Efeitos do anfenaco e de sua combinação com bevacizumabe sobre a secreção de citocinas pró-angiogênicas e inflamatórias pelas células do epitélio pigmentado retiniano

• Autor(a) principal: Miyamoto, Cristina [UNIFESP]
• Data de Publicação: 2015
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: Repositório Institucional da UNIFESP
• Texto Completo: https://sucupira.capes.gov.br/sucupira/public/consultas/coleta/trabalhoConclusao/viewTrabalhoConclusao.jsf?popup=true&id_trabalho=2728322; https://repositorio.unifesp.br/handle/11600/46209
• topic: amfenac; bevacizumab; cytokines; angiogenesis; inflammation; anfenaco; bevacizumabe; citocinas; angiogênese; inflamação

OBJECTIVE: To evaluate the effects of amfenac (the active component of nepafenac) and its combination with bevacizumab on the secretion of pro-angiogenic and inflammatory cytokines by human retinal pigment epithelial cells. Additionally, to evaluate if chemical hypoxia with cobalt chloride, a hypoxia-mimicking agent, produces the same effect of the real hypoxia (hypoxia chamber) on the secretion of these cytokines by human retinal epithelial cells. METHODS: ARPE-19 cells were incubated under normoxia, chemical hypoxia, and real hypoxia. The cells were treated as follows: control, bevacizumab (0,25 mg/mL), amfenac (2 ng/mL), and the combination of bevacizumab (0,25 mg/mL) and amfenac (2,0 ng/mL). Media was harvested after 24h for sandwich ELISA-based angiogenesis and inflammation assays. The secretion of 10 pro-angiogenic cytokines: angiogenin, ANG-2, EGF, bFGF, HB-EGF, PDGF-BB, leptin, PIGF, HGF e VEGF; as well as of 10 inflammatory cytokines: IL-1?, IL-1?, IL-4, IL-6, IL-8, IL-10, IL-13, IFN-?, TNF-?, MCP-1, were measured. The viability/cytotoxicity was also assessed. RESUTS: Under normoxia, amfenac increased the secretion of ANG-2, HB-EGF, leptin, PDGF-BB, and PIGF, almost did not changed the secretion of EGF, and reduced the secretion of angiogenin, bFGF, HGF, and VEGF by ARPE-19 cells; the combination of the drugs elevated the secretion of all pro-angiogenic cytokines, except angiogenin, bFGF, and VEGF, whose secretion was diminished. Under chemical hypoxia, amfenac decreased the secretion of all pro-angiogenic cytokines, except ANG-2, whose secretion was increased, EGF whose secrection was practically unchanged and PIGF whose secretion was not detected by the method after the three treatments; the combination of the drugs reduced the secrection of all pro-angiogenic cytokines, except ANG-2 and HGF whose secretion were raised, and PIGF whose secretion was virtually the same. Under real hypoxia, amfenac increased the secretion of all pro-angiogenic cytokines, except angiogenin, bFGF and VEGF, whose secretion was decreased; the combination of the drugs increased the secretion of all pro-angiogenic cytokines, except VEGF. Under normoxia, amfenac practically did not altered the secretion of IL-1? and IL-13, reduced the secretion of IL-1?, IL-4, IL-8, and IL-10, and increased the secretion of IL-6, MCP-1, and TNF-?; the combination of the drugs virtually did not modified the secretion of IL-1?, IL-8, and IL-10, diminished the secretion of IL-1?, and increased the secretion of the remaining inflammatory cytokines. Under chemical hypoxia, amfenac practically did not changed the secretion of IL-1?, IL-1?, and IFN-?, reduced the secretion of IL-10, IL-13, and TNF-?, and elevated the secretion of all other inflammatory cytokines; the combination of the drugs virtually did not modified the secretion of IL-1?, decreased the secretion of IL-10, IL-13, and TNF-?, and increased the secretion of the remaining inflammatory cytokines. Under real hypoxia, amfenac reduced the secretion of all inflammatory cytokines; the combination of the drugs also decreased the secretion of the inflammatory cytokines apart from IFN-?, whose secretion increased; the combination of the drugs caused a more pronounced reduction in the secretion of the inflammatory cytokines (except MCP-1) compared to bevacizumab. These results were not statistically significant. All the treatments did not reduced the viability of the ARPE-19 cells under normoxia, chemical hypoxia and real hypoxia. Comparing the chemical hypoxia with cobalt chloride and the real hypoxia, the chemical hypoxia produced statistically different results regarding the secretion of 8 out of 10 pro-angiogenic cytokines (except EGF and HB-EGF), and the secretion of two inflammatory cytokines (IL-6 and IL-8). CONCLUSIONS: Amfenac reduced the secretion of pro-angiogenic cytokines (angiogenin, bFGF, and VEGF) and the secretion of all inflammatory cytokines under real hypoxia. Besides, except from MCP-1 and IFN-?, the combination of amfenac with bevacizumab tends to cause higher reduction of the levels of inflammatory cytokines secreted by ARPE-19 cells under real hypoxia than bevacizumab alone. Nepafenac may be a worth therapy for exudative AMD in combination with bevacizumab with additional benefits like the reduction of the frequency of intravitreal injections of anti-VEGF. Further studies are necessary to confirm this hypothesis. Compared with real hypoxia, the chemical hypoxia produced different results, sometimes statistically significant, regarding the levels of the pro-angiogenic and inflammatory cytokines secreted by ARPE-19 cells. This suggests that some cytokines function primarily associated with HIF-1?, while others do not.