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2.
Zhonghua Shao Shang Za Zhi ; 36(3): 224-233, 2020 Mar 20.
Artigo em Chinês | MEDLINE | ID: mdl-32241049

RESUMO

Objective: To explore the transcriptional regulation mechanism of transforming growth factor ß(1) (TGF-ß(1)) on Meox1 and its effect on cell migration of adult human dermal fibroblasts (HDF-a). Methods: (1) HDF-a cells were cultured in RPMI 1640 complete medium (hereinafter referred to as routinely cultured). The cells were divided into TGF-ß(1) stimulation group and blank control group. The cells in TGF-ß(1) stimulation group were stimulated with 10 µL TGF-ß(1) in the mass concentration of 1 mg/µL, while the cells in blank control group were stimulated with the equal volume of phosphate buffer solution. After 72 hours in culture, partial cells in both groups were collected for transcriptome sequencing. The genes with differential expression ratio greater than or equal to 2 and P<0.01 between the two groups were selected to perform enrichment analysis and analysis of metabolic pathways of the Kyoto Gene and Genome Encyclopedia with, and the expression value of Meox1 per million transcripts (TPM) was recorded (n=3). Partial cells from the two groups were used to detect the Meox1 mRNA expression by real-time fluorescent quantitative reverse transcription polymerase chain reaction (RT-PCR) (n=3). (2) Cultured HDF-a cells in the logarithmic growth phase (the same growth phase of cells below) were divided into empty plasmid group, Smad2 overexpression (OE) group, Smad3 OE group, and Smad4 OE group, which were transfected respectively with 2 µg empty pcDNA3.1 plasmid and pcDNA3.1 plasmids separately carrying Smad2, Smad3, and Smad4 for 6 hours, and then were routinely cultured for 48 hours. The Meox1 mRNA expression in the transfected cells of each group was detected by real-time fluorescent quantitative RT-PCR (n=3). (3) HDF-a cells were routinely cultured and grouped the same as in experiment (1). After 72 hours in culture, the enrichment of Smad2, Smad3, and Smad4 protein on the Meox1 promoter in the cells of each group was detected by chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) (n=3). (4) HDF-a cells were routinely cultured and divided into negative interference group, small interference RNA (siRNA)-Smad2 group, siRNA-Smad3 group, siRNA-Smad4 group, empty plasmid group, Smad2 OE group, Smad3 OE group, and Smad4 OE group, which were transfected respectively with 50 µmol/L random siRNA, siRNA-Smad2, siRNA-Smad3, siRNA-Smad4, 2 µg empty pcDNA3.1 plasmid and pcDNA3.1 plasmids separately carrying Smad2, Smad3, and Smad4 for 6 hours and then routinely cultured for 48 hours. The enrichment of Smad2, Smad3, and Smad4 protein on the Meox1 promoter in the cells of corresponding group was detected by ChIP-qPCR (n=3). (5) Two batches of HDF-a cells were cultured and divided into negative interference group, siRNA-Meox1 group, empty plasmid group, and Meox1 OE group, which were transfected respectively with 50 µmol/L random siRNA, siRNA-Meox1, 2 µg empty pcDNA3.1 plasmid and pcDNA3.1 plasmid carrying Meox1 for 6 hours and then routinely cultured for 24 hours. One batch of cells were subjected to scratch test with the scratch width being observed 24 hours after scratching and compared with the initial width for scratch wound healing; the other batch of cells were subjected to Transwell assay, in which the migrated cells were counted after being routinely cultured for 24 hours (n=3). (6) From January 2018 to June 2019, 3 hypertrophic scar patients (2 males and 1 female, aged 35-56 years) were admitted to the First Affiliated Hospital of Army Medical University (the Third Military Medical University) 8-12 months after burns. The scar tissue and normal skin tissue along the scar margin resected during surgery were taken, and immunohistochemical staining was performed to observe the distribution of Meox1 protein expression. Data were statistically analyzed with one-way analysis of variance and independent sample t test. Results: (1) After 72 hours in culture, a total of 843 genes were obviously differentially expressed between the two groups, being related to tissue repair, cell migration, inflammatory cell chemotaxis induction process and potential signaling pathways such as tumor necrosis factor, interleukin 17, extracellular matrix receptor. The TPM value of Meox1 in the cells of blank control group was 45.9±1.9, which was significantly lower than 163.1±29.5 of TGF-ß(1) stimulation group (t=6.88, P<0.01) with RNA-sequencing. After 72 hours in culture, the Meox1 mRNA expression levels in the cells of blank control group was 1.00±0.21, which was significantly lower than 11.00±3.61 of TGF-ß(1) stimulation group (t=4.79, P<0.01). (2) After 48 hours in culture, the Meox1 mRNA expression levels in the cells of Smad2 OE group, Smad3 OE group, and Smad4 OE group were 198.70±11.02, 35.47±4.30, 20.27±2.50, respectively, which were significantly higher than 1.03±0.19 of empty plasmid group (t=31.07, 13.80, 13.12, P<0.01). (3) After 72 hours in culture, the enrichment of Smad2, Smad3, and Smad4 protein on the promoter of Meox1 in the cells of TGF-ß(1) stimulation group was significantly higher than that of blank control group respectively (t=12.99, 41.47, 29.10, P<0.01). (4) After 48 hours in culture, the enrichment of Smad2 protein on the promoter of Meox1 in the cells of negative interference group was (0.200 000±0.030 000)%, significantly higher than (0.000 770±0.000 013)% of siRNA-Smad2 group (t=11.67, P<0.01); the enrichment of Smad2 protein on the promoter of Meox1 in the cells of empty plasmid group was (0.200 000±0.040 000)%, significantly lower than (0.700 000±0.090 000)% of Smad2 OE group (t=8.85, P<0.01). The enrichment of Smad3 protein on the promoter of Meox1 in the cells of negative interference group was (0.500 0±0.041 3)%, significantly higher than (0.006 0±0.001 3)% of siRNA-Smad3 group (t=17.79, P<0.01); the enrichment of Smad3 protein on the promoter of Meox1 in the cells of empty plasmid group was (0.470 0±0.080 0)%, which was significantly lower than (1.100 0±0.070 0)% of Smad3 OE group (t=9.93, P<0.01). The enrichment of Smad4 protein on the promoter of Meox1 in the cells of negative interference group was similar to that of siRNA-Smad4 group (t=2.11, P>0.05); the enrichment of Smad4 protein on the promoter of Meox1 in the cells of empty plasmid group was similar to that of Smad4 OE group (t=0.60, P>0.05). (5) Twenty-four hours after scratching, the scratch healing width of cells in siRNA-Meox1 group was narrower than that of negative interference group, while that of Meox1 OE group was wider than that of empty plasmid group. After 24 hours in culture, the number of migration cells in negative interference group was significantly higher than that in siRNA-Meox1 group (t=9.12, P<0.01), and that in empty plasmid group was significantly lower than that in Meox1 OE group (t=8.99, P<0.01). (6) The expression of Meox1 protein in the scar tissue was significantly higher than that in normal skin of patients with hypertrophic scars. Conclusions: TGF-ß(1) transcriptionally regulates Meox1 expression via Smad2/3 in HDF-a cells, thus promoting cell migration.


Assuntos
Movimento Celular , Cicatriz Hipertrófica , Fibroblastos/metabolismo , Proteínas de Homeodomínio , Fatores de Transcrição , Fator de Crescimento Transformador beta1/metabolismo , Adulto , Regulação da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Transdução de Sinais , Fator de Crescimento Transformador beta
3.
Anticancer Res ; 40(4): 1915-1920, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32234880

RESUMO

BACKGROUND/AIM: New anticancer drugs are usually tested on cancer cells in culture in a standard medium. We stimulated immune polynuclear cells by lipopolysaccharides to obtain an enriched medium (EM) containing inflammatory cytokines more closely reflecting the tumor microenvironment and tested a rhenium-diselenium (Re-diSe) drug in this new model. Concentrations of cytokines were compared with a control medium (CM). MATERIALS AND METHODS: Human-derived breast cancer cells were grown in culture either in CM or EM with or without Re-diSe. Assays of tumor necrosis factor alpha (TNFα), interleukin 6 (IL6), intereukin 1 beta (IL1ß), transforming growth factor-beta (TGFß), insulin growth factor 1 (IGF1) and vascular epidermal growth factor A (VEGFA) were performed by enzyme-linked immunosorbent assays. The production of reactive oxygen species (ROS) was determined by 2,7-dichlorofluorescein test. The cell growth was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide tests. RESULTS: Concentrations of TNFα, IL6 and Il1ß were observed to be significantly higher in EM than in CM. There was no difference for TGFß, IGF1 and VEGFA. The cells were sensitive to Re-diSe, with reduced concentrations of TGFß, IGF1, VEGFA and ROS, but the half-maximal inhibitory concentration was significantly higher in EM than in CM. CONCLUSION: The efficacy of the Re-diSe drug was confirmed in this model of aggressive cancer.


Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Rênio/farmacologia , Selênio/farmacologia , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proliferação de Células/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Fator de Crescimento Insulin-Like I/genética , Interleucina-1beta/genética , Interleucina-6/genética , Lipopolissacarídeos/farmacologia , Cultura Primária de Células , Espécies Reativas de Oxigênio , Fator de Crescimento Transformador beta/genética , Microambiente Tumoral/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/genética
4.
Med Sci Monit ; 26: e920943, 2020 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-32248202

RESUMO

BACKGROUND Bone morphogenetic proteins (BMPs) are widely involved in cancer development. However, a wealth of conflicting data raises the question of whether BMPs serve as oncogenes or as cancer suppressors. MATERIAL AND METHODS By integrating multi-omics data across cancers, we comprehensively analyzed the genomic and pharmacogenomic landscape of BMP genes across cancers. RESULTS Surprisingly, our data indicate that BMPs are globally downregulated in cancers. Further genetics and epigenetics analyses show that this abnormal expression is driven by copy number variations, especially heterozygous amplification. We next assessed the BMP-associated pathways and demonstrated that they suppress cell cycle and estrogen hormone pathways. Bone morphogenetic protein interacts with 58 compounds, and their dysfunction can induce drug sensitivity. CONCLUSIONS Our results define the landscape of the BMP family at a systems level and open potential therapeutic opportunities for cancer patients.


Assuntos
Proteínas Morfogenéticas Ósseas , Neoplasias/metabolismo , Animais , Proteínas Morfogenéticas Ósseas/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Proteínas Morfogenéticas Ósseas/fisiologia , Variações do Número de Cópias de DNA , Humanos , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta/fisiologia
5.
Basic Res Cardiol ; 115(3): 27, 2020 03 07.
Artigo em Inglês | MEDLINE | ID: mdl-32146539

RESUMO

Heart failure is a major health problem worldwide with a significant morbidity and mortality rate. Although studied extensively in animal models, data from patients at the compensated disease stage are lacking. We sampled myocardium biopsies from aortic stenosis patients with compensated hypertrophy and moderate heart failure and used transcriptomics to study the transition to failure. Sequencing and comparative analysis of analogous samples of mice with transverse aortic constriction identified 25 candidate genes with similar regulation in response to pressure overload, reflecting highly conserved molecular processes. The gene cysteine-rich secretory protein LCCL domain containing 1 (CRISPLD1) is upregulated in the transition to failure in human and mouse and its function is unknown. Homology to ion channel regulatory toxins suggests a role in Ca2+ cycling. CRISPR/Cas9-mediated loss-of-function leads to dysregulated Ca2+ handling in human-induced pluripotent stem cell-derived cardiomyocytes. The downregulation of prohypertrophic, proapoptotic and Ca2+-signaling pathways upon CRISPLD1-KO and its upregulation in the transition to failure implicates a contribution to adverse remodeling. These findings provide new pathophysiological data on Ca2+ regulation in the transition to failure and novel candidate genes with promising potential for therapeutic interventions.


Assuntos
Sinalização do Cálcio , Cálcio/metabolismo , Moléculas de Adesão Celular/metabolismo , Evolução Molecular , Insuficiência Cardíaca/metabolismo , Sequência de Aminoácidos , Animais , Estenose da Valva Aórtica/complicações , Estenose da Valva Aórtica/genética , Estenose da Valva Aórtica/metabolismo , Apoptose , Biópsia , Moléculas de Adesão Celular/química , Moléculas de Adesão Celular/deficiência , Moléculas de Adesão Celular/genética , Sequência Conservada , Regulação para Baixo , Feminino , Insuficiência Cardíaca/complicações , Insuficiência Cardíaca/genética , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Masculino , Camundongos , Miocárdio/metabolismo , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Transcriptoma , Fator de Crescimento Transformador beta/metabolismo
6.
Sichuan Da Xue Xue Bao Yi Xue Ban ; 51(2): 165-170, 2020 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-32220183

RESUMO

Objective: To investigate the regulation of fibromodulin (FMOD) on proliferation, adhesion and migration of non-small cell lung cancer cell line H322, and discuss its action mechanism. Methods: H322 cells were randomly divided into control group, small interfering RNA (siRNA) silencing FMOD ( FMOD siRNA) group and control siRNA (Con siRNA) group. FMOD siRNA and Con siRNA were transfected into H322 cells. The cell viability of each group was detected by CCK-8 method. The adhesion ability of cells was detected by fluorescein diacetate (FDA) fluorescent staining. The cell migration ability was detected by Transwell method. Real time-PCR was used to detect the mRNA expressions of Cyclin D1, intercellular adhesion molecule -1 (ICAM-1), E-cadherin, FMOD, transforming growth factor-ß (TGF-ß), Smad2, Smad3, Smad4 and Smad7 in cells. The protein expressions of Cyclin D1, ICAM-1, E-cadherin, FMOD, TGF-ß1, Smad2, Smad3, Smad4 and Smad7 were detected by Western blot. Results: Compared with the Con siRNA group, the cell viability, cell adhesion and migration ability of the FMOD siRNA group were decreased, and the difference was statistically significant ( P<0.01). There was no significant difference between the control group and the Con siRNA group. Real time-PCR and Western blot results showed that the mRNA and protein expression levels of Cyclin D1, ICAM-1, TGF-ß1, Smad2, Smad3 and Smad4 were decreased in FMOD siRNA group, compared with Con siRNA group, while the mRNA and protein expression levels of E-cadherin and Smad7 are elevated. Conclusion: Silencing of the FMOD gene significantly reduces the proliferation, adhesion and migration of H322 cells, which may be conducted by inhibiting the TGF-ß/Smad signaling pathway.


Assuntos
Carcinoma Pulmonar de Células não Pequenas , Fibromodulina/genética , Inativação Gênica , Neoplasias Pulmonares , Proteínas Smad , Fator de Crescimento Transformador beta , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Adesão Celular , Movimento Celular , Proliferação de Células , Fibromodulina/fisiologia , Expressão Gênica , Humanos , Neoplasias Pulmonares/metabolismo , RNA Interferente Pequeno , Transdução de Sinais , Proteínas Smad/fisiologia , Fator de Crescimento Transformador beta/fisiologia
8.
Life Sci ; 250: 117552, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32179074

RESUMO

AIMS: This study aimed to explore the possible mechanism of trauma-induced laryngotracheal stenosis and potential protective and therapeutic efficacy of quercetin on trauma-induced laryngotracheal stenosis. MAIN METHODS: The expression and activity of fibrotic factors [interleukin (IL)-6, IL-8, autophagy related 5 (ATG5), collagen (COL)-1, tumor growth factor (TGF)-ß COL-3, microtubule-associated proteins 1A/1B light chain 3A (LC3), and vascular endothelial growth factor (VEGF)] and fibrotic signaling mediators [mammalian target of rapamycin (mTOR) and phosphorylated AKT (pAKT)] were detected by real-time quantitative PCR (qRT-PCR), ELISA, Western blot, and immunohistochemical staining, respectively, in the lipopolysaccharide (LPS)-induced WI-38 (a human embryonic lung fibroblast cell line) cellular fibrotic model and a trauma-induced rabbit tracheal stenosis model, with and without quercetin treatment. KEY FINDINGS: Pre-treatment with quercetin significantly reversed the LPS-induced upregulation of pro-fibrotic factors (IL-6, IL-8, COL-1, COL-3, LC3) and fibrotic signaling mediators (mTOR and AKT), and it induced the downregulation of ATG5 in the WI-38 cells. Furthermore, the anti-fibrotic activity of quercetin was confirmed in the trauma-induced rabbit tracheal stenosis model. Thus, the nasogastric administration of quercetin attenuated the tracheal stenosis of the rabbit tracheal stenosis model, in addition to effectively reversing an increase in pro-fibrotic factors (VEGF, IL-6, TGF-ß, COL-1, and COL-3) and fibrotic signaling mediators (mTOR and AKT), as well as downregulating ATG5 of the rabbit tracheal stenosis model. SIGNIFICANCE: Quercetin exhibits anti-fibrotic activity by inhibiting pro-fibrotic factors and AKT/mTOR signaling pathway, in addition to activating autophagy activity. This study provided experimental evidence supporting the application of quercetin in tracheal stenosis, clinically.


Assuntos
Proteínas Proto-Oncogênicas c-akt/metabolismo , Quercetina/farmacologia , Serina-Treonina Quinases TOR/metabolismo , Traqueia/efeitos dos fármacos , Fator de Crescimento Transformador beta/metabolismo , Animais , Anti-Inflamatórios/farmacologia , Autofagia , Linhagem Celular , Sobrevivência Celular , Constrição Patológica/tratamento farmacológico , Regulação para Baixo , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Lipopolissacarídeos , Masculino , Coelhos , Transdução de Sinais , Traqueia/patologia
9.
Invest Ophthalmol Vis Sci ; 61(2): 1, 2020 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-32031573

RESUMO

Purpose: This study aimed to explore the role of the protein kinase A (PKA) pathway in proliferative vitreoretinopathy (PVR) and the effect of the PKA inhibitor H89 on experimental PVR. Methods: Epiretinal membranes (ERMs) were acquired from PVR patients and analyzed by frozen-section immunofluorescence. An in vivo model was developed by intravitreal injecting rat eyes with ARPE-19 cells and platelet-rich plasma, and changes in eye structures and vision function were observed. An in vitro epithelial-mesenchymal transition (EMT) cell model was established by stimulating ARPE-19 cells with transforming growth factor (TGF)-ß. Alterations in EMT-related genes and cell function were detected. Mechanistically, PKA activation and activity were explored to assess the relationship between TGF-ß1 stimulation and the PKA pathway. The effect of H89 on the TGF-ß-Smad2/3 pathway was detected. RNA sequencing was used to analyze gene expression profile changes after H89 treatment. Results: PKA was activated in human PVR membranes. In vivo, H89 treatment protected against structural changes in the retina and prevented decreases in electroretinogram b-wave amplitudes. In vitro, H89 treatment inhibited EMT-related gene alterations and partially reversed the functions of the cells. TGF-ß-induced PKA activation was blocked by H89 pretreatment. H89 did not affect the phosphorylation or nuclear translocation of regulatory Smad2/3 but increased the expression of inhibitory Smad6. Conclusions: PKA pathway activation is involved in PVR pathogenesis, and the PKA inhibitor H89 can effectively inhibit PVR, both in vivo and in vitro. Furthermore, the protective effect of H89 is related to an increase in inhibitory Smad6.


Assuntos
Isoquinolinas/antagonistas & inibidores , Sulfonamidas/antagonistas & inibidores , Vitreorretinopatia Proliferativa/tratamento farmacológico , Idoso , Animais , Células Cultivadas , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/metabolismo , Eletrorretinografia , Membrana Epirretiniana/metabolismo , Células Epiteliais/metabolismo , Feminino , Humanos , Isoquinolinas/farmacologia , Sistema de Sinalização das MAP Quinases/fisiologia , Masculino , Pessoa de Meia-Idade , Epitélio Pigmentado da Retina/efeitos dos fármacos , Proteínas Smad/fisiologia , Sulfonamidas/farmacologia , Fator de Crescimento Transformador beta/antagonistas & inibidores
10.
J Cancer Res Clin Oncol ; 146(5): 1205-1215, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32034483

RESUMO

Non-coding RNAs (ncRNAs) are reported to be regulators of signaling pathways that are involved in colorectal cancer (CRC) progression. Aiming at finding ncRNAs (miRNAs) that are differentially expressed in tumor versus normal colorectal tissue samples, online RNA-seq data were analyzed. Of between 18 candidate miRNAs, hsa-miR-29b-1 (miR-29b-1) represented the highest fold change of expression level. Hsa-miR-29b-1 is encoded from the third intron of LOC646329 long ncRNA gene. Surprisingly, two miR-29b sponging sites were predicted within exons of LOC646329 gene. Then, dual luciferase assay supported the interaction of miR-29b-1 with LOC646329-variant D transcript. Also, a direct indication of miR-29b-1 with 3'UTR sequence of SMAD3 gene was verified through dual luciferase assay and RT-qPCR analysis. Furthermore, a reverse pattern of expression was detected between miR-29b-1 and LOC646329-variant D transcript in about 25 pairs of CRC tumor samples, detected by RTqPCR. Consistently, overexpression of LOC646329-variant D transcript was followed by increased SMAD3 and p21 genes expression level and downregulation of CyclinD1 genes in HCT116 cells, detected by RT-qPCR, and western analysis. Also, overexpression of it was followed by increased G1 cell population of HCT-116 cells. All of these data suggested a tumor suppressor effect for LOC646329-variant D in CRC tumor tissue samples, consistent to its reduced expression level at late stages of CRC progression. Data also indicated that LOC646329-variant D exerts its suppression effect on CRC progression through sponging miR-29b, which in turn regulates Wnt and TGFB signaling pathways. This makes LOC646329-variant D transcript as a novel potential therapy target.


Assuntos
Neoplasias Colorretais/metabolismo , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Biologia Computacional , Células HCT116 , Células HEK293 , Células HT29 , Humanos , MicroRNAs/genética , RNA Longo não Codificante/genética , Transdução de Sinais
11.
Cancer Sci ; 111(4): 1241-1253, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32012400

RESUMO

We previously revealed that tumor cell-derived angiopoietin-like protein 2 (ANGPTL2) accelerates the metastatic capacity of tumors in an autocrine/paracrine manner by activating tumor cell motility and invasiveness and the epithelial-mesenchymal transition. However, the effects of ANGPTL2 on cancer cell glycolytic metabolism, which is a hallmark of tumor cells, are unknown. Here we report evidence supporting a role for tumor cell-derived ANGPTL2 in establishing a preference for glycolytic metabolism. We report that a highly metastatic lung cancer cell subline expressing abundant ANGPTL2 showed upregulated expression of the glucose transporter GLUT3 as well as enhanced glycolytic metabolism relative to a less metastatic parental line. Most notably, ANGPTL2 overexpression in the less metastatic line activated glycolytic metabolism by increasing GLUT3 expression. Moreover, ANGPTL2 signaling through integrin α5ß1 increased GLUT3 expression by increasing transforming growth factor-ß (TGF-ß) signaling and expression of the downstream transcription factor zinc finger E-box binding homeobox 1 (ZEB1). Conversely, ANGPTL2 knockdown in the highly metastatic subline decreased TGF-ß1, ZEB1, and GLUT3 expression and antagonized glycolytic metabolism. In primary tumor cells from patients with lung cancer, ANGPTL2 expression levels correlated with GLUT3 expression. Overall, this work suggests that tumor cell-derived ANGPTL2 accelerates activities associated with glycolytic metabolism in lung cancer cells by activating TGF-ß-ZEB1-GLUT3 signaling.


Assuntos
Proteínas Semelhantes a Angiopoietina/genética , Transportador de Glucose Tipo 3/genética , Neoplasias Pulmonares/genética , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética , Comunicação Autócrina/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Glicólise/genética , Humanos , Integrina alfa5beta1/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Masculino , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Metástase Neoplásica , Comunicação Parácrina/genética , Fator de Crescimento Transformador beta/genética
13.
Gene ; 738: 144483, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-32070750

RESUMO

TGFß signaling pathway is critical for the cell division, differentiation and apoptosis, the aberrant regulation of which will result in severe diseases including cancer. N6-methyl-adenosine (m6A) is one of the most abundant modifications on mRNA, it is unclear yet how m6A epitranscriptome response to stimulation of TGFß. Here, we found that cellular m6A level of RNA was elevated after TGFß treatment, which might be regulated by upregulation of WTAP and METTL3. MeRIP-Seq of mRNAs of MCF7 with or without treated by TGFß showed that mRNA with upregulated m6A modification level after TGFß treatment were enriched in TGFß signaling pathway. Phosphorylated level of SMAD2 or SMAD3 induced by TGFß was impaired when WTAP was silenced. Moreover, the m6A modification and mRNA level of JunB, which is known as a cell cycle inhibitor, both were increased after induction of TGFß and decreased after knockdown of WTAP. Intriguingly, growth inhibition caused by TGFß was rescued in WTAP-knockdown cells. Collectively, these results reveal the key role that m6A pathway playing in the cell cycle arrest induced by TGFß signaling, providing new mechanisms explanation for growth inhibition mediated by TGFß.


Assuntos
Adenosina/análogos & derivados , Ciclo Celular/genética , Fator de Crescimento Transformador beta/metabolismo , Adenosina/metabolismo , Adenosina/fisiologia , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular/genética , Divisão Celular , Linhagem Celular Tumoral , Humanos , Células MCF-7 , Metiltransferases/genética , Metiltransferases/metabolismo , Proteínas Nucleares/genética , Fosforilação , RNA/genética , Fatores de Processamento de RNA/metabolismo , RNA Mensageiro/genética , Transdução de Sinais/fisiologia , Proteína Smad2/metabolismo , Proteína Smad3/metabolismo
14.
Nat Commun ; 11(1): 785, 2020 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-32034145

RESUMO

Extracellular signals such as TGF-ß can induce epithelial-to-mesenchymal transition (EMT) in cancers of epithelial origin, promoting molecular and phenotypical changes resulting in pro-metastatic characteristics. We identified C/EBPα as one of the most TGF-ß-mediated downregulated transcription factors in human mammary epithelial cells. C/EBPα expression prevents TGF-ß-driven EMT by inhibiting expression of known EMT factors. Depletion of C/EBPα is sufficient to induce mesenchymal-like morphology and molecular features, while cells that had undergone TGF-ß-induced EMT reverted to an epithelial-like state upon C/EBPα re-expression. In vivo, mice injected with C/EBPα-expressing breast tumor organoids display a dramatic reduction of metastatic lesions. Collectively, our results show that C/EBPα is required for maintaining epithelial homeostasis by repressing the expression of key mesenchymal markers, thereby preventing EMT-mediated tumorigenesis. These data suggest that C/EBPα is a master epithelial "gatekeeper" whose expression is required to prevent unwarranted mesenchymal transition, supporting an important role for EMT in mediating breast cancer metastasis.


Assuntos
Neoplasias da Mama/patologia , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Transição Epitelial-Mesenquimal/fisiologia , Glândulas Mamárias Humanas/patologia , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Proteínas Estimuladoras de Ligação a CCAAT/genética , Células Cultivadas , Células Epiteliais/metabolismo , Feminino , Regulação da Expressão Gênica , Humanos , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/secundário , Glândulas Mamárias Humanas/metabolismo , Camundongos SCID , Proteína Smad3/genética , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Nat Commun ; 11(1): 635, 2020 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-32005814

RESUMO

Multipotent Nkx2-1-positive lung epithelial primordial progenitors of the foregut endoderm are thought to be the developmental precursors to all adult lung epithelial lineages. However, little is known about the global transcriptomic programs or gene networks that regulate these gateway progenitors in vivo. Here we use bulk RNA-sequencing to describe the unique genetic program of in vivo murine lung primordial progenitors and computationally identify signaling pathways, such as Wnt and Tgf-ß superfamily pathways, that are involved in their cell-fate determination from pre-specified embryonic foregut. We integrate this information in computational models to generate in vitro engineered lung primordial progenitors from mouse pluripotent stem cells, improving the fidelity of the resulting cells through unbiased, easy-to-interpret similarity scores and modulation of cell culture conditions, including substratum elastic modulus and extracellular matrix composition. The methodology proposed here can have wide applicability to the in vitro derivation of bona fide tissue progenitors of all germ layers.


Assuntos
Células Epiteliais/citologia , Pulmão/citologia , Camundongos/genética , Células-Tronco Pluripotentes/citologia , Animais , Técnicas de Cultura de Células , Diferenciação Celular , Células Epiteliais/metabolismo , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Feminino , Camadas Germinativas/embriologia , Camadas Germinativas/metabolismo , Pulmão/embriologia , Pulmão/metabolismo , Masculino , Camundongos/embriologia , Camundongos/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células-Tronco Pluripotentes/metabolismo , Transdução de Sinais , Fator Nuclear 1 de Tireoide/genética , Fator Nuclear 1 de Tireoide/metabolismo , Transcriptoma , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo
16.
Ann Hematol ; 99(4): 703-714, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32103323

RESUMO

Monoclonal gammopathy of renal significance (MGRS) is characterized by the nephrotoxic monoclonal immunoglobulin secreted by an otherwise asymptomatic or indolent B cell or plasma cell clone, without hematologic criteria for treatment. These MGRS-associated diseases can involve one or more renal compartments, including glomeruli, tubules, and vessels. Hydrophobic residue replacement, N-glycosylated, increase in isoelectric point in monoclonal immunoglobulin (MIg) causes it to transform from soluble form to tissue deposition, and consequently resulting in glomerular damage. In addition to MIg deposition, complement deposition is also found in C3 glomerulopathy with monoclonal glomerulopathy, which is caused by an abnormality of the alternative pathway and may involve multiple factors including complement component 3 nephritic factor, anti-complement factor auto-antibodies, or MIg which directly cleaves C3. Furthermore, inflammatory factors, growth factors, and virus infection may also participate in the development of the diseases. In this review, for the first time, we discussed current highlights in the mechanism of MGRS-related lesions.


Assuntos
Anticorpos Monoclonais/metabolismo , Nefropatias/etiologia , Paraproteinemias/etiologia , Paraproteínas/metabolismo , Autoanticorpos/imunologia , Autoanticorpos/metabolismo , Complemento C3/metabolismo , Fator Nefrítico do Complemento 3/metabolismo , Convertases de Complemento C3-C5/antagonistas & inibidores , Convertases de Complemento C3-C5/metabolismo , Via Alternativa do Complemento , Crioglobulinemia/etiologia , Crioglobulinemia/metabolismo , Glicosilação , Humanos , Amiloidose de Cadeia Leve de Imunoglobulina/etiologia , Amiloidose de Cadeia Leve de Imunoglobulina/metabolismo , Mediadores da Inflamação/metabolismo , Glomérulos Renais/metabolismo , Glomérulos Renais/patologia , Proteínas de Neoplasias/metabolismo , Paraproteinemias/complicações , Paraproteinemias/genética , Paraproteinemias/metabolismo , Processamento de Proteína Pós-Traducional , Fator de Crescimento Transformador beta/metabolismo
17.
Adv Exp Med Biol ; 1227: 81-94, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32072500

RESUMO

Gremlin is a member of the TGF-ß superfamily that can act as a BMP antagonist, and recently, has been described as a ligand of the vascular endothelial growth factor receptor 2 (VEGFR2). Gremlin shares properties with the Notch signaling pathway. Both participate in embryonic development and are reactivated in pathological conditions. Gremlin is emerging as a potential therapeutic target and biomarker of renal diseases. Here we review the role of the Gremlin-VEGFR2 axis in renal damage and downstream signaling mechanisms, such as Notch pathway.


Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Receptores Notch/metabolismo , Transdução de Sinais , Animais , Humanos , Rim/metabolismo , Rim/patologia , Fator de Crescimento Transformador beta/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
18.
Braz Oral Res ; 34: e014, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32074214

RESUMO

Although dental implants and bone regenerative procedures are important approaches for the reestablishment of esthetics and function in young patients with a history of generalized aggressive periodontitis (GAP), no predictable outcomes have been reported, and the host osteo-immunoinflammatory response may play a relevant role in this context. In view of the lack of molecular investigations into the bone tissue condition of young patients with periodontitis, the aim of this study was to evaluate the gene expression of bone-related factors in this population. Bone biopsies were obtained from the posterior mandible in 16 individuals previously diagnosed with GAP and on periodontal support therapy and from 17 periodontally healthy (PH) patients. The gene expression of tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-ß, receptor activator of the NF-κB ligand (RANKL), osteoprotegerin (OPG), osteocalcin (OC), bone sialoprotein (BSP), and type I collagen (COL-I), important biomarkers of bone turnover, was evaluated by qRT-PCR. Lower TGF-ß and OPG mRNA levels were observed in GAP patients compared to PH individuals (p ≤ 0.05). There were no between-group differences in levels of TNF-α, BSP, RANKL, OC, or COL-I mRNA (p>0.05). In young adults, a history of periodontal disease can negatively modulate the gene expression of important bone-related factors in alveolar bone tissue. These molecular outcomes may contribute to the future development of therapeutic approaches to benefit bone healing in young patients with history of periodontitis via modulation of osteo-immuno-inflammatory biomarkers.


Assuntos
Periodontite Agressiva/genética , Expressão Gênica , Adulto , Periodontite Agressiva/metabolismo , Processo Alveolar/química , Biomarcadores , Colágeno Tipo I/análise , Colágeno Tipo I/genética , Estudos Transversais , Feminino , Humanos , Sialoproteína de Ligação à Integrina/análise , Sialoproteína de Ligação à Integrina/genética , Masculino , Osteocalcina/análise , Osteocalcina/genética , Osteoprotegerina/análise , Osteoprotegerina/genética , Ligante RANK/análise , Ligante RANK/genética , Reação em Cadeia da Polimerase em Tempo Real , Valores de Referência , Método Simples-Cego , Estatísticas não Paramétricas , Fator de Crescimento Transformador beta/análise , Fator de Crescimento Transformador beta/genética , Fator de Necrose Tumoral alfa/análise , Fator de Necrose Tumoral alfa/genética , Adulto Jovem
19.
Adv Exp Med Biol ; 1202: 179-201, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32034714

RESUMO

Transforming growth factor beta (TGF-ß) signaling is involved in the regulation of proliferation, differentiation and survival/or apoptosis of many cells, including glioma cells. TGF-ß acts via specific receptors activating multiple intracellular pathways resulting in phosphorylation of receptor-regulated Smad2/3 proteins that associate with the common mediator, Smad4. Such complex translocates to the nucleus, binds to DNA and regulates transcription of many genes. Furthermore, TGF-ß-activated kinase-1 (TAK1) is a component of TGF-ß signaling and activates mitogen-activated protein kinase (MAPK) cascades. Negative regulation of TGF-ß/Smad signaling may occur through the inhibitory Smad6/7. While genetic alterations in genes related to TGF-ß signaling are relatively rare in gliomas, the altered expression of those genes is a frequent event. The increased expression of TGF-ß1-3 correlates with a degree of malignancy of human gliomas. TGF-ß may contribute to tumor pathogenesis in many ways: by direct support of tumor growth, by maintaining self-renewal of glioma initiating stem cells and inhibiting anti-tumor immunity. Glioma initiating cells are dedifferentiated cells that retain many stem cell-like properties, play a role in tumor initiation and contribute to its recurrence. TGF-ß1,2 stimulate expression of the vascular endothelial growth factor as well as the plasminogen activator inhibitor and some metalloproteinases that are involved in vascular remodeling, angiogenesis and degradation of the extracellular matrix. Inhibitors of TGF-ß signaling reduce viability and invasion of gliomas in animal models and show a great promise as novel, potential anti-tumor therapeutics.


Assuntos
Glioma/metabolismo , Glioma/patologia , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Animais , Carcinogênese , Glioma/tratamento farmacológico , Humanos , Fosforilação , Receptores de Fatores de Crescimento Transformadores beta/metabolismo
20.
Artigo em Inglês | MEDLINE | ID: mdl-32032399

RESUMO

Growth factors have been used in numerous oral applications to aid in bone formation after tooth extraction. Bone morphogenetic proteins (BMPs) are members of the transforming growth factor-b superfamily and are involved in the differentiation of pluripotent mesenchymal cells, leading to new bone formation through osteoblastic induction. This study examined histologic wound healing following extraction and ridge preservation using recombinant human BMP-2 (rhBMP-2) and a collagen sponge. Formation of new vital bone was seen, suggesting that this material is a viable option for ridge preservation in preparation for implant placement.


Assuntos
Aumento do Rebordo Alveolar , Implantes Dentários , Processo Alveolar , Proteína Morfogenética Óssea 2 , Proteínas Morfogenéticas Ósseas , Estética Dentária , Humanos , Osteogênese , Proteínas Recombinantes , Fator de Crescimento Transformador beta
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