RESUMO
The human kallikrein-related peptidase family is comprised of 15 serine protease genes on chromosome 19q13.4. Our previous microarray analyses showed that the gene kallikrein-related peptidase 13 (KLK13) was down-regulated in oral squamous cell carcinoma (OSCC) cell lines. We evaluated the expression status of KLK13 in primary OSCCs and performed functional molecular experiments in OSCC cell lines. In 102 primary tumors studied, KLK13 expression significantly (P < 0.05) decreased compared with matched normal counterparts. Interestingly, KLK13-negative cases correlated significantly (P < 0.05) with regional lymph node metastasis. In vitro, cells overexpressing KLK13 (oeKLK13) had decreased invasiveness and motility and up-regulation of adhesion molecules (E-cadherin, α-catenin, ß-catenin, junction plakoglobin, plakophilin4, desmocollin2, desmoglein3, and desmoplakin) compared with control cells. A rescue experiment that transfected oeKLK13 cells with siRNA against KLK13 restored invasiveness and migration activities with down-regulated adhesion molecules. Based on our results, we concluded that KLK13 may play an important role in regulating cellular migration and invasiveness, making the loss of KLK13 a potential biomarker for early detection of lymph node metastasis in OSCCs.
Assuntos
Biomarcadores Tumorais/biossíntese , Carcinoma de Células Escamosas/enzimologia , Carcinoma de Células Escamosas/patologia , Moléculas de Adesão Celular/biossíntese , Calicreínas/biossíntese , Neoplasias Bucais/enzimologia , Neoplasias Bucais/patologia , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células , Regulação para Baixo , Humanos , Calicreínas/genética , Metástase Linfática , Invasividade Neoplásica , Interferência de RNA , RNA Interferente Pequeno/genéticaRESUMO
PURPOSE: Tripeptidyl peptidase II (TPP2), a member of the family of eukaryotic serine peptidase, has been implicated in DNA repair, cellular division, and apoptosis. The aim of this study was to examine TPP2 expression and its functional mechanisms in oral squamous cell carcinoma (OSCC). METHODS: TPP2 mRNA and protein expression in seven OSCC-derived cells (Ca9-22, HSC-2, HSC-3, HSC-4, HO-1-N-1, H1, and Sa3) was analyzed by quantitative reverse transcriptase-polymerase chain reaction and immunoblotting analyses. Since previous studies indicated that TPP2 might control chromosomal division, we investigated cellular proliferation and spindle assembly checkpoint (SAC) molecules, MAD2 and CCNB1. In addition, we evaluated the correlation between TPP2 expression levels in primary OSCCs (n = 108 specimens) and the clinicopathologic status by immunohistochemistry (IHC). RESULTS: TPP2 mRNA and protein were significantly (P < 0.05) up-regulated in OSCC-derived cells compared with human normal oral keratinocytes. Suppression of TPP2 expression with shRNA significantly (P < 0.05) inhibited cellular proliferation compared with the control cells. In addition, appropriate localization of MAD2 and up-regulation of CCNB1 were observed in TPP2 knockdown OSCC cells. IHC showed that TPP2 expression in primary OSCCs was significantly (P < 0.001) greater than that in the normal oral counterparts, and the TPP2-positive cases were significantly (P < 0.05) correlated with tumor size. CONCLUSION: The current study showed that overexpression of TPP2 occurs frequently during oral carcinogenesis and might be associated with OSCC progression via SAC activation.
Assuntos
Aminopeptidases/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Carcinoma de Células Escamosas/enzimologia , Proteínas de Ciclo Celular/metabolismo , Ciclina B1/metabolismo , Dipeptidil Peptidases e Tripeptidil Peptidases/metabolismo , Pontos de Checagem da Fase M do Ciclo Celular , Neoplasias Bucais/enzimologia , Proteínas Repressoras/metabolismo , Serina Endopeptidases/metabolismo , Idoso , Aminopeptidases/genética , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Proliferação de Células , Dipeptidil Peptidases e Tripeptidil Peptidases/genética , Regulação para Baixo/efeitos dos fármacos , Feminino , Imunofluorescência , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Immunoblotting , Japão , Metástase Linfática , Proteínas Mad2 , Masculino , Pessoa de Meia-Idade , Neoplasias Bucais/metabolismo , Neoplasias Bucais/patologia , Estadiamento de Neoplasias , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Serina Endopeptidases/genética , Transfecção , Regulação para CimaRESUMO
We previously reported that human squamous cell carcinoma (SCC) cell lines refractory to cis-diaminedichloro-platinum II (cisplatin [CDDP]) had significant upregulation of the phosphodiesterase 3B gene (PDE3B), suggesting that inhibiting PDE3B suppresses CDDP resistance. shRNA-mediated PDE3B depletion in CDDP-resistant cells derived from SCC cells and Hela cells and induced CDDP sensitivity and inhibited tumor growth with elevated cyclic GMP induction resulting in upregulation of the multidrug-resistant molecule, but this did not occur in the 5-fluorouracil-resistant hepatocellular carcinoma cell lines. Furthermore, the antitumor growth effect of the combination of a PDE3B inhibitor (cilostazol) and CDDP in vivo was also greater than with either cilostazol or CDDP alone, with a significant increase in the number of apoptotic and cell growth-suppressive cancer cells in CDDP-resistance cell lines. Our results provided novel information on which to base further mechanistic studies of CDDP sensitization by inhibiting PDE3B in human cancer cells and for developing strategies to improve outcomes with concurrent chemotherapy.
Assuntos
Carcinoma de Células Escamosas/tratamento farmacológico , Cisplatino/farmacologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 3/biossíntese , Inibidores da Fosfodiesterase 3/farmacologia , Neoplasias do Colo do Útero/tratamento farmacológico , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Apoptose/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Cilostazol , Cisplatino/administração & dosagem , AMP Cíclico/metabolismo , GMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 3/genética , Nucleotídeo Cíclico Fosfodiesterase do Tipo 3/fisiologia , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feminino , Regulação Enzimológica da Expressão Gênica , Inativação Gênica , Células HeLa , Humanos , Camundongos , Camundongos Nus , Inibidores da Fosfodiesterase 3/administração & dosagem , RNA Mensageiro/genética , Tetrazóis/administração & dosagem , Tetrazóis/farmacologia , Células Tumorais Cultivadas , Neoplasias do Colo do Útero/metabolismo , Neoplasias do Colo do Útero/patologia , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Salivary α-amylase is the most important enzyme for oral digestion of dietary starch. Therefore, regeneration of the salivary glands via a tissue engineering approach is clearly required for patients with salivary gland dysfunction. Early during seed germination, the embryo synthesizes gibberellic acid (GA3), a plant hormone that activates the synthesis and secretion of α-amylase. The purpose of this study was to explore an approach for differentiation of stem cells into salivary glands using GA3. We isolated adipose-derived stem cells (ASCs), which are positive for mesenchymal stem cell markers (CD73, CD90 and CD105) and possess pluripotency to osteoblasts, adipocytes and neural cells, from human buccal fat pads, which are a readily available source for dentists and oral surgeons. In addition, we investigated the cytotoxicity of GA3 for human ASCs. GA3 neither affects cell morphology nor cell viability in a dose- or time-dependent manner. ASCs were incubated with GA3 to assess mRNA and protein expression of α-amylase by reverse transcriptase-polymerase chain reaction and western blot analyses. α-amylase mRNA expression on 21 days after treatment with GA3 (1 mM) was seven times greater than that in resting condition (Day 0). While we did not detect α-amylase bands on Day 0, α-amylase protein was detectable 7 days after treatment with GA3, reaching a maximal level on Day 21. Our results indicated that GA3 can increase cellular α-amylase expression and that our induction method would be useful for therapeutic application for salivary gland regeneration.
Assuntos
Tecido Adiposo/metabolismo , Giberelinas/farmacologia , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , alfa-Amilases/metabolismo , Tecido Adiposo/citologia , Diferenciação Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Expressão Gênica , Giberelinas/toxicidade , Humanos , RNA Mensageiro/metabolismo , Células-Tronco/citologia , alfa-Amilases/genéticaRESUMO
Heat shock factor 1 (HSF1) is responsible for expres-- sion of a large class of heat shock proteins that have been implicated in the malignant phenotype of human cancers. Little is known about the effect of a high level of HSF1 on the behavior of oral squamous cell carcinoma (OSCC). In this study, we assessed the value of HSF1 for predicting clinical outcomes in OSCC. Quantitative reverse transcriptase-polymerase chain reaction and Western blotting showed that the expressions of HSF1 mRNA and protein in OSCC-derived cell lines (HSC-2, HSC-3, HSC-4, Sa3, Ca9-22, KON and Ho-1-u-1) were elevated compared with those in human normal oral keratinocytes (P<0.05). Similar to in vitro data, HSF1 mRNA expression in primary OSCCs (n=50) was significantly greater than in normal counterparts (P<0.05). Since HSF1 was observed in the nucleus and cytoplasm by immu-- nohistochemistry, we investigated the correlation between the HSF1 expression status at each subcellular location and the clinical behavior of OSCCs. Among the clinical classifications, higher nuclear HSF1 expression was closely related to tumor size and histopathologic types (P<0.05). These results showed for the first time that nuclear HSF1 expression may contribute to cancer progression and that HSF1 might be a potential diagnostic biomarker and a therapeutic target for OSCCs.