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1.
Biochimie ; 162: 229-238, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30954547

RESUMO

Lipopolysaccharide (LPS) as a component of the outer structure of cell wall of gram-negative bacteria, could induce apoptosis in the intestinal endocrine cell line STC-1. However, the signaling cascades involved in this process have not been elucidated. Hence, we investigated the mechanism of cell apoptosis and hyposecretion of glucagon-like peptide 1 (GLP-1) induced by LPS in the GLUTag enteroendocrine cell line. LPS decreased the cell viability of GLUTag cells, up-regulated the TNF-α level, induced the apoptosis and down-regulated the mRNA and protein levels of GLP-1. In addition, TNF-α promoted LPS-induced apoptosis of GLUTag cells through mediating the formation of the RIP1/RIP3 necrosome. RIP1 and RIP3 knockdown increased cell viability, the mRNA and protein levels of GLP-1 and the mTOR signaling pathway-related proteins (p-mTOR and p-S6), and decreased the relative caspase 3/7 activity, cell apoptosis and ROS production. Further studies showed that ROS inhibited the mTOR signaling pathway. Moreover, the antioxidant N-acetyl-l-cysteine increased cell viability, GLP-1 expressions and the mTOR signaling pathway-related proteins, and inhibited the ROS production. However, the mTOR specific inhibitor (Rapa) reversed all these above effects. Taken together, our result revealed that LPS induced the apoptosis of GLUTag cells and GLP-1 hyposecretion through the RIP/ROS/mTOR pathway.

2.
Oncol Rep ; 40(5): 2608-2618, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30226614

RESUMO

The purpose of the present study was to identify the potential function of Kruppel-like factor 5 (KLF5) in thyroid cancer and investigate the underlying mechanisms. The protein levels of KLF5 in 98 thyroid cancer tissues were analyzed using an immunohistochemistry assay. SW579 cells transfected with small interfering RNA against KLF5 and B-CPAP cells transfected with KLF5 expressing vectors were used for functional studies. Western blot analysis, immunofluorescence and co-immunoprecipitation assays were used to investigate the mechanisms of KLF5. In vivo tumorigenicity was assessed using a subcutaneous xenograft experiment. The results revealed that KLF5 was highly expressed in thyroid cancer tissues and associated with lymph node metastasis. Knockdown of KLF5 in SW579 cells suppressed proliferation, anchorage-independent growth, migration and invasion in vitro, while the overexpression of KLF5 resulted in opposite effects in B-CPAP cells. Mechanistically, it was demonstrated that KLF5 promoted the cytoplasm-nuclear translocation of nuclear factor-κB. Additionally, it was revealed that insufficient F-box/WD repeat-containing protein 7 expression may be responsible for the dysfunction of KLF5 in thyroid cancer. These results revealed that KLF5 promotes the tumorigenesis and metastasis of thyroid cancer cells and may be a potential therapeutic target in patients with thyroid cancer.

3.
Biochem Biophys Res Commun ; 478(2): 612-7, 2016 09 16.
Artigo em Inglês | MEDLINE | ID: mdl-27475499

RESUMO

Podocyte apoptosis induced by high levels of glucose is a key event in the development and prognosis of diabetic nephropathy (DN). Forkhead transcription factor O1 (FoxO1) has been defined as a critical mediator of oxidative stress in animal models of diabetes and is involved in mitophagy. To test the role of FoxO1 in regulating podocyte apoptosis both in vivo and in vitro, we generated FoxO1 overexpression models. High-glucose (HG) induced podocyte apoptosis with decreased mitophagy. These changes were accompanied by mitochondrial dysfunction and more severe podocyte loss in mouse kidney. FoxO1 overexpression prevented the apoptosis induced by HG. Reduction of cell apoptosis and renal damage depended upon the expression of PTEN-induced putative kinase 1 (PINK1). These findings suggest that specific overexpression of renal FoxO1 decreases podocyte apoptosis, which may be explained in part by its regulation of PINK1, and that targeting FoxO1 may represent a novel therapeutic approach for DN.


Assuntos
Apoptose , Nefropatias Diabéticas/genética , Proteína Forkhead Box O1/genética , Glucose/metabolismo , Glomérulos Renais/patologia , Podócitos/patologia , Regulação para Cima , Animais , Linhagem Celular , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/patologia , Proteína Forkhead Box O1/metabolismo , Rim/metabolismo , Rim/patologia , Glomérulos Renais/citologia , Glomérulos Renais/metabolismo , Masculino , Camundongos , Podócitos/citologia , Podócitos/metabolismo , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , RNA Mensageiro/genética , Transdução de Sinais
4.
Biochem Biophys Res Commun ; 471(4): 416-22, 2016 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-26902117

RESUMO

Accumulating evidence has suggested that the epithelial-mesenchymal transition (EMT) is a pathway that potentially leads to podocyte depletion and proteinuria in diabetic nephropathy (DN). Therefore, this study was designed to investigate the protective effects of forkhead transcription factor O1 (FOXO1) on podocyte EMT, under high-glucose (HG) conditions in vitro and under diabetic conditions in vivo. The results showed that HG-induced podocyte EMT was associated with FOXO1 inactivation, which was accompanied by activation of the transforming growth factor (TGF)-ß1/SMAD3/integrin-linked kinase (ILK) pathway. Accordingly, constitutive FOXO1 activation suppressed the TGF-ß1/Smad3/ILK pathway and partially reversed EMT, similar to the effects observed after treatment with SIS3 or QLT0267, which are selective inhibitors of TGF-ß1-dependent SMAD3 phosphorylation and ILK, respectively. In addition, lentiviral-mediated FOXO1 overexpression in the kidneys of diabetic mice considerably increased FOXO1 expression and activation, while decreasing proteinuria and renal pathological injury. These data suggested that forced FOXO1 activation inhibited HG-induced podocyte EMT and ameliorated proteinuria and renal injury in diabetic mice. Our findings further highlighted that FOXO1 played a protective role against diabetes in mice and may potentially be used as a novel therapeutic target for treating diabetic nephropathy.


Assuntos
Transição Epitelial-Mesenquimal/fisiologia , Fatores de Transcrição Forkhead/metabolismo , Glucose/metabolismo , Podócitos/patologia , Animais , Compostos Azo/administração & dosagem , Desmina/metabolismo , Diabetes Mellitus Experimental/metabolismo , Nefropatias Diabéticas/patologia , Proteína Forkhead Box O1 , Fatores de Transcrição Forkhead/genética , Regulação da Expressão Gênica , Isoquinolinas/farmacologia , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Podócitos/efeitos dos fármacos , Podócitos/fisiologia , Proteínas Serina-Treonina Quinases/metabolismo , Pirazóis/administração & dosagem , Piridinas/farmacologia , Pirróis/farmacologia , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/metabolismo
5.
J Exp Clin Cancer Res ; 34: 146, 2015 Dec 04.
Artigo em Inglês | MEDLINE | ID: mdl-26637328

RESUMO

BACKGROUND: The incidence of thyroid cancer has progressively increased over the past few decades, and the most frequent types of this cancer are papillary thyroid carcinoma (PTC) and small primary tumors. In PTC, oncogene activation is known to occur at a high frequency. However, the potential roles of tumor suppressor genes in thyroid carcinogenesis remain unclear. LDOC1 was first identified as a gene encoding a leucine zipper protein whose expression was decreased in a series of pancreatic and gastric cancer cell lines. In this study, we aimed to determine the status of LDOC1 in PTC and identify its mechanistic role in PTC pathogenesis. METHODS: LDOC1 expression was evaluated in fresh samples and stored specimens of human PTC and contralateral normal tissues by performing quantitative reverse transcription-PCR and immunohistochemical staining. The correlation to nuclear p65 content in the stored specimens was analyzed. Moreover, the basal level of LDOC1 in two human PTC-derived cell lines (BCPAP and TPC-1) compared with normal thyroid tissue was determined. Human LDOC1 cDNA was inserted into a lentiviral vector and transduced into TPC-1 cells. TPC-1 cells overexpressing LDOC1/GFP (Lv-LDOC1) or negative control GFP (Lv-NC) were stimulated with TNFα or recombinant TGF-ß1, and then cell proliferation, cell cycle distribution, and apoptosis were assessed. Western blotting was used to examine the expression of p65, IκBα, c-Myc, Bax, and Bcl-xL, and a luciferase reporter assay was used to measure NF-κB activity stimulated by TNFα. Statistical significance was determined using Student's t tests or ANOVA and Newman-Keuls multiple comparison tests. Pearson chi-square test was used to analyze possible associations. RESULTS: LDOC1 expression was significantly downregulated in PTC specimens as compared with the expression in normal thyroid tissues, and this downregulation was associated with an increase in tumor size (P < 0.05). There is a correlation between LDOC1 and nuclear P65 expression in human PTC tissues (P < 0.01). Lentivirus-mediated restoration of LDOC1 expression in TPC-1 cells characterized by low level of LDOC1 expression suppressed proliferation and induced apoptosis by inhibiting NF-κB activation, and LDOC1-overexpressing TPC-1 cells recovered responsiveness to TGF-ß1 antiproliferative signaling. CONCLUSIONS: LDOC1 might function as a tumor suppressor gene in PTC by inhibiting NF-κΒ signaling, and thus might represent a promising therapeutic target in patients with PTC.


Assuntos
Apoptose/genética , Carcinoma/genética , Carcinoma/metabolismo , NF-kappa B/metabolismo , Proteínas Nucleares/genética , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/metabolismo , Proteínas Supressoras de Tumor/genética , Carcinoma/patologia , Carcinoma Papilar , Ciclo Celular/genética , Linhagem Celular Tumoral , Núcleo Celular , Proliferação de Células/genética , Ativação Enzimática/genética , Expressão Gênica , Humanos , Transporte Proteico , Transdução de Sinais , Câncer Papilífero da Tireoide , Neoplasias da Glândula Tireoide/patologia , Fator de Transcrição RelA/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Carga Tumoral
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