RESUMEN
Recent studies have provided evidence for tumor suppressive function of the embryonic stem cell-specific miR-302/367 cluster through induction of a reprogramming process. Aspirin has been found to induce reprogramming factors of mesenchymal-to-epithelial transition in breast cancer cells. Therefore, we aimed to investigate whether overexpression of miR-302/367 cluster and aspirin treatment cooperate in the induction of reprogramming and tumor suppression in breast cancer cells. MDA-MB-231 and SK-BR-3 human breast cancer cell lines were transfected with a miR-302/367 expressing vector and treated with aspirin. The cells were evaluated for indices of apoptosis, proliferation, migration, and invasion. In both cell lines, treatment of miR-302/367-transfected cells with aspirin upregulated expression of some main pluripotency factors such as OCT4, SOX2, NANOG, and KLF4, and downregulated expression of some invasion and angiogenesis markers at gene and protein levels. Aspirin increased the apoptotic rate in both cell lines transfected with miR-302/367. Both miR-302/367 and aspirin upregulated the expression of FOXD3 protein which is a known inducer of OCT4 and NANOG. Our results demonstrate that aspirin can enhance miR-302/367-induced reprogramming of breast cancer cells possibly through upregulation of FOXD3 expression. This can further augment the reversal of epithelial-mesenchymal transition and inhibits migration, invasion, and angiogenic signaling in breast cancer cells reprogrammed by miR-302/367. Therefore, aspirin may serve as a useful adjuvant for reprogramming of cancer cells.
Asunto(s)
Neoplasias de la Mama , MicroARNs , Aspirina/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Transición Epitelial-Mesenquimal/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , MicroARNs/genética , MicroARNs/metabolismoRESUMEN
Overexpression of either miR-302 or miR-302/367 cluster induces reprogramming of cancer cells and exerts tumor-suppressive effects by induction of mesenchymal-to-epithelial transition, apoptosis and a less proliferative capacity. Several reports have described miR-16 as a tumor suppressor microRNA (miRNA). Here, we studied the impact of exogenous induction of miR-16 in MDA-MB-231 and SK-BR-3 breast cancer cells following overexpression of miR-302/367 cluster and investigated whether transfection of these cells by a mature miR-16 mimic could affect the reprogramming state of the cells and their tumorigenicity. miR-16 enhanced the expression levels of OCT4A, SOX2, and NANOG, generally known as transcription or pluripotency factors, and suppressed proliferation and invasiveness of these cells. Meanwhile, inhibition of miR-16 counteracted both the reprogramming effect and the antitumor function of miR-302/367 in the breast cancer cells. Current results indicate that miR-16 can work as an adjuvant to improve both cancer cell reprogramming and tumor-suppressive function of miR-302/367 cluster in MDA-MB-231 and SK-BR-3 cells, while its inhibition counteracts all of these effects. Combined application of miRNAs that share some common targets in cancer cell signaling pathways may provide new approaches for repression of multiple hallmarks of cancer.
Asunto(s)
Reprogramación Celular/genética , Células Epiteliales/metabolismo , MicroARNs/genética , Apoptosis/genética , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Células Epiteliales/patología , Transición Epitelial-Mesenquimal/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Glándulas Mamarias Humanas/metabolismo , Glándulas Mamarias Humanas/patología , MicroARNs/metabolismo , Proteína Homeótica Nanog/genética , Proteína Homeótica Nanog/metabolismo , Factor 3 de Transcripción de Unión a Octámeros/genética , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Factores de Transcripción SOXB1/genética , Factores de Transcripción SOXB1/metabolismo , Transducción de Señal , Esferoides Celulares/metabolismo , Esferoides Celulares/patologíaRESUMEN
Recent investigations have shown tumor-suppressive roles for miR-16 and miR-34a. They also share some features in regard to targeting cancer cell signaling pathways which they control. Therefore, in this study, we aimed to further scrutinize whether exogenous induction of mature miR-34a and miR-16 can collaborate in breast tumor suppression. MDA-MB-231 and SK-BR-3 human breast cancer cell lines were cultured and transfected twice with hsa-miR-16-5p and hsa-miR-34a-5p mimics individually or in combination. The cells were analyzed for apoptosis rate and cell cycle indices by flow cytometry. Also, the expression of several invasion and the epithelial-mesenchymal transition markers was evaluated at gene and protein levels by quantitative real-time polymerase chain reaction and western blot analysis, respectively. Assessment of invasiveness and migratory potential of the transfected cells was performed using three-dimensional spheroid formation and wound-healing assay, respectively. In both cell lines, miR-16 and miR-34a induced apoptosis and cell-cycle arrest and also suppressed invasion and migration. Some of these effects, like cell-cycle arrest and induction of apoptosis, were significantly higher when using both microRNAs than when using them individually for transfection of the cells. Our results are indicating that miR-16 and miR-34a can collaborate in breast tumor suppression.
Asunto(s)
Neoplasias de la Mama/metabolismo , MicroARNs/metabolismo , Western Blotting , Neoplasias de la Mama/genética , Técnicas de Cultivo de Célula , Línea Celular Tumoral , Transformación Celular Neoplásica/metabolismo , Transición Epitelial-Mesenquimal/genética , Transición Epitelial-Mesenquimal/fisiología , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , MicroARNs/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Cicatrización de Heridas/genética , Cicatrización de Heridas/fisiologíaRESUMEN
Epigenetic reprogramming by embryonic stem cell-specific miR-302/367 cluster has shown some tumor suppressive effects in cancer cells of different tissues such as skin, colon, and cervix. Vitamin C has been known as a reprogramming enhancer of human and mouse somatic cells. In this study, first we aimed to investigate whether exogenous induction of miR-302/367 in breast cancer cells shows the same tumor suppressive effects previously observed in other cancer cells lines, and whether vitamin C can enhance reprogramming of breast cancer cells and also improve the tumor suppressive function of miR-302/367 cluster. Overexpression of miR-302/367 cluster in MDA-MB-231 and SK-BR-3 breast cancer cells upregulated expression of miR-302/367 members and also some core pluripotency factors including OCT4A, SOX2 and NANOG, induced mesenchymal to epithelial transition, suppressed invasion, proliferation, and induced apoptosis in the both cell lines. However, treatment of the miR-302/367 transfected cells with vitamin C suppressed the expression of pluripotency factors and augmented the tumorigenicity of the breast cancer cells by restoring their proliferative and invasive capacity and compromising the apoptotic effect of miR-302/367. Supplementing the culture medium with vitamin C downregulated expression of TET1 gene which seems to be the reason behind the negative impact of vitamin C on the reprogramming efficiency of miR-302/367 cluster and its anti-tumor effects. Therefore application of vitamin C may not always serve as a reprogramming enhancer depending on its switching function on TET1. This phenomenon should be carefully considered when considering a reprogramming strategy for tumor suppression.