RESUMO
AIM: To investigate the potential roles and mechanisms of miR-17-5p/20a in human gastric cancer development and progression. METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to determine miR-17-5p/20a expression profiles in 110 gastric cancer tissues. microRNAs' (miRNAs) mimics and inhibitors were used to reveal their function in gastric cancer. Antagomirs were applied to treating gastric cancer cell derived xenograft in vivo. Western blot and luciferase assays were performed to uncover the targets and mechanisms of miR-17-5p/20a. RESULTS: miR-17-5p/20a levels were upregulated in human gastric cancer tissues. Overexpression of miR-17-5p/20a promoted gastric cancer cell cycle progression and inhibited cell apoptosis, whereas knockdown of miR-17-5p/20a resulted in cell cycle arrest and increased apoptosis. p21 and tumour protein p53-induced nuclear protein 1 (TP53INP1) were validated as the targets of miR-17-5p/20a. Antagomirs against miR-17-5p/20a significantly inhibited gastric cancer growth via upregulation of p21 and TP53INP1 in a mouse xenograft model. The negative relationship between miR-17-5p/20a and TP53INP1 was observed in patient gastric cancer tissues. Murine double minute 2 (MDM2) was found to be involved in miRNA regulation and function. Targeted inhibition of MDM2 in a miRNA mimic-transfected gastric cancer cell line abolished miR-17-5p/20a function and inhibition of p21 expression. MDM2 restoration by pCMV-MDM2 rescued the functionality. CONCLUSIONS: Our findings indicate that miR-17-5p/20a promote gastric cancer cell proliferation and inhibit cell apoptosis via post-transcriptional modulation of p21 and TP53INP1. They may be promising therapeutic markers for gastric cancer. MDM2 contributes to miR-17-5p/20a function and inhibition of p21 in gastric cancer, and may be a novel mechanism underlying the oncogenic roles of miR-17-5p/20a.
Assuntos
Biomarcadores Tumorais/genética , MicroRNAs/genética , Proteínas Proto-Oncogênicas c-mdm2/genética , Neoplasias Gástricas/genética , Animais , Apoptose/genética , Western Blotting , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Oligonucleotídeos/genética , Oligonucleotídeos/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Neoplasias Gástricas/patologia , Neoplasias Gástricas/prevenção & controle , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Circulating miR-17-5p and miR-20a (miR-17-5p/20a) have been demonstrated to be elevated in the plasma of gastric cancer patients. However, the clinical significance of the circulating levels of these microRNAs (miRNAs), the predictive power for prognosis and application for monitoring of chemotherapeutic effects remain unclear. To this end, we measured plasma miR-17-5p/20a levels in unpaired pre-operative (n=65), post-operative (n=16) and relapse (n=6) gastric cancer patient groups. The 3-year overall survival rate for the unpaired pre-operative patients was recorded. The circulating levels of miR-17-5p/20a were also tested in paired pre-operative and post-operative plasma from 14 gastric cancer patients. We found that the concentrations of miR-17-5p/20a were significantly associated with the differentiation status and TNM stages of gastric cancer. The miRNA levels in the different groups reflected pathological tumor progression. Kaplan-Meier curve analysis revealed that high expression levels of miR-17-5p/20a were significantly correlated with poor overall survival. Cox regression analysis demonstrated that the level of plasma miR-20a was an independent risk predictor for prognosis. An in vivo mouse tumor model was established and antagomirs against miR-17-5p/20a were applied as chemotherapeutics to perform tumor treatment. An assay of serum miR-17-5p/20a levels showed that the levels of serum miRNAs were notably reduced in post-treated mice with tumor volume regression. Taken together, the levels of circulating miR-17-5p/20a may be a promising non-invasive molecular marker for pathological progression, prediction of prognosis and monitoring of chemotherapeutic effects for gastric cancer.
Assuntos
Biomarcadores/sangue , Regulação Neoplásica da Expressão Gênica , MicroRNAs/sangue , Neoplasias Gástricas/sangue , Idoso , Animais , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Estimativa de Kaplan-Meier , Masculino , Camundongos , MicroRNAs/antagonistas & inibidores , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Oligonucleotídeos Antissenso/farmacologia , Prognóstico , Análise de Regressão , Fatores de Risco , Neoplasias Gástricas/mortalidade , Transplante HeterólogoRESUMO
BACKGROUND: Mesenchymal stem cells (MSCs) promote tumor growth by differentiating into carcinoma-associated fibroblasts (CAFs) and composing the tumor microenvironment. However, the mechanisms responsible for the transition of MSCs to CAFs are not well understood. Exosomes regulate cellular activities by mediating cell-cell communication. In this study, we aimed to investigate whether cancer cell-derived exosomes were involved in regulating the differentiation of human umbilical cord-derived MSCs (hucMSCs) to CAFs. METHODOLOGY/PRINCIPAL FINDINGS: We first showed that gastric cancer cell-derived exosomes induced the expression of CAF markers in hucMSCs. We then demonstrated that gastric cancer cell-derived exosomes stimulated the phosphorylation of Smad-2 in hucMSCs. We further confirmed that TGF-ß receptor 1 kinase inhibitor attenuated Smad-2 phosphorylation and CAF marker expression in hucMSCs after exposure to gastric cancer cell-derived exosomes. CONCLUSION/SIGNIFICANCE: Our results suggest that gastric cancer cells triggered the differentiation of hucMSCs to CAFs by exosomes-mediated TGF-ß transfer and TGF-ß/Smad pathway activation, which may represent a novel mechanism for MSCs to CAFs transition in cancer.