RESUMO
OBJECTIVES: The aim of this study was to determine expression levels of miR-433 in oral squamous cell carcinomas (OSCCs) and adjacent normal tissues, and explore its biological functions in OSCCs. METHODS: miR-433 level in oral squamous cell carcinomas (OSCCs) and adjacent normal tissues was tested by real-time qPCR. The effect of miR-433 on cell growth was detected by MTT and colony formation assays. The tumorigenicity of miR-433 transfected OSCCs was evaluated in nude mice model. Transwell and wound healing assays were performed to detect the effect of miR-433 on OSCCs cell invasion and migration. Luciferase reporter gene assays were performed to identify the interaction between miR-433 and 3'UTR of HDAC6 mRNA. The protein level of HDAC6, BCL2, CCNE1, MMP1 and MMP9 was determined by Western blotting. Immunohistochemistry analysis was performed to detect the expression of HDAC6 in oral squamous cell carcinomas (OSCCs) and adjacent normal tissues. RESULTS: We found that miR-433 was frequently down-regulated in OSCCs compared with adjacent normal tissues. Restoring miR-433 expression in OSCC cells dramatically suppressed cells growth, invasion and migration. Importantly, our data showed that miR-433 downregulated the expression of HDAC6 through directly targeting its 3'UTR. CONCLUSION: Our data suggest that miR-433 exerts its tumor suppressor function by targeting HDAC6, leading to the inhibition of OSCC cell growth, invasion and migration, which suggest that miR-433 may be potential target for diagnostic and therapeutic applications in OSCC.
Assuntos
Carcinoma de Células Escamosas/metabolismo , Histona Desacetilases/metabolismo , Neoplasias Bucais/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Adulto , Idoso , Animais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Desacetilase 6 de Histona , Humanos , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Neoplasias Bucais/genética , Neoplasias Bucais/patologia , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Icariin (ICA), a flavanoid isolated from herbal Epimedium, has multiple biological activities. The present study investigated the effects of ICA on the proliferation and alkaline phosphatase (ALP) activity (an index for PDLC differentiation) of human periodontal ligament cells (hPDLCs) inhibited by lipopolysaccharide (LPS). hPDLCs were cultured in vitro and stimulated with various concentrations of ICA. The proliferation ability of hPDLCs was detected by an MTT assay. The activity of ALP was determined by the p-Nitrophenyl phosphate method, and the expression of ALP was analyzed by reverse transcription polymerase chain reaction and western blot analysis. ICA exhibited a dose-dependent effect on the proliferation of hPDLCs in a suitable concentration range, from 10-6 to 10-8 mol/l, and with a mediate optimal concentration (10-6 mol/l). The alkaline phosphatase activity was markedly inhibited in 10 µg/ml LPS-treated PDLCs and this inhibition was suppressed in the presence of icariin at a concentration of 10-6 mol/L following prolonged treatment (96 h). Therefore, this study provided insight into the use of ICA for periapical tissue regeneration.
Assuntos
Fosfatase Alcalina/metabolismo , Diferenciação Celular/efeitos dos fármacos , Flavonoides/farmacologia , Lipopolissacarídeos/farmacologia , Ligamento Periodontal/efeitos dos fármacos , Fosfatase Alcalina/genética , Apoptose/efeitos dos fármacos , Western Blotting , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Medicamentos de Ervas Chinesas , Humanos , Ligamento Periodontal/citologia , Ligamento Periodontal/enzimologia , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
The easily accessible mesenchymal stem cells in the Wharton's jelly of human umbilical cord tissue (hUCMSCs) have excellent proliferation and differentiation potential, but it remains unclear whether hUCMSCs can differentiate into odontoblasts. In this study, mesenchymal stem cells were isolated from the Wharton's jelly of human umbilical cord tissue using the simple method of tissue blocks culture attachment. UCMSC surface marker expression was then evaluated for the isolated cells using flow cytometry. The third-passage hUCMSCs induced by conditioned medium from developing tooth germ cells (TGC-CM) displayed high alkaline phosphatase (ALP) levels (P < 0.001), an enhanced ability to proliferate (P < 0.05), and the presence of mineralized nodules. These effects were not observed in cells treated with regular medium. After induction of hUCMSCs, the results of reverse transcriptional polymerase chain reaction (PCR) indicated that the dentin sialophosphoprotein (DSPP) and dentin matrix protein 1 (DMP1) genes were significantly tested. Additionally, dentin sialoprotein (DSP) and DMP1 demonstrated significant levels of staining in an immunofluorescence analysis. In contrast, the control cells failed to display the characteristics of odontoblasts. Taken together, these results suggest that hUCMSCs can be induced to differentiate into odontoblast-like cells with TGC-CM and provide a novel strategy for tooth regeneration research.
