RESUMO
OBJECTIVES: Verify the in-vitro effect of triiodothyronine (T3) on the chondrogenic differentiation of female rat bone marrow mesenchymal stem cells (BMMSCs) over several time periods and at several doses. METHODS: CD54 + /CD73 + /CD90 + BMMSCs from Wistar female rats were cultured in chondrogenic medium with or without T3 (0.01; 1; 100; 1000 nm). At seven, 14 and 21 days, the cell morphology, chondrogenic matrix formation and expression of Sox9 and collagen II were evaluated. KEY FINDINGS: The dose of 100 nm did not alter the parameters evaluated in any of the periods studied. However, the 0.01 nm T3 dose improved the chondrogenic potential by increasing the chondrogenic matrix formation and expression of Sox9 and collagen II in at least one of the evaluated periods; the 1 nm T3 dose also improved the chondrogenic potential by increasing the chondrogenic matrix formation and the expression of collagen II in at least one of the evaluated periods. The 1000 nm T3 dose improved the chondrogenic potential by increasing the chondrogenic matrix formation and Sox9 expression in at least one of the evaluated periods. CONCLUSIONS: T3 has a dose-dependent effect on the differentiation of BMMSCs from female rats.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Condrogênese/efeitos dos fármacos , Células-Tronco Mesenquimais/efeitos dos fármacos , Tri-Iodotironina/farmacologia , Animais , Células Cultivadas , Condrócitos/citologia , Colágeno Tipo II/genética , Relação Dose-Resposta a Droga , Feminino , Células-Tronco Mesenquimais/citologia , Ratos , Ratos Wistar , Fatores de Transcrição SOX9/genética , Fatores de Tempo , Tri-Iodotironina/administração & dosagemRESUMO
AIMS: Polymorphisms in cell cycle genes are considered prognostic as radiosensitivity markers in patients with head and neck squamous cell carcinoma. Therefore, we aimed to investigate the relationship of ATM 5557G>A, ATM IVS62 + 60G>A, TP53 215G>C, BCL2-938C>A, TGFß-509C>T, and TGFß 29C>T with radiotherapy response. MATERIALS AND METHODS: Genotyping was performed by polymerase chain reaction followed by restriction fragment length polymorphism in 210 patients with oral cavity/oropharyngeal carcinoma and 101 patients with laryngeal tumors. RESULTS: In irradiated oral cavity/oropharyngeal tumors, the ATM IVS62 + 60G>A AA genotype significantly increased local recurrence risk (odds ratio [OR] = 4.43; confidence interval [CI] = 1.22-16.13) and the BCL2-938C>A C allele and the TGFß-509C>T T allele were associated with worse disease-specific survival (hazard ratio [HR] = 0.46; CI = 0.24-0.90 and HR = 2.20; CI = 1.12-4.29, respectively). In irradiated laryngeal carcinoma, the TGFß 29C>T C allele was associated with increased local recurrence risk (OR = 0.09; CI = 0.02-0.53), death rate (OR = 0.18; CI = 0.04-0.86), and worse local disease-free and disease-specific survival rates (HR = 0.13; CI = 0.03-0.59 and HR = 0.21; CI = 0.07-0.60, respectively), while the BCL2-938C>A C allele was related to a worse disease-specific survival (HR = 0.32; CI = 0.12-0.83). DISCUSSION: These results can help individualize treatment according to a patient's genetic markers. We demonstrated that ATM IVS62 + 60G>A, TGFß 29C>T, TGFß-509C>T, and BCL2-938C>A can function as biomarkers of tumor radiosensitivity, being candidates for a predictive genetic profile of radiotherapy response.
Assuntos
Biomarcadores Tumorais/genética , Neoplasias de Cabeça e Pescoço/genética , Adulto , Idoso , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Carcinoma de Células Escamosas/genética , Feminino , Genótipo , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Polimorfismo de Nucleotídeo Único/genética , Prognóstico , Regiões Promotoras Genéticas/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Radioterapia , Taxa de Sobrevida , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo , Resultado do TratamentoRESUMO
Head and neck squamous cell carcinoma (HNSCC) is a heterogeneous disease affecting the epithelium of the oral cavity, pharynx and larynx. Conditions of most patients are diagnosed at late stages of the disease, and no sensitive and specific predictors of aggressive behavior have been identified yet. Therefore, early detection and prognostic biomarkers are highly desirable for a more rational management of the disease. Hypermethylation of CpG islands is one of the most important epigenetic mechanisms that leads to gene silencing in tumors and has been extensively used for the identification of biomarkers. In this study, we combined rapid subtractive hybridization and microarray analysis in a hierarchical manner to select genes that are putatively reactivated by the demethylating agent 5-aza-2'-deoxycytidine (5Aza-dC) in HNSCC cell lines (FaDu, UM-SCC-14A, UM-SCC-17A, UM-SCC-38A). This combined analysis identified 78 genes, 35 of which were reactivated in at least 2 cell lines and harbored a CpG island at their 5' region. Reactivation of 3 of these 35 genes (CRABP2, MX1, and SLC15A3) was confirmed by quantitative real-time polymerase chain reaction (PCR; fold change, >or=3). Bisulfite sequencing of their CpG islands revealed that they are indeed differentially methylated in the HNSCC cell lines. Using methylation-specific PCR, we detected a higher frequency of CRABP2 (58.1% for region 1) and MX1 (46.3%) hypermethylation in primary HNSCC when compared with lymphocytes from healthy individuals. Finally, absence of the CRABP2 protein was associated with decreased disease-free survival rates, supporting a potential use of CRABP2 expression as a prognostic biomarker for HNSCC patients.
