing mice, functional miRNA liposomes exerted a stronger anticancer effect than functional vinorelbine liposomes, and combination therapy with these two formulations resulted in nearly complete inhibition of tumor growth. Preliminary safety evaluations indicated that the functional miRNA liposomes did not affect body weight or cause damage to any major organs. Furthermore, the functional liposomes significantly increased the half-life of the drug in the blood of cancer-bearing nude mice, and increased drug accumulation in breast cancer tissues. Conclusion: In this study, we constructed novel functional miRNA liposomes. These liposomes silenced Slug expression and inhibited the TGF-ß1/Smad pathway in TNBC cells, and enhanced anticancer efficacy in mice using combined chemotherapy. Hence, the present study demonstrated a promising strategy for gene therapy of invasive breast cancer.


Assuntos
Inativação Gênica , MicroRNAs/metabolismo , Nanopartículas/química , Tamanho da Partícula , Fatores de Transcrição da Família Snail/genética , Neoplasias de Mama Triplo Negativas/terapia , Animais , Antineoplásicos/farmacologia , Apoptose , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Endocitose/efeitos dos fármacos , Feminino , Humanos , Lipossomos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , Invasividade Neoplásica , Fosfatidiletanolaminas/química , Polietilenoglicóis/química , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Neoplasias de Mama Triplo Negativas/patologia
9.
Hua Xi Kou Qiang Yi Xue Za Zhi ; 37(2): 162-167, 2019 Apr 01.
Artigo em Chinês | MEDLINE | ID: mdl-31168982

RESUMO

OBJECTIVE: The purpose of this study is to investigate the potential effects of sclerostin (SOST) on the biological funtions and related mechanisms of cementoblasts under mechanical stress. METHODS: OCCM-30 cells were treated with varying doses of SOST (0, 25, 50, and 100 ng·mL⁻¹) and were loaded with uniaxial compressive stress (2 000 µ strain with a frequency of 0.5 Hz) for six hours. Western blot was utilized to detect the expressions of ß-catenin, p-smad1/5/8, and smad1/5/8 proteins. Alkaline phosphatase (ALP) activity was determined, and reverse transcription polymerase chain reaction was used to measure the expressions of runt-related transcription factor 2 (Runx-2), osteocalcin (OCN), bone sialoproteins (BSP), receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG) mRNA. RESULTS: The expression of p-smad
1/5/8 was significantly downregulated with increasing SOST. ß-catenin and smad1/5/8 exhibited no difference. ALP activity decreased under mechanical compressive stress with increasing SOST concentrations. Runx-2 expression was reduced with increasing SOST concentrations, and a similar trend was observed for the BSP and OCN expressions. When the SOST concentration was enhanced, RANKL expression gradually increased, whereas the expression of OPG decreased. CONCLUSIONS: Under mechanical comprehensive stress, SOST can adjust the bone morphogenetic protein (BMP) /smad signal pathway. Osteosclerosis inhibits the mineralization of cementoblasts under mechanical compressive stress, which may be achieved by inhibiting the expressions of osteogenesis factors (Runx2, OCN, BSP, and others) and by promoting the ratio of cementoclast-related factors (RANKL/OPG) through BMP signal pathways.


Assuntos
Proteínas Morfogenéticas Ósseas , Cemento Dentário , Proteínas Morfogenéticas Ósseas/metabolismo , Subunidade alfa 1 de Fator de Ligação ao Core , Osteocalcina , Proteínas Smad/metabolismo , Estresse Mecânico
10.
Nat Commun ; 10(1): 2824, 2019 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-31249305

RESUMO

The fibrogenic response in tissue-resident fibroblasts is determined by the balance between activation and repression signals from the tissue microenvironment. While the molecular pathways by which transforming growth factor-1 (TGF-ß1) activates pro-fibrogenic mechanisms have been extensively studied and are recognized critical during fibrosis development, the factors regulating TGF-ß1 signaling are poorly understood. Here we show that macrophage hypoxia signaling suppresses excessive fibrosis in a heart via oncostatin-m (OSM) secretion. During cardiac remodeling, Ly6Chi monocytes/macrophages accumulate in hypoxic areas through a hypoxia-inducible factor (HIF)-1α dependent manner and suppresses cardiac fibroblast activation. As an underlying molecular mechanism, we identify OSM, part of the interleukin 6 cytokine family, as a HIF-1α target gene, which directly inhibits the TGF-ß1 mediated activation of cardiac fibroblasts through extracellular signal-regulated kinase 1/2-dependent phosphorylation of the SMAD linker region. These results demonstrate that macrophage hypoxia signaling regulates fibroblast activation through OSM secretion in vivo.


Assuntos
Fibrose/metabolismo , Hipóxia/metabolismo , Macrófagos/metabolismo , Oncostatina M/metabolismo , Animais , Antígenos Ly/genética , Antígenos Ly/metabolismo , Feminino , Fibroblastos/metabolismo , Fibrose/genética , Fibrose/patologia , Hipóxia/genética , Hipóxia/patologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Oncostatina M/genética , Fosforilação , Transdução de Sinais , Proteínas Smad/genética , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta1/metabolismo
11.
Parasit Vectors ; 12(1): 279, 2019 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-31151477

RESUMO

BACKGROUND: Activation of hepatic stellate cells is the dominant pathogenic event during the process of liver fibrosis. Bone morphogenic protein (BMP)-7 has recently been identified as an anti-fibrotic factor and leads to phosphorylation of Smad1/5/8 in activated hepatic stellate cells. Its expression can be upregulated by the transcriptional activator, Y-Box protein-1 (YB1). Previous studies have found that the recombinant Schistosoma japonicum protein p40 (rSjp40) can inhibit the activation of hepatic stellate cells, and based on this evidence we attempted to investigate whether or not BMP-7 is involved in rSjp40's inhibition. METHODS: A human hepatic stellate cell line, the LX-2 cell line, was cultured and treated with rSjp40. The role of BMP-7 was analyzed by Western blot. RESULTS: Our findings testified that knockdown of BMP-7 impaired rSjp40-induced downregulation of α-SMA and phosphorylation of Smad1/5/8 in LX-2 cells. Furthermore, rSjp40 upregulated expression of BMP-7 at both mRNA and protein levels depending on YB1. Interestingly, YB1 was translocated from the cytoplasm to the nucleus upon treatment of rSjp40. CONCLUSIONS: These results suggest that rSjp40 inhibits the activation of hepatic stellate cells by promoting nuclear translocation of YB1 and inducing BMP-7/Smad1/5/8 pathway, which provide a new clue to guide ongoing research into the anti-fibrosis of rSjp40.


Assuntos
Antígenos de Helmintos/metabolismo , Proteína Morfogenética Óssea 7/metabolismo , Proteínas de Helminto/metabolismo , Células Estreladas do Fígado/parasitologia , Transdução de Sinais , Proteínas Smad/metabolismo , Proteína 1 de Ligação a Y-Box/metabolismo , Animais , Linhagem Celular , Núcleo Celular/metabolismo , Humanos , Cirrose Hepática/patologia , Fosforilação , Transporte Proteico , Interferência de RNA , Proteínas Recombinantes/farmacologia , Schistosoma japonicum , Proteína Smad1/metabolismo , Proteína Smad5/metabolismo , Proteína Smad8/metabolismo , Proteína 1 de Ligação a Y-Box/genética
12.
Cell Prolif ; 52(4): e12631, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31038259

RESUMO

OBJECTIVES: Growth differentiation factor 11 (GDF11), an emerging secreted member of the TGF-beta superfamily, plays essential roles in development, physiology and multiple diseases; however, its role during adipogenic differentiation and the underlying mechanisms remains poorly understood. MATERIALS AND METHODS: Bone marrow-derived human mesenchymal stem cells (hMSCs) and 3T3-L1 pre-adipocytes were induced with adipogenic culture medium supplementing with different concentrations of recombinant GDF11 (rGDF11 0, 10, 50, 100 ng mL-1 ). Oil Red O staining, qRT-PCR analysis, Western blot analysis and immunofluorescence staining were performed to assay adipogenesis. RESULTS: For both hMSCs and 3T3-L1 pre-adipocytes, the presence of rGDF11 leads to a dose-dependent reduction of intracellular lipid droplet accumulation and suppressed adipogenic-related gene expression. Mechanically, GDF11 inhibits adipogenesis by activating Smad2/3-dependent TGF-beta signalling pathway, and these inhibitory effects could be restored by SB-431542, a pharmacological TGF-beta type I receptor inhibitor. CONCLUSIONS: Taken together, our data indicates that GDF11 inhibits adipogenic differentiation in both hMSCs and 3T3-L1 pre-adipocytes by activating Smad2/3-dependent TGF-beta signalling pathway.


Assuntos
Adipócitos/metabolismo , Adipogenia/fisiologia , Proteínas Morfogenéticas Ósseas/metabolismo , Diferenciação Celular/fisiologia , Fatores de Diferenciação de Crescimento/metabolismo , Transdução de Sinais/fisiologia , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Células 3T3-L1 , Animais , Linhagem Celular , Expressão Gênica/fisiologia , Humanos , Células-Tronco Mesenquimais/metabolismo , Camundongos
13.
Biomed Pharmacother ; 114: 108801, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30928803

RESUMO

BACKGROUND: Diabetic nephropathy (DN) is one of the most serious and dangerous chronic complications of diabetes mellitus.Panax notoginseng has been widely used with great efficacy in the long-term treatment of kidney disease. However, the mechanism by which it exerts its effects has not been fully elucidated. AIM: We sought to identify the major components ofPanax notoginseng that are effective in reducing the symptoms of DN in vitro and in vivo. METHODS: Inhibition of cell proliferation and collagen secretion were used to screen the ten most highly concentrated components ofPanax notoginseng. The STZ-induced DN rat model on a high-fat-high-glucose diet was used to investigate the renal protective effect of Panax notoginseng and dencichine and their underlying molecular mechanisms. RESULTS: Among the ten components analysed, dencichine (ß-N-oxalyl-L-α,ß-diaminopropionic acid) was the most protective against DN. Dencichine andPanax notoginseng attenuated glucose and lipid metabolic disorders in STZ-induced DN rats on a high-fat-high-glucose diet. In the untreated DN rats, we observed albuminuria, renal failure, and pathological changes. However, treatment with dencichine and Panax notoginseng alleviated these symptoms. We also observed that dencichine suppressed the expression of TGF-ß1 and Smad2/3, which mediates mesangial cell proliferation and extracellular matrix (ECM) accumulation in the glomerulus, and enhanced the expression of Smad7, the endogenous inhibitor of the TGF-ß1/Smad signalling pathway. CONCLUSION: From these results, we concluded that dencichine is the main compound inPanax notoginseng that is responsible for alleviating renal injury in the experimental DN model. Its mechanism may be related to the reduction of the deposition of ECM in glomeruli and inhibition of the epithelial mesenchymal transformation (EMT) by inhibition of the TGF-ß1/Smad signalling pathway.


Assuntos
Diamino Aminoácidos/farmacologia , Diabetes Mellitus Tipo 2/complicações , Nefropatias Diabéticas/tratamento farmacológico , Medicamentos de Ervas Chinesas/farmacologia , Cinuramina/farmacologia , Panax notoginseng/química , Animais , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Nefropatias Diabéticas/metabolismo , Matriz Extracelular/efeitos dos fármacos , Rim/efeitos dos fármacos , Rim/metabolismo , Masculino , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , beta-Alanina/análogos & derivados , beta-Alanina/farmacologia
14.
Med Sci Monit ; 25: 3069-3076, 2019 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-31023998

RESUMO

BACKGROUND Diabetic nephropathy (DN) is the leading cause of end-stage renal disease. Mesenchymal stem cells (MSCs) treatment has been proved to be effective in DN models by protecting renal function and preventing fibrosis. However, the underlying mechanism is unclear. Previous research indicated diabetes and associated complications may be attributed to failed resolution of inflammation, which is deliberately regulated by pro-resolving lipids, including lipoxins (LXs), resolvins (Rv) D and E series, protectins, and maresins. In this study, we monitored pro-resolving mediators in a DN model to explore the mechanism of MSCs treatment. MATERIAL AND METHODS The DN model was induced by STZ injection in SD rats. UPLC-MS/MS was performed to determine pro-resolving lipids in kidney tissue and serum of DN model before and after MSCs treatment, as well as in supernatants of HBZY-1-MSCs co-culture. RESULTS LXA4 was highly accumulated in renal tissue of DN rats with MSCs treatment; ex vivo, LXA4 was significantly increased in the supernatants of HBZY-1 cells co-cultured with MSCs in a high-glucose (HG) medium. Western blot analysis indicated that ALX/FPR2, the receptor of LXA4, was markedly expressed in renal tissue of the DN-MSC group and HBZY-1 after incubating with MSCs in HG. Intraperitoneal injection of LXA4 inhibited renal fibrosis by targeting TGF-ß/Smad signaling and downregulated serum TNF-alpha, IL-6, IL-8, and IFN-γ in DN rats. Notably, all the protective effects induced by MSCs or LXA4 were abolished by ALX/FRP2 blocking. CONCLUSIONS Our results demonstrate that MSCs intervention prevented DN procession via the LXA4-ALX/FPR2 axis, which inhibited glomerulosclerosis and pro-inflammatory cytokines, eventually contributing to kidney homeostasis.


Assuntos
Citocinas/metabolismo , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/terapia , Lipoxinas/metabolismo , Transplante de Células-Tronco Mesenquimais/métodos , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Nefropatias Diabéticas/patologia , Ácidos Docosa-Hexaenoicos/metabolismo , Fibrose/metabolismo , Fibrose/patologia , Mediadores da Inflamação/metabolismo , Masculino , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais
15.
Gynecol Oncol ; 153(3): 639-650, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30928020

RESUMO

OBJECTIVE: To test whether platelets could increase invasion potential and initiate EMT in ovarian cancer cells via a TGF-ß signaling pathway. METHODS: Blood samples were collected in 69 patients with ovarian cancer, 16 patients with benign ovarian tumor and 64 healthy donors. SK-OV-3 and OVCAR-3 ovarian cancer cells were treated with platelets. Transwell assays were used to analyze the invasive capacity, and EMT was assessed by microarray analysis, quantitative real-time PCR (qPCR) and Western blotting. Activation of TGF-ß pathway was examined by ELISA and Western blotting. TGF-ß type I receptor (TßR I) inhibitor A83-01 was used to confirm the role of TGF-ß pathway in vitro and in vivo. RESULTS: Clinical data showed ovarian cancer patients with elevated platelet counts had a higher incidence of advanced stages. Treatment with platelets increased the invasive properties of both cell lines. Mesenchymal markers (snail family transcriptional repressor-1, vimentin, neural cadherin, fibronectin-1 and matrix metalloproteinase-2) were up-regulated in platelet-treated cells, while the epithelial marker (epithelial cadherin) was down-regulated. Higher TGF-ß level was observed in patients with elevated platelet counts when compared to the subjects. Higher levels of TGF-ß were also found in culture medium treated with platelets, and cells treated with platelets also showed increased phosphorylation of Smad2. TßR I inhibitor A83-01 reversed the EMT-like alterations and inhibited platelet-induced invasion in vitro and in vivo. CONCLUSION: Platelet increased invasion potential and induced EMT in ovarian cancer cells in a TGF-ß dependent pathway. Platelet-derived TGF-ß may be useful as a new target treatment for ovarian cancer.


Assuntos
Plaquetas , Transição Epitelial-Mesenquimal , Neoplasias Ovarianas/sangue , Fator de Crescimento Transformador beta/metabolismo , Adulto , Animais , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Técnicas de Cocultura , Transição Epitelial-Mesenquimal/genética , Feminino , Fibronectinas/genética , Expressão Gênica , Voluntários Saudáveis , Humanos , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , Camundongos , Pessoa de Meia-Idade , Invasividade Neoplásica , Transplante de Neoplasias , Neoplasias Ovarianas/patologia , Contagem de Plaquetas , Pirazóis/farmacologia , Receptor do Fator de Crescimento Transformador beta Tipo I/antagonistas & inibidores , Transdução de Sinais , Proteínas Smad/metabolismo , Fatores de Transcrição da Família Snail/genética , Tiossemicarbazonas/farmacologia , Fator de Crescimento Transformador beta/sangue , Regulação para Cima , Vimentina/genética
16.
Pharmazie ; 74(4): 239-242, 2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30940309

RESUMO

Diabetic nephropathy (DN) is a common cause of end-stage kidney disease (ESKD) all over the world. Sitagliptin, an inhibitor of DPP-IV plays a beneficial role in type 2 diabetic nephropathy. The purpose of this study was to explore the effect and mechanism of sitagliptin on renal injury in type 1 diabetic mice. Streptozotocin (STZ) induced type 1 diabetic mice were treated with oral administration of sitagliptin (15 mg/kg/ day) for 4 weeks. The results showed that sitagliptin treatment did not change the levels of blood glucose in STZ induced type 1 diabetic mice. Sitagliptin attenuates diabetic nephropathy by significantly inhibiting 24 h proteinuria, renal injury and fibrosis. Sitagliptin can inhibit the expression level of TGF-ß1 and the other related fibrosis factors in renal tissue of type 1 diabetic mice while delaying the progression of type 1 diabetic nephropathy. These results indicated that sitagliptin treatment is potentially a new strategy for treating type 1 diabetic nephropathy.


Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Tipo 1/tratamento farmacológico , Nefropatias Diabéticas/prevenção & controle , Inibidores da Dipeptidil Peptidase IV/farmacologia , Fosfato de Sitagliptina/farmacologia , Administração Oral , Animais , Glicemia/efeitos dos fármacos , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Tipo 1/complicações , Hipoglicemiantes/farmacologia , Masculino , Camundongos , Proteinúria/prevenção & controle , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Estreptozocina , Fator de Crescimento Transformador beta1/genética
17.
Nat Commun ; 10(1): 1089, 2019 03 06.
Artigo em Inglês | MEDLINE | ID: mdl-30842446

RESUMO

The essential roles played by Nodal and Bmp signalling during early mouse development have been extensively documented. Here we use conditional deletion strategies to investigate functional contributions made by Nodal, Bmp and Smad downstream effectors during primordial germ cell (PGC) development. We demonstrate that Nodal and its target gene Eomes provide early instructions during formation of the PGC lineage. We discover that Smad2 inactivation in the visceral endoderm results in increased numbers of PGCs due to an expansion of the PGC niche. Smad1 is required for specification, whereas in contrast Smad4 controls the maintenance and migration of PGCs. Additionally we find that beside Blimp1, down-regulated phospho-Smad159 levels also distinguishes PGCs from their somatic neighbours so that emerging PGCs become refractory to Bmp signalling that otherwise promotes mesodermal development in the posterior epiblast. Thus balanced Nodal/Bmp signalling cues regulate germ cell versus somatic cell fate decisions in the early posterior epiblast.


Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Diferenciação Celular/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Células Germinativas/fisiologia , Proteína Nodal/metabolismo , Animais , Proteínas Morfogenéticas Ósseas/genética , Linhagem Celular , Movimento Celular/fisiologia , Embrião de Mamíferos , Endoderma/citologia , Endoderma/fisiologia , Feminino , Técnicas de Inativação de Genes , Masculino , Camundongos , Camundongos Knockout , Células-Tronco Embrionárias Murinas , Proteína Nodal/genética , Transdução de Sinais/genética , Proteínas Smad/genética , Proteínas Smad/metabolismo
18.
Environ Toxicol Pharmacol ; 68: 109-119, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30884453

RESUMO

Di(2-ethylhexyl) phthalate (DEHP) is an environmental pollutant that is widely used in medical and consumer products. An epidemiological study has suggested that a large daily intake of DEHP from phthalate-contaminated food may be a risk factor for liver dysfunction. Long-term exposure to DEHP is associated with liver disease and exacerbates the progression of chronic liver injury. However, the effect of DEHP on hepatic fibrosis is rarely studied. In the present study, we sought to determine the effect of DEHP on carbon tetrachloride (CCl4)-induced liver fibrosis, and to further examine the molecular mechanisms. We found that DEHP exposure remarkably promoted liver inflammation, necrosis and fibrosis, and increased expression of the protein associated with liver inflammation and fibrogenesis, including α-SMA, COL-Ⅰ, COL-Ⅲ, TGF-ß1, P-Smad2, P-Smad3, P-p38 and P-p65. The similar trend was observed in the LX-2 cells. Furthermore, DEHP exposure induced oxidative stress and inflammatory cytokine production. Taken together, DEHP might play a fibrotic role in hepatic fibrosis rats and TGF-ß1-stimulated LX-2 cells in vitro which was related to TGF-ß1/Smad and p38MAPK/NF-κB signal pathway.


Assuntos
Dietilexilftalato/toxicidade , Cirrose Hepática/induzido quimicamente , Plastificantes/toxicidade , Animais , Inflamação/induzido quimicamente , Inflamação/metabolismo , Inflamação/patologia , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
19.
J Cancer Res Ther ; 15(1): 192-203, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30880778

RESUMO

Background: Promoter methylation of tumor suppressor genes (TSGs) is a well-reported portent in carcinogenesis; hence, it is worthy to investigate this in high-risk Northeast population of India. The study was designed to investigate methylation status of 94 TSGs in esophageal squamous cell carcinoma (ESCC). Further, the effect of OPCML promoter methylation on gene expression was analyzed by immunohistochemistry. Moreover, in silico protein-protein interactions were examined among 8 TSGs identified in the present study and 23 epigenetically regulated genes reported previously by our group in ESCC. Materials and Methods: Methylation profiling was carried out by polymerase chain reaction array and OPCML protein expression was examined by tissue microarray-based immunohistochemistry. Results: OPCML, NEUROG1, TERT, and WT1 genes were found hypermethylated and SCGB3A1, CDH1, THBS1, and VEGFA were hypomethylated in Grade 2 tumor. No significant change in OPCML expression was observed among control, Grade 1, and Grade 2 tumor. Conclusively, hypermethylation of the studied OPCML promoter in Grade 2 tumor produced no effect on expression. Unexpectedly, OPCML expression was downregulated in Grade 3 tumor in comparison to other groups signifying that downregulation of OPCML expression may lead to higher grade of tumor formation at the time of diagnosis of ESCC in patients. Significant interactions at protein level were found as VEGFA:PTK2, CTNNB1:CDH1, CTNNB1:VEGFA, CTNNB1:NEUROG1, CTNND2:CDH1, and CTNNB1:TERT. These interactions are pertinent to Wnt/ß-catenin and TGF-ß-Smad pathways. Conclusions: Deranged OPCML expression may lead to high-grade ESCC as well as epigenetically regulated genes, that is, CDH1, CTNNB1, CTNND2, THBS1, PTK2, WT1, OPCML, TGFB1, and SMAD4 may alter the Wnt/ß-catenin and TGF-ß-Smad pathways in ESCC. Further study of these genes could be useful to understand the molecular pathology of ESCC with respect to epithelial-mesenchymal transition (EMT) mediated by Wnt/ß-catenin and TGF-ß signaling pathways.


Assuntos
Moléculas de Adesão Celular/genética , Neoplasias Esofágicas/genética , Carcinoma de Células Escamosas do Esôfago/genética , Regulação Neoplásica da Expressão Gênica , Moléculas de Adesão Celular/metabolismo , Metilação de DNA/genética , Regulação para Baixo , Epigênese Genética/genética , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/patologia , Esôfago/patologia , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/metabolismo , Humanos , Masculino , Gradação de Tumores , Regiões Promotoras Genéticas/genética , Transdução de Sinais/genética , Proteínas Smad/metabolismo , Análise Serial de Tecidos , Fator de Crescimento Transformador beta/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Proteínas Wnt/metabolismo , beta Catenina/metabolismo
20.
Molecules ; 24(6)2019 Mar 21.
Artigo em Inglês | MEDLINE | ID: mdl-30901941

RESUMO

Metastasis is a major cause of death in patients with breast cancer. In the process of cancer development, epithelial-mesenchymal transition (EMT) is crucial to promoting the invasion and migration of tumor cells. In a previous study, the role of resveratrol in migration and metastasis was investigated in MDA-MB-231 (MDA231) human breast cancer cells and a xenograft-bearing mouse model. Additionally, the related mechanism was explored. In the present study, in vitro Transwell assays showed that resveratrol can inhibit the migration of transforming growth factor (TGF)-ß1-induced MDA231 cells in a concentration-dependent manner. An enzyme-linked immunosorbent assay (ELISA) showed that resveratrol can reduce the secretion of matrix metalloproteinase (MMP)-2 and MMP-9. Immunofluorescence was performed to confirm the expression of EMT-related markers. Immunofluorescence assays confirmed that resveratrol changed the expression of the EMT-related markers E-cadherin and vimentin. Western blot analysis demonstrated that resveratrol decreased the expression levels of MMP-2, MMP-9, Fibronectin, α-SMA, P-PI3K, P-AKT, Smad2, Smad3, P-Smad2, P-Smad3, vimentin, Snail1, and Slug, as well as increased the expression levels of E-cadherin in MDA231 cells. In vivo, resveratrol inhibited lung metastasis in a mouse model bearing MDA231 human breast cancer xenografts without marked changes in body weight or liver and kidney function. These results indicate that resveratrol inhibits the migration of MDA231 cells by reversing TGF-ß1-induced EMT and inhibits the lung metastasis of MDA231 human breast cancer in a xenograft-bearing mouse model.


Assuntos
Neoplasias da Mama/metabolismo , Movimento Celular/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Resveratrol/farmacologia , Fator de Crescimento Transformador beta1/metabolismo , Animais , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Humanos , Metaloproteinases da Matriz/metabolismo , Camundongos , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta1/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
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