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1.
Life Sci ; 244: 117342, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31978450

RESUMO

AIMS: Microvascular endothelial cell dysfunction is a leading cause of radiation-induced heart disease (RIHD). BRCA1 plays an important role in DNA damage repair. The study aims to explore the effect of BRCA1 in endothelial cells involved in RIHD. MATERIALS AND METHODS: BRCA1 and p21 expression were detected in human umbilical vein endothelial cells (HUVECs) and in mouse heart tissue after irradiation exposure. The effects of BRCA1 on cell proliferation, cell cycle and radiosensitivity were determined in HUVECs with overexpression and knockdown of BRCA1. A mouse model of RIHD was established. Heart damage was detected in C57BL/6J mice and endothelial cell specific knockout BRCA1 mice (EC-BRCA1-/-). KEY FINDINGS: BRCA1 and p21 expression was significantly increased both in vitro and vivo response to irradiation. BRCA1 overexpression in endothelial cells enhanced cell growth and G1/S phase arrest, and the opposite results were observed in BRCA1 knockdown endothelial cells. BRCA1 downregulated endothelial cell cycle-related genes cyclin A, cyclin D1, cyclin E and p-Rb through increasing p21 expression, and HUVECs with BRCA1 gene knockdown were more sensitive to radiation. In vivo, a decrease in cardiac microvascular density, as well as cardiomyocyte hypoxia and apoptosis were observed in a time-dependent manner. EC-BRCA1-/- mice were more prone to severe RIHD than EC-BRCA1+/- mice after 16Gy radiation exposure due to endothelial dysfunction caused by loss of BRCA1, and p21 was declined in EC-BRCA1-/- mice heart. SIGNIFICANCE: These findings indicate that BRCA1 plays a protective role in RIHD by regulating endothelial cell cycle arrest mediated by p21 signal.


Assuntos
Proteína BRCA1/metabolismo , Pontos de Checagem do Ciclo Celular , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Células Endoteliais da Veia Umbilical Humana/citologia , Neovascularização Patológica/prevenção & controle , Substâncias Protetoras/administração & dosagem , Tolerância a Radiação , Animais , Proteína BRCA1/genética , Proteína BRCA1/fisiologia , Proliferação de Células , Células Cultivadas , Inibidor de Quinase Dependente de Ciclina p21/genética , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos da radiação , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neovascularização Patológica/etiologia , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Radiação Ionizante
2.
Life Sci ; 241: 117134, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31811854

RESUMO

AIMS: Non-small cell lung cancer (NSCLC), characterized by extensive metastasis and poor prognosis, is the most common type of lung cancer. Dysregulation of certain lncRNAs is known to be linked to the tumorigenesis of NSCLC. However, the specific roles in NSCLC for many other lncRNAs, such as linc01088, remain largely unknown. MATERIALS AND METHODS: The expression patterns of linc01088, p21 and EZH2 were examined both in NSCLC tissues and cell lines using RT-qPCR assay. CCK-8, colony formation, immunofluorescence staining, and flow cytometry assays were employed to evaluate the effects of linc01088 on NSCLC cell proliferation properties. RNA immunoprecipitation (RIP) assay was performed to determine the direct binding relationship between linc01088 and zeste homolog 2 (EZH2). Western blot and RT-qPCR analysis were performed to assess p21 level within knockdown of either linc01088 or EZH2. Nude mouse subcutaneous NSCLC models were constructed for further validating the effects and mechanisms of linc01088 in vivo. KEY FINDINGS: linc01088 and EZH2 were highly expressed both in NSCLC tissues and cell lines. Knockdown of linc01088 suppressed the proliferation of NSCLC cells, and prolonged the G1 phase while shortened S and G2-M phases. RIP assay revealed the direct binding relationship between linc01088 and EZH2. Knockdown of either linc01088 or EZH2 induced up-regulation of p21 expression, which subsequently inhibited the tumor growth. SIGNIFICANCE: We demonstrated that linc01088 could promote cell proliferation via binding with EZH2 to repress p21, which aggravates the tumorigenesis of NSCLC. Therefore, linc01088 might be a potential oncogene and target for novel anti-tumor therapies.


Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma Pulmonar de Células não Pequenas/secundário , Inibidor de Quinase Dependente de Ciclina p21/antagonistas & inibidores , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/patologia , RNA Longo não Codificante/genética , Animais , Apoptose , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Proliferação de Células , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Feminino , Seguimentos , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Invasividade Neoplásica , Metástase Neoplásica , Prognóstico , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
3.
Cancer Sci ; 111(1): 175-185, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31715070

RESUMO

Neurogenic differentiation factor 1 (NeuroD1) is a transcription factor critical for promoting neuronal differentiation and maturation. NeuroD1 is involved in neuroblastoma and medulloblastoma; however, its molecular mechanism in promoting tumorigenesis remains unclear. Furthermore, the role of NeuroD1 in non-neural malignancies has not been widely characterized. Here, we found that NeuroD1 is highly expressed in colorectal cancer. NeuroD1-silencing induces the expression of p21, a master regulator of the cell cycle, leading to G2 -M phase arrest and suppression of colorectal cancer cell proliferation as well as colony formation potential. Moreover, NeuroD1-mediated regulation of p21 expression occurs in a p53-dependent manner. Through chromatin immunoprecipitation and point mutation analysis in the predicted NeuroD1 binding site of the p53 promoter, we found that NeuroD1 directly binds to the p53 promoter and suppresses its transcription, resulting in increased p53 expression in NeuroD1-silenced colorectal cancer cells. Finally, xenograft experiments demonstrated that NeuroD1-silencing suppresses colorectal cancer cell tumorigenesis potential by modulating p53 expression. These findings reveal NeuroD1 as a novel regulator of the p53/p21 axis, underscoring its importance in promoting non-neural malignancies. Furthermore, this study provides insight into the transcriptional regulation of p53.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Carcinogênese/genética , Neoplasias Colorretais/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Proteína Supressora de Tumor p53/genética , Carcinogênese/patologia , Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias Colorretais/patologia , Regulação Neoplásica da Expressão Gênica/genética , Células HCT116 , Humanos , Neuroblastoma/genética , Neuroblastoma/patologia , Regiões Promotoras Genéticas/genética , Fatores de Transcrição/genética
4.
Chemosphere ; 238: 124863, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31551201

RESUMO

Peripheral blood leukocyte telomere length in omethoate-exposed workers is related to environmental exposure and single nucleotide polymorphisms (SNPs) in genes including p21, GSTM1, miR-145, etc. However, the roles of SNPs in tankyrase (TNKS) gene in telomere length are still unknown. The aim of this study was to explore the association between SNPs in TNKS gene and telomere length in omethoate-exposed workers. Telomere length in peripheral blood leukocyte DNA from 180 omethoate-exposed workers and 115 healthy controls was measured using Real-time quantitative polymerase chain reaction (PCR). Genotyping of the selected functional and susceptible SNPs was performed by the flight mass spectrometry based on PCR and single-base extension. The analysis of covariance was performed to find effects of SNPs on telomere length. Generalized linear models were used to analyze the environment, gene, and interaction on telomere length. The results showed that telomere length in the CG + CC genotypes in rs1055328 in TNKS gene was significantly longer than that in the wild homozygous GG genotype both in exposure group (P = 0.017) and in control group (P = 0.038) after adjusting the covariates. The variables kept in the generalized linear models included omethoate-exposure (ß = 0.580, P = 0.001) and rs1055328 (CG + CC) in TNKS gene (ß = 0.339, P = 0.002). The study suggests that the prolongation of telomere length is associated with omethoate-exposure and the CG + CC genotypes in rs1055328 in TNKS gene.


Assuntos
Dimetoato/análogos & derivados , Exposição Ocupacional/efeitos adversos , Tanquirases/genética , Homeostase do Telômero/efeitos dos fármacos , Telômero/fisiologia , Adulto , Inibidor de Quinase Dependente de Ciclina p21/genética , DNA/genética , Dano ao DNA/efeitos dos fármacos , Dimetoato/toxicidade , Feminino , Genótipo , Glutationa Transferase/genética , Humanos , Leucócitos/citologia , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único/genética , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Telômero/genética
5.
Nucleic Acids Res ; 47(17): 9087-9103, 2019 09 26.
Artigo em Inglês | MEDLINE | ID: mdl-31400114

RESUMO

Most human cancers acquire mutations causing defects in the p53 signaling pathway. The tumor suppressor p53 becomes activated in response to genotoxic stress and is essential for arresting the cell cycle to facilitate DNA repair or to initiate apoptosis. p53-induced cell cycle-arrest is mediated by expression of the CDK inhibitor p21WAF1/Cip1, which prevents phosphorylation and inactivation of the pocket proteins RB, p130, and p107. In a hypophosphorylated state, pocket proteins bind to E2F factors forming RB-E2F and DREAM transcriptional repressor complexes. Here, we analyze the influence of RB and DREAM on p53-induced gene repression and cell-cycle arrest. We show that abrogation of DREAM function by knockout of the DREAM component LIN37 results in a reduced repression of cell-cycle genes. We identify the genes repressed by the p53-DREAM pathway and describe a set of genes that is downregulated by p53 independent of LIN37/DREAM. Most strikingly, p53-dependent repression of cell-cycle genes is completely abrogated in LIN37-/-;RB-/- cells leading to a loss of the G1/S checkpoint. Taken together, we show that DREAM and RB are key factors in the p53 signaling pathway to downregulate a large number of cell-cycle genes and to arrest the cell cycle at the G1/S transition.


Assuntos
Pontos de Checagem do Ciclo Celular/genética , Regulação da Expressão Gênica , Proteínas Interatuantes com Canais de Kv/metabolismo , Proteínas Repressoras/metabolismo , Proteína do Retinoblastoma/genética , Transativadores/fisiologia , Proteína Supressora de Tumor p53/metabolismo , Animais , Células Cultivadas , Proteína Substrato Associada a Crk/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Fatores de Transcrição E2F/genética , Fatores de Transcrição E2F/metabolismo , Fibroblastos/metabolismo , Genes cdc , Células HCT116 , Humanos , Proteínas Interatuantes com Canais de Kv/genética , Camundongos , Proteínas Repressoras/genética , Proteína do Retinoblastoma/metabolismo , Proteína p107 Retinoblastoma-Like/genética , Transativadores/genética , Transativadores/metabolismo , Proteína Supressora de Tumor p53/genética
6.
Artif Cells Nanomed Biotechnol ; 47(1): 2891-2899, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31307234

RESUMO

JHDM1A participates in cancer development via demethylate dimethyl histone H3 lysine 36 (H3K36me2). p300 is an intrinsic acetyltransferase. This study explored the acetyltransferase activity of p300 on JHDM1A and analyzed the JHDM1A acetylation on H3K36me2 demethylation in osteosarcoma. Co-immunoprecipitation (CoIP) and immunoblotting assay found that p300 directly acetylated JHDM1A at K409 residue in osteosarcoma MG-63 and HOS cells. Nucleosomes and mononucleosomes were prepared and found that acetylation of JHDMIA disrupted its association with nucleosomes and thereby impaired its capability to induce H3K36me2 demethylation. Moreover, chromatin immunoprecipitation (ChIP) assay discovered that the input levels of H3K36me2 in the promoter regions of p21 and puma were increased after acetylation of JHDM1A, which raised the p21 and puma mRNA levels in the cells. Finally, the analysis of JHDM1A acetylation on osteosarcoma cell proliferation and invasion, along with tumor growth pointed out that acetylation of JHDMIA inhibited the proliferation and invasion of osteosarcoma HOS cells, as well as suppressed the tumor growth of osteosarcoma. In conclusion, the outcomes of our research verified that p300 could directly acetylate JHDM1A at K409 site, which reduces the demethylation of H3K36me2, enhanced the transcription of p21 and puma, and thereby inhibited the growth and metastasis of osteosarcoma.


Assuntos
Carcinogênese , Proteína p300 Associada a E1A/metabolismo , Proteínas F-Box/metabolismo , Histona Desmetilases com o Domínio Jumonji/metabolismo , Osteossarcoma/patologia , Acetilação , Animais , Proteínas Reguladoras de Apoptose/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Proteínas F-Box/química , Histonas/metabolismo , Humanos , Histona Desmetilases com o Domínio Jumonji/química , Lisina/metabolismo , Metilação , Camundongos , Nucleossomos/metabolismo , Proteínas Proto-Oncogênicas/genética , Transcrição Genética
7.
BMC Cancer ; 19(1): 698, 2019 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-31311512

RESUMO

BACKGROUND: This research provides the first evidence of CDK5 in ccRCC prognosis and correlation with different p21 expression in overall survival (OS) analysis. METHODS: The data from both of The Cancer Genome Atlas (TCGA) and Gene Expression of Normal and Tumor Tissue (GENT) were analyzed for determining the expression of CDK5 in kidney cancer. Tissue microarray that made by using 150 ccRCC samples was used in immunohistochemistry (IHC) analysis. A validation of OS cohort was extracted from Oncomine database. RESULTS: The CDK5 expression was significantly lower in cancer tissue compared with normal in TCGA (p < 0.0001), GENT database also showed a relative low expression in kidney cancer. Among 150 ccRCC patients, low CDK5 was detected in 83 cases (55.3%), low p21 in 97 cases (64.7%). CDK5 was associated with the advanced TNM stage (p = 0.042), and Fuhrman grade (p = 0.035). Patients with lower CDK5 might be more likely to be aggressive status. According to the combination analysis of CDK5 and p21, patients in CDK5 low/p21 low group showed poorer survival rate, and no significant survival difference was observed in other groups. In the Cox multivariate analysis, the co-expression of CDK5 low/p21 low was identified as an independent prognostic factor in ccRCC patients. CONCLUSIONS: Together, our findings provided the first evidence that CDK5 was acting as a promising biomarker in ccRCC patients, and co-expression of CDK5 and p21 is an independent prognostic for overall survival. IHC analysis of CDK5 and p21 on cancer tissues after surgery may help to evaluate and predict the outcome of ccRCC patients.


Assuntos
Carcinoma de Células Renais/metabolismo , Quinase 5 Dependente de Ciclina/metabolismo , Neoplasias Renais/metabolismo , Adulto , Idoso , Biomarcadores , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/mortalidade , Carcinoma de Células Renais/patologia , Quinase 5 Dependente de Ciclina/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Neoplasias Renais/genética , Neoplasias Renais/mortalidade , Neoplasias Renais/patologia , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico
8.
Anticancer Res ; 39(7): 3493-3498, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31262873

RESUMO

BACKGROUND/AIM: Pancreatic cancer is the most lethal cancer of the digestive system. IL-29 is a new member of the IFNλ family and well-known for its strong antiviral activity. However, its direct effect on pancreatic cancer is still unclear. This study was performed to investigate if IL-29 has any direct effect on Pan-48 pancreatic cancer cells. MATERIALS AND METHODS: Clonogenic survival assay, cell proliferation, and caspase-3 activity kits were used to evaluate the effects of IL-29 on cell survival, proliferation, and apoptosis of Pan-48 pancreatic cancer cells. RT-PCR and IHC were subsequently performed to explore IL-29's potential molecular mechanisms. RESULTS: The percentage of colonies of Pan-48 cells was decreased following the addition of IL-29. This was consistent with a decreased optical density (OD) value of cancer cells. Furthermore, the relative caspase-3 activity in cancer cells was increased after the addition of IL-29, indicating increased apoptosis of cancer cells. The anti-proliferative effect of IL-29 on cancer cells correlated with increased expression of the anti-proliferative molecule p21. The pro-apoptotic effect of IL-29 on cancer cells correlated with an increased expression of the pro-apoptotic molecule Bax. CONCLUSION: IL-29 constrains Pan-48 pancreatic cell growth via up-regulation of p21 and Bax. Our study suggests a potential use of IL-29 in immunotherapy for pancreatic cancer treatment.


Assuntos
Antineoplásicos/farmacologia , Inibidor de Quinase Dependente de Ciclina p21/genética , Interleucinas/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Proteína X Associada a bcl-2/genética , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Interferons , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , RNA Mensageiro/metabolismo , Regulação para Cima , Proteína X Associada a bcl-2/metabolismo
9.
Mol Med Rep ; 20(2): 1569-1574, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31257531

RESUMO

Short­chain fatty acids (SCFAs; butyrate, propionate and acetate) are metabolites derived from the gut microbiota via dietary fiber fermentation. In colon cancer, treatment with SCFAs, mainly butyrate and propionate, suppresses cell proliferation, migration and invasion. Furthermore, although sodium butyrate is known to induce cell apoptosis in lung cancer, the anticancer effects of sodium propionate (SP) on lung cancer are not well understood. In the present study, SP treatment induced cell cycle arrest, especially in the G2/M phase, and cell apoptosis in the H1299 and H1703 lung cancer cell lines. As determined by reverse transcription­quantitative PCR and western blotting, Survivin and p21 expression levels were significantly affected by SP treatment, suggesting that SP treatment suppressed cell proliferation in these lung cancer cell lines. Thus, it was proposed that the SP­mediated regulation of Survivin and p21 in lung cancer may be applicable to lung cancer therapy.


Assuntos
Antineoplásicos/farmacologia , Ácido Butírico/farmacologia , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Propionatos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p21/agonistas , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Microbioma Gastrointestinal/fisiologia , Humanos , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Transdução de Sinais , Survivina/antagonistas & inibidores , Survivina/genética , Survivina/metabolismo
10.
Talanta ; 204: 20-28, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31357283

RESUMO

Hepatocarcinoma is the second leading cause of cancer-related death worldwide accompanied by the aberrant expression of many genes. Among of them, p53 and p21 genes played a vital role in the development of liver cancer. Thus, monitoring mRNA levels of the two markers is urgently needed for early diagnosis and understanding the molecular mechanism of hepatocarcinoma development. Herein, a functional nanosystem for in situ and real-time monitoring levels of p53 and p21 mRNA was constructed using reduced graphene oxide nanosheet assisted fluorescence probes. Comparing with traditional methods, the new method indicated high sensitivity and outstanding selectivity towards p53 and p21 mRNA assay in vitro. Moreover, the functional nanosystem was successfully used for monitoring dynamic change of mRNAs in HepG2 cells caused by cisplatin treatment, the reliability of which was confirmed by RT-PCR. In summary, this strategy provides a promising tool to reveal p53 and p21 mRNA regulatory process in cancer cells, which is practicable for drug screening and therapy evaluation on clinic.


Assuntos
Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Grafite/química , Nanoestruturas/química , RNA Mensageiro/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Adsorção , Antineoplásicos/farmacologia , Técnicas Biossensoriais/métodos , Linhagem Celular Tumoral , Cisplatino/farmacologia , Inibidor de Quinase Dependente de Ciclina p21/genética , DNA/química , DNA/genética , Sondas de DNA/química , Sondas de DNA/genética , Corantes Fluorescentes/química , Humanos , Limite de Detecção , Microscopia Confocal/métodos , Microscopia de Fluorescência/métodos , Hibridização de Ácido Nucleico , RNA Mensageiro/genética , Proteína Supressora de Tumor p53/genética
11.
Med Sci Monit ; 25: 4207-4216, 2019 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-31169265

RESUMO

BACKGROUND The role of the ubiquitin-specific peptidase 9 X-linked (USP9X) gene in breast cancer remains poorly understood. This study aimed to investigate the role of USP9X in breast cancer tissue and cell lines. MATERIAL AND METHODS Immunohistochemistry was used to examine the expression levels of USP9X in 102 breast cancer tissue samples and 41 normal breast tissue samples. Overexpression of USP9X in MCF-7 and MDA-MB-231 breast cancer cell lines were studied by USP9X lentivirus vector transfection. Clustered regularly interspaced short palindromic repeats (CRISPR)/caspase-9 USP9X gene knockout was performed. Cell proliferation, growth, and survival were examined using the cell counting kit-8 (CCK-8) assay, the colony formation assay, flow cytometry assays, and a tumor xenograft study. RESULTS Immunohistochemistry showed that USP9X was significantly overexpressed in 93 of 102 (91.1%) breast cancer tissue samples compared with 41 normal breast tissue samples and was associated with tumor size ≥5.0 cm (P<0.05). USP9X overexpression in MCF-7 and MDA-MB-231 breast cancer increased cell proliferation and survival, significantly reduced the number of cells in the G1-phase cells and increased the number of cells in the S-phase cells, which were reversed by CRISPR/caspase-9 USP9X gene knockout. Overexpression of USP9X upregulated the CCND1 gene encoding cyclin D1 and downregulated cyclin-dependent inhibitor kinase 1A (CDKN1A) gene in breast cancer cells, which were reversed by USP9X knockout. CONCLUSIONS Overexpression of USP9X was associated with upregulation of the CCND1 gene and downregulation of the CDKN1A gene in breast cancer tissue and cell lines.


Assuntos
Neoplasias da Mama/genética , Ciclina D1/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Ubiquitina Tiolesterase/genética , Adulto , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Carcinogênese , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Ciclina D1/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Xenoenxertos , Humanos , Imuno-Histoquímica , Células MCF-7 , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Ubiquitina Tiolesterase/biossíntese , Ubiquitina Tiolesterase/metabolismo
12.
Eur J Med Chem ; 177: 302-315, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31158746

RESUMO

Betulin-1,4-quinone hybrids were obtain by connecting two active structures with a linker. This strategy allows for obtaining compounds showing a high biological activity and better bioavailability. In this research, synthesis, anticancer activity and molecular docking study of betulin-1,4-quinone hybrids are presented. Newly synthesized compounds were characterized by 1H, 13C NMR, IR and HR-MS. Hybrids were tested in vitro against a panel of human cell lines including glioblastoma, melanoma, breast and lung cancer. They showed a high cytotoxic activity depending on the type of 1,4-quinone moiety and the applied tumor cell lines. It was found that cytotoxic activities of the studied hybrids were increasing against the cell line with higher NQO1 protein level, like melanoma (C-32), breast (MCF-7) and lung (A-549) cancer. Selected hybrids were tested on the transcriptional activity of the gene encoding a proliferation marker (H3 histone), a cell cycle regulators (p53 and p21) and an apoptosis pathway (BCL-2 and BAX). The obtained results suggested that the tested compounds caused a mitochondrial apoptosis pathway in A549 and MCF-7 cell lines. The molecular docking was used to examine the probable interaction between the hybrids and human NAD[P]H-quinone oxidoreductase (NQO1) protein. The computational studies showed that the type of the 1,4-quinone moiety affected the location of the compound in the active site of the enzyme. Moreover, it was shown that an interaction of 1,4-quinone fragment with the hydrophobic matrix of the active site near Tyr128, Phe178, Trp105 and FAD cofactor could explain the observed increase of TP53 gene expression.


Assuntos
Antineoplásicos/farmacologia , NAD(P)H Desidrogenase (Quinona)/metabolismo , Quinonas/farmacologia , Triterpenos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/metabolismo , Apoptose/efeitos dos fármacos , Betula/química , Domínio Catalítico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p21/genética , Ensaios de Seleção de Medicamentos Antitumorais , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , NAD(P)H Desidrogenase (Quinona)/química , Ligação Proteica , Quinonas/síntese química , Quinonas/química , Quinonas/metabolismo , Relação Estrutura-Atividade , Triterpenos/síntese química , Triterpenos/química , Triterpenos/isolamento & purificação , Triterpenos/metabolismo , Proteína Supressora de Tumor p53/genética
13.
Int J Mol Sci ; 20(11)2019 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-31146388

RESUMO

Stem cells undergo senescence both in vivo, contributing to the progressive decline in self-healing mechanisms, and in vitro during prolonged expansion. Here, we show that an early developmental zebrafish embryo extract (ZF1) could act as a modulator of senescence in human mesenchymal stem cells (hMSCs) isolated from both adult tissues, including adipose tissue (hASCs), bone marrow (hBM-MSCs), dental pulp (hDP-MSCs), and a perinatal tissue such as the Wharton's Jelly (hWJ-MSCs). In all the investigated hMSCs, ZF1 decreased senescence-associated ß-galactosidase (SA ß-gal) activity and enhanced the transcription of TERT, encoding the catalytic telomerase core. In addition, it was associated, only in hASCs, with a transcriptional induction of BMI1, a pleiotropic repressor of senescence. In hBM-MSCs, hDP-MSCs, and hWJ-MSCs, TERT over-expression was concomitant with a down-regulation of two repressors of TERT, TP53 (p53), and CDKN1A (p21). Furthermore, ZF1 increased the natural ability of hASCs to perform adipogenesis. These results indicate the chance of using ZF1 to modulate stem cell senescence in a source-related manner, to be potentially used as a tool to affect stem cell senescence in vitro. In addition, its anti-senescence action could also set the basis for future in vivo approaches promoting tissue rejuvenation bypassing stem cell transplantation.


Assuntos
Senescência Celular , Embrião não Mamífero/química , Células-Tronco Mesenquimais/efeitos dos fármacos , Extratos de Tecidos/farmacologia , Animais , Células Cultivadas , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Complexo Repressor Polycomb 1/genética , Complexo Repressor Polycomb 1/metabolismo , Telomerase/genética , Telomerase/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Peixe-Zebra
14.
EBioMedicine ; 45: 58-69, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31202814

RESUMO

BACKGROUND: Long noncoding RNAs (lncRNAs) have been identified as regulators of a number of developmental and tumorigenic processes. However, the functions of most lncRNAs in glioma remain unknown and the mechanisms governing the proliferation of tumor cells remain poorly defined. METHODS: Both in vitro and in vivo assays were performed to investigate the roles of lncRNAs in the pathophysiology of gliomas. lncRNA arrays were used to identify differentially expressed lncRNAs. Subcutaneous tumor formation and a brain orthotopic tumor model in nude mice were used to investigate the functions of lncRNAs in vivo. The in vitro functions of lncRNAs were analyzed by fluorescence-activated cell sorting, colony formation, and western blot analyses. RNA fluorescence in situ hybridization and immunoprecipitation were used to explore the underlying mechanisms. FINDINGS: Here, we describe the newly discovered noncoding RNA RP11-732M18.3, which is highly overexpressed in glioma cells and interacts with 14-3-3ß/α to promote glioma growth, acting as an oncogene. Overexpression of lncRNA RP11-732 M18.3 was associated with the proliferation of glioma cells and tumor growth in vitro and in vivo. Remarkably, lncRNA RP11-732M18.3 promoted cell proliferation and G1/S cell cycle transition. lncRNA RP11-732M18.3 is predominately localized in the cytoplasm. Mechanistically, the interaction of lncRNA RP11-732M18.3 with 14-3-3ß/α increases the degradation of the p21 protein. lncRNA RP11-732M18.3 promoted the recruitment of ubiquitin-conjugating enzyme E2 E1 to 14-3-3ß/α and the binding of 14-3-3ß/α with ubiquitin-conjugating enzyme E2 E1 (UBE2E1) promoted the degradation of p21. INTERPRETATION: Overall these data demonstrated that lncRNA RP11-732M18.3 regulates glioma growth through a newly described lncRNA-protein interaction mechanism. The inhibition of lncRNA RP11-732M18.3 could provide a novel therapeutic target for glioma treatment.


Assuntos
Proteínas 14-3-3/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Glioma/tratamento farmacológico , RNA Longo não Codificante/genética , Animais , Apoptose/genética , Carcinogênese/genética , Movimento Celular/genética , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Glioma/genética , Glioma/patologia , Xenoenxertos , Humanos , Hibridização in Situ Fluorescente , Camundongos , Terapia de Alvo Molecular , Ligação Proteica/genética , Proteólise , Enzimas de Conjugação de Ubiquitina/genética
15.
Nat Commun ; 10(1): 2016, 2019 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-31043605

RESUMO

Appropriate therapeutic modulation of endothelial proliferation and sprouting is essential for the effective inhibition of angiogenesis in cancer or its induction in cardiovascular disease. The current view is that an increase in growth factor concentration, and the resulting mitogenic activity, increases both endothelial proliferation and sprouting. Here, we modulate mitogenic stimuli in different vascular contexts by interfering with the function of the VEGF and Notch signalling pathways at high spatiotemporal resolution in vivo. Contrary to the prevailing view, our results indicate that high mitogenic stimulation induced by VEGF, or Notch inhibition, arrests the proliferation of angiogenic vessels. This is due to the existence of a bell-shaped dose-response to VEGF and MAPK activity that is counteracted by Notch and p21, determining whether endothelial cells sprout, proliferate, or become quiescent. The identified mechanism should be considered to achieve optimal therapeutic modulation of angiogenesis.


Assuntos
Endotélio Vascular/efeitos dos fármacos , Mitógenos/farmacologia , Neovascularização Patológica/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Animais , Proliferação de Células/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Endotélio Vascular/patologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Camundongos , Camundongos Knockout , Neovascularização Patológica/patologia , Receptores Notch/antagonistas & inibidores , Receptores Notch/metabolismo , Retina , Vasos Retinianos , Transdução de Sinais/genética , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/metabolismo
16.
Int J Mol Sci ; 20(9)2019 Apr 28.
Artigo em Inglês | MEDLINE | ID: mdl-31035388

RESUMO

Transactivation of p21 (cyclin-dependent kinase inhibitor 1A, CDKN1A) is closely related to the recruitment of transcription cofactors at the p53 responsive elements (p53REs) in its promoter region. Human chromatin remodeling enzyme INO80 can be recruited to the p53REs of p21 promoter and negatively regulates p21. As one of the key subunits of the INO80 complex, YY1 has also been confirmed to bind to the p53RE sites of p21 promoter. Importantly, YY1 was recently reported to be bound and stabilized by BCCIP (BRCA2 and CDKN1A-interacting protein). Therefore, we hypothesized that the YY1/BCCIP complex plays an important role in regulating the transactivation of p21. Here we present evidence that the YY1/BCCIP complex coordinatively regulates p53RE-mediated p21 transactivation. We first confirmed the cross-interaction between YY1, BCCIP, and p53, suggesting an intrinsic link between three proteins in the regulation of p21 transcription. In dual luciferase assays, YY1 inhibited p53RE-mediated luciferase activity, whereas BCCIP revealed the opposite effect. More interestingly, the region 146-270 amino acids of YY1, which bound to BCCIP, increased p53-mediated luciferase activity, indicating the complexity of the YY1/BCCIP complex in co-regulating p21 transcription. Further in-depth research confirmed the co-occupancy of YY1/BCCIP with p53 at the p53RE-proximal region of p21. Lentiviral-mediated knockdown of BCCIP inhibited the recruitment of p53 and YY1 at the p53RE proximal region of p21; however, this phenomenon was reversed by expressing exogenous YY1, suggesting the collaborative regulation of YY1/BCCIP complex in p53RE-mediated p21 transcription. These data provide new insights into the transcriptional regulation of p21 by the YY1/BCCIP complex.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ciclo Celular/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Proteínas Nucleares/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Fator de Transcrição YY1/metabolismo , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ciclo Celular/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Células HCT116 , Humanos , Proteínas Nucleares/genética , Ativação Transcricional/genética , Ativação Transcricional/fisiologia , Fator de Transcrição YY1/genética
17.
Nat Commun ; 10(1): 1969, 2019 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-31036808

RESUMO

Long noncoding RNAs (lncRNAs) are emerging as regulators of fundamental biological processes. Here we report on the characterization of an intergenic lncRNA expressed in epithelial tissues which we termed EPR (Epithelial cell Program Regulator). EPR is rapidly downregulated by TGF-ß and its sustained expression largely reshapes the transcriptome, favors the acquisition of epithelial traits, and reduces cell proliferation in cultured mammary gland cells as well as in an animal model of orthotopic transplantation. EPR generates a small peptide that localizes at epithelial cell junctions but the RNA molecule per se accounts for the vast majority of EPR-induced gene expression changes. Mechanistically, EPR interacts with chromatin and regulates Cdkn1a gene expression by affecting both its transcription and mRNA decay through its association with SMAD3 and the mRNA decay-promoting factor KHSRP, respectively. We propose that EPR enables epithelial cells to control proliferation by modulating waves of gene expression in response to TGF-ß.


Assuntos
Estabilidade de RNA/genética , RNA Longo não Codificante/genética , Proteína Smad3/metabolismo , Transcriptoma/genética , Fator de Crescimento Transformador beta/farmacologia , Animais , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Células Cultivadas , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Humanos , RNA Longo não Codificante/efeitos dos fármacos
18.
Mol Med Rep ; 19(6): 5195-5202, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31059098

RESUMO

MicroRNAs (miRNAs) are considered to be critical mediators of gene expression with respect to tumor progression, although their role in ischemia­induced angiogenesis is poorly characterized, including in peripheral arterial disease (PAD). Furthermore, the underlying mechanism of action of specific miRNAs in PAD remains unknown. Reverse transcription­quantitative polymerase chain reaction analysis revealed that microRNA­93 (miR­93) was significantly upregulated in patients with PAD and in the EA.hy926 endothelial cells in response to hypoxia. Additionally, miRNA (miR)­93 promoted angiogenesis by enhancing proliferation, migration and tube formation. Cyclin dependent kinase inhibitor 1A (CDKN1A), verified as a potential target gene of miR­93, was inhibited by overexpressed miR­93 at the protein and mRNA expression levels. Furthermore, a hind­limb ischemia model served to evaluate the role of miR­93 in angiogenesis in vivo, and the results demonstrated that miR­93 overexpression enhanced capillary density and perfusion recovery from hind­limb ischemia. Taken together, miR­93 was indicated to be a promising target for pharmacological regulation to promote angiogenesis, and the miR­93/CDKN1A pathway may function as a novel therapeutic approach in PAD.


Assuntos
Inibidor de Quinase Dependente de Ciclina p21/metabolismo , MicroRNAs/metabolismo , Neovascularização Fisiológica , Doença Arterial Periférica/patologia , Regiões 3' não Traduzidas , Idoso , Animais , Hipóxia Celular , Linhagem Celular , Proliferação de Células , Inibidor de Quinase Dependente de Ciclina p21/química , Inibidor de Quinase Dependente de Ciclina p21/genética , Modelos Animais de Doenças , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Feminino , Membro Posterior/patologia , Humanos , Isquemia/metabolismo , Isquemia/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/química , MicroRNAs/genética , Pessoa de Meia-Idade , Doença Arterial Periférica/sangue , Doença Arterial Periférica/genética
19.
Ann Endocrinol (Paris) ; 80(3): 153-158, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31072588

RESUMO

Gastroenteropancreatic Neuroendocrine Neoplasms (GEP-NENs) arise throughout the gut and feature varying biological behaviour and malignant potential. GEP-NENs include two genetically different entities, well-differentiated neuroendocrine tumours (NETs) and poorly differentiated neuroendocrine carcinomas (NEC). NECs are characterized by a dismal prognosis and by distinctive TP53 and RB1 inactivation which sets them apart from NETs. The latter, conversely, have a wide spectrum of aggressiveness and molecular alterations. Knowledge on their biology has recently expanded thanks to high-throughput studies focused on two important groups of well-differentiated neuroendocrine neoplasms: pancreatic (PanNETs) and small intestinal (SiNETs) tumours. PanNETs have been among the most studied also due to genetic syndromes featuring their onset. Research stemming from this observation has uncovered the inactivation of MEN1, VHL, TSC1/2, and the hyperactivation of the PI3K/mTOR pathway as distinctive biological features of these neoplasms. Next-Generation Sequencing added information on the role of telomere lengthening via ATRX/DAXX inactivation in a fraction of PanNETs, while other display shortened telomeres and recurrent chromosomal alterations. The data so far disclosed a heterogeneous combination of driver events, yet converging into four pathways including DNA damage repair, cell cycle regulation, PI3K/mTOR signalling and telomere maintenance. SiNETs showed a lesser relationship with mutational driver events, even in the case of familial cases. High throughput studies identified putative driver mutations in CDKN1 and APC which, however, were reported in a minor fraction (∼10%) of cases. Tumorigenesis of SiNETs seems to depend more on chromosomal alterations (loss of chromosome 8, gains at 4, 5 and 20) and epigenetic events, which converge to hyperactivate the PI3K/mTOR, MAPK and Wnt pathways. While calling for further integrative studies, these data lay previous and recent findings in a more defined frame and provide clinical research with several candidate markers for patient stratification and companion diagnostics.


Assuntos
Neoplasias Intestinais/genética , Intestino Delgado , Tumores Neuroendócrinos/genética , Neoplasias Pancreáticas/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Epigênese Genética , Amplificação de Genes , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Neoplasias Intestinais/fisiopatologia , Proteínas Quinases Ativadas por Mitógeno/genética , Terapia de Alvo Molecular , Mutação , Tumores Neuroendócrinos/fisiopatologia , Neoplasias Pancreáticas/fisiopatologia , Fosfatidilinositol 3-Quinases/genética , Transdução de Sinais , Síndrome , Serina-Treonina Quinases TOR/genética , Via de Sinalização Wnt/genética
20.
DNA Cell Biol ; 38(7): 651-659, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31140859

RESUMO

Breast cancer is the leading cause of death in women. Although numerous clinical regimens are used to treat breast cancer and manifest satisfied efficacy, drug resistance is emerging as the major obstacle to their long-term use. It is critically necessary to decipher the molecular mechanism underlying this process to obtain improved and long-term use of each regimen. In the present study, we showed the negative relationship between EZH2 and chemoresistance to taxol in breast cancer cells. EZH2 interference was capable of decreasing while overexpression increasing apoptosis of breast cancer cells challenged with taxol. Meanwhile, p21, the inhibitor of cell cycle entry, interference upregulated, while overexpression downregulated apoptosis induced by taxol. Mechanistically, EZH2 was recruited to the promoter of p21 accompanied with H3K27me3 enrichment and transcription silencing. Collectively, EZH2 attenuates chemoresistance of breast cancer cells to taxol by dampening p21 epigenetically.


Assuntos
Antineoplásicos Fitogênicos/toxicidade , Neoplasias da Mama/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Epigênese Genética , Paclitaxel/toxicidade , Apoptose/efeitos dos fármacos , Neoplasias da Mama/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Resistencia a Medicamentos Antineoplásicos , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Feminino , Humanos , Células MCF-7
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