RESUMEN
Objective To investigate the effect of SMAD family member 3(SMAD3) silenced by small interfering RNA (siRNA) on macrophage polarization and transforming growth factor β1 (TGF-β1)/ SMAD family signaling pathway in rheumatoid arthritis (RA). Methods RA macrophages co-cultured with rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS) were used as a cell model. TGF-β1 was used to stimulate macrophages, and SMAD3-specific siRNA (si-SMAD3) and negative control siRNA (si-NC) were transfected into human RA macrophages co-cultured in TranswellTM chamber. The expression of SMAD3 mRNA was detected by real-time fluorescence quantitative PCR, and the expression of TGF-β1, SMAD3 and SMAD7 protein was detected by Western blot analysis. The contents of TGF-β1 and IL-23 in cell culture supernatant were determined by ELISA. Cell proliferation was detected by CCK-8 assay. TranswellTM chamber was used to measure cell migration. Results Compared with the model group and the si-NC group, the expression of TGF-β1, SMAD3 mRNA and protein in RA macrophages decreased significantly after silencing SMAD3. In addition, the secretion of IL-23 decreased significantly, and the cell proliferation activity and cell migration were inhibited, with high expression of SMAD7. Conclusion Knockdown of SMAD3 can promote M2 polarization and SMAD7 expression in RA macrophages.
Asunto(s)
Humanos , Artritis Reumatoide/genética , Interleucina-23 , Macrófagos , ARN Mensajero , ARN Interferente Pequeño/genética , Proteína smad7/genética , Factor de Crecimiento Transformador beta1/genética , Proteína smad3/genética , Silenciador del GenRESUMEN
MicroRNAs (miRNAs) have recently been recognized to have a role in human orthopedic disorders. The objective of our study was to explore the expression profile and biological function of miRNA-17-5p (miR-17-5p), which is well known to be related to cancer cell proliferation and invasion, in osteoblastic differentiation and in cell proliferation. The expression levels of miR-17-5p in the femoral head mesenchymal stem cells of 20 patients with non-traumatic osteonecrosis (ON) and 10 patients with osteoarthritis (OA) were examined by quantitative reverse transcription-PCR (qRT-PCR). Furthermore, the interaction between miR-17-5p and SMAD7 was observed. We found that in non-traumatic ON samples the level of mature miR-17-5p was significantly lower than that of OA samples (P=0.0002). By targeting SMAD7, miR-17-5p promoted nuclear translocation of beta-catenin, enhanced expression of COL1A1 and finally facilitated the proliferation and differentiation of HMSC-bm cells. We also demonstrated that restoring expression of SMAD7 in HMSC-bm cells partially reversed the function of miR-17-5p. Together, our data suggested a theory that dysfunction of a network containing miR-17-5p, SMAD7 and beta-catenin could contribute to ON pathogenesis. The present study prompts the potential clinical value of miR-17-5p in non-traumatic ON.
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Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Secuencia de Bases , Proteína Morfogenética Ósea 2/metabolismo , Diferenciación Celular , Línea Celular , Proliferación Celular , Regulación de la Expresión Génica , MicroARNs/genética , Osteoartritis/genética , Osteoblastos/citología , Osteogénesis , Osteonecrosis/genética , Transducción de Señal , Proteína smad7/genética , beta Catenina/metabolismoRESUMEN
One of the major signaling pathways that determine the tumor aggression and patient outcome in pancreatic cancer is the transforming growth factor-beta (TGF-ß) pathway. It is inactivated at various levels in pancreatic cancer and plays a dual role in tumor initiation and progression. The Smad family of proteins transduce signals from the TGF-ß superfamily ligands that regulate cell proliferation, differentiation and death through activation of receptor serine/threonine kinases. This review discusses the structure, function and regulation of various participating Smad family members, and their individual roles in determining the progression and outcome of pancreatic cancer patients, with a special emphasis on Smad4.
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Diferenciación Celular , Proliferación Celular , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Humanos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Fosforilación , Receptores de Factores de Crecimiento Transformadores beta/genética , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Transducción de Señal , Proteína Smad4/química , Proteína Smad4/genética , Proteína Smad4/metabolismo , Proteína smad6/genética , Proteína smad6/metabolismo , Proteína smad7/genética , Proteína smad7/metabolismo , Factor de Crecimiento Transformador beta/genética , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
Background: Smad4, Smad6 and Smad7 are important molecules in TGF-beta pathway, which plays an important role in pancreatic ductal adenocarcinoma (PDAC) biology. Aims : This study examined the expression profiles of Smad4, Smad6 and Smad7 mRNA in patient samples of PDAC and their relationship to Smad protein expression, SMAD4 gene mutations, clinicopathological parameters and patient survival. Settings and Design: Surgically resected, paired normal and tumor tissues of 25 patients of PDAC were studied. Materials and Methods: Protein and mRNA levels were assessed by immunohistochemistry and RT-PCR, respectively. Statistical Methods: Statistical analysis was done using Student's t-test, Pearson's chi-square test, Spearman's Rank Correlation, Pearson's Correlation test and Kaplan-Meier Logrank test. Results: While there was a highly significant difference in the protein levels of all three Smads in tumor as compared to normal samples, mRNA levels were significantly different only for Smad4. Protein levels did not correlate significantly with mRNA levels for any of the three Smads. The mRNA levels of Smad4 and Smad6, Smad4 and Smad7, and Smad6 and Smad7 in tumor samples showed a significant positive correlation. The relationship of Smad4 mRNA expression to SMAD4 gene status and Smad4 protein expression was discordant and there was no significant correlation between mRNA expression and clinicopathological parameters and patient survival. Conclusion : The absence of concordance between SMAD4 gene status, mRNA expression and Smad4 protein expression suggests the presence of other regulatory mechanisms in Smad4 transcription and translation in PDAC.
Asunto(s)
Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/secundario , Adulto , Anciano , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/secundario , Femenino , Humanos , Técnicas para Inmunoenzimas , Masculino , Persona de Mediana Edad , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Pronóstico , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteína Smad4/genética , Proteína Smad4/metabolismo , Proteína smad6/genética , Proteína smad6/metabolismo , Proteína smad7/genética , Proteína smad7/metabolismo , Tasa de SupervivenciaRESUMEN
Idiopathic pulmonary fibrosis (IPF) is a lethal parenchymal lung disease characterized by myofibroblast proliferation. Alveolar epithelial cells (AECs) are thought to produce myofibroblasts through the epithelial to mesenchymal transition (EMT). Receptor for advanced glycation end products (RAGE) is a member of the immunoglobulin superfamily of cell surface receptors whose activation is associated with renal fibrosis during diabetes and liver fibrosis. RAGE is expressed at low basal levels in most adult tissues except the lung. In this study, we evaluated the interaction of ligand advanced glycation end products (AGE) with RAGE during the epithelial to myofibroblast transition in rat AECs. Our results indicate that AGE inhibited the TGF-beta-dependent alveolar EMT by increasing Smad7 expression, and that the effect was abolished by RAGE siRNA treatment. Thus, the induction of Smad7 by the AGE-RAGE interaction limits the development of pulmonary fibrosis by inhibiting TGF-beta-dependent signaling in AECs.