Efeito do orlistat sobre a proliferação e apoptose de células derivadas de carcinoma espinocelular bucal humano

A enzima ácido graxo sintase (FASN) desempenha papel chave na lipogênese de células neoplásicas e apresenta alta expressão em vários tumores malignos humanos. Um grande número de evidências sugere que FASN seja um oncogene metabólico com papel importante no crescimento e sobrevivência de células tumorais. Diferentes inibidores da atividade de FASN têm sido utilizados, como a droga Orlistat, a qual demonstrou efeito anti-tumoral em algumas neoplasias, incluindo a redução em até 50% no número de metástases experimentais em melanomas. O objetivo deste trabalho foi avaliar as conseqüências do tratamento com Orlistat sobre proliferação e apoptose das linhagens celulares derivadas de CECs bucais humanos SCC-9 e SCC-25. As células foram tratadas com concentrações de 50, 100 e 200 uM de Orlistat na presença de 10% de soro fetal bovino pelos períodos de 12, 24, 48 e 72 horas, sendo então marcadas com anexina V para avaliação das taxas de apoptose ou com iodeto de propídio para análise do ciclo celular. O efeito do Orlistat sobre o ciclo celular foi dose dependente, ocorrendo uma inibição de aproximadamente 70% da fase S para as duas linhagens após 72 horas de tratamento na concentração de 200 uM. Entretanto, não observamos aumento significativo dos níveis de apoptose, como ocorre em células derivadas de outros tipos de tumores. Concluímos que o uso do inibidor da atividade de FASN Orlistat em células de CECs bucais humanos causa inibição significativa da progressão do ciclo celular, o que faz desta enzima um alvo terapêutico em potencial para estes tumores.The enzyme fatty acid synthase (FASN) plays a key role in lipogenesis in tumor cells and shows high expression in several human malignancies. A growing body of evidence suggests that FASN is a metabolic oncogene plays a major role in the growth and survival of tumor cells. Different activity of FASN inhibitors have been used as the drug Orlistat, which demonstrated anti-tumor effect in some cancers, including reducing by 50% in the number of metastases in experimental melanomas. The aim of this study was to evaluate the consequences of treatment with Orlistat on proliferation and apoptosis of cell lines derived from human oral SCC SCC-9 and SCC-25. Cells were treated with concentrations of 50, 100 and 200 uM of Orlistat in the presence of 10% fetal bovine serum for periods of 12, 24, 48, 72 hours, and then stained with annexin V to assess the rates of apoptosis or propidium iodide for cell cycle analysis. The effect of Orlistat on the cell cycle was dose-dependent inhibition occurring at approximately 70% of S phase for both strains after 72 hours of treatment at a concentration of 200 uM. However, we observed no significant increase in levels of apoptosis, as occurs in cells derived from other tumor types. We conclude that use of the FASN inhibitor Orlistat activity in cells of human oral SCCs cause significant inhibition of cell cycle progression, which makes this enzyme a potential therapeutic target for these tumors

Pro-inflammatory Analysis of Macrophages in Contact with Titanium Particles and Porphyromonas gingivalis

<div><p>Abstract During insertion of titanium dental implants, particles may shear from the implant to the periimplant region causing osteolysis, and their association with bacteria can exacerbate the inflammatory reaction. However, the association of a high invasive bacterium from the oral cavity, Porphyromonas gingivalis (Pg), and titanium particles remains unknown. This study evaluated pro-inflammatory reaction of human macrophages in contact with micro and nanoparticles of titanium associated with Porphyromonas gingivalis lipopolysaccharide (PgLPS). THP-1 cell were used and treated for 12, 24 and 48 h following 6 groups: Control(C), PgLPS (L); Microparticles (M); Nanoparticles (N); PgLPS and microparticles (LM); PgLPS and nanoparticles (LN). The following assays were carried out: i) cell viability using MTS, ii) cell morphology by SEM and iii) expression of tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β) and interleukin-6 (IL-6) by qRT-PCR and ELISA. For statistics two-way ANOVA followed by Tukey’s test was used (p<0.05). After treatment, cells presented similar viability and morphology demonstrating that the treatments were not able to induce cell death. Gene expression was significantly higher for TNF-α and IL1-β after 12 h, and for IL-6 after 24 h in the N and LN groups. Cytokine production over time was an ascending curve for TNF-α with the peak at 48 h and IL1-β and IL-6 had a straight line among the time points, although cells from N group presented a significant production of IL-6 at 48 h. In conclusion, these results suggest that titanium nanoparticles stimulate stronger pro-inflammatory response in macrophages, independent of their association with LPS from P.gingivalis.</p></div