RESUMO
PURPOSE: This paper deals with the capability of the ciliary epithelium to express neurolysin, involved in the inactivation of numerous neuropeptides. METHODS: Total RNAs from ciliary body (CB) were processed for RT-PCR, and the amplification products were sequenced. The whole-protein extracts of CBs were analyzed using the Western blot. The CBs were processed for neurolysin immunolocalization. RESULTS: The RT-PCR detected the presence of neurolysin mRNA in the ciliary body. The Western blot assays demonstrated immunochemical cross-reactivity with neurolysin. The immunoreactivity to neurolysin was observed in ciliary epithelium. CONCLUSIONS: These results indicate that the ciliary epithelium expresses neurolysin.
Assuntos
Corpo Ciliar/metabolismo , Metaloendopeptidases/metabolismo , Animais , Western Blotting , Bovinos , Corpo Ciliar/citologia , Reações Cruzadas , Células Epiteliais/metabolismo , Epitélio/metabolismo , Imunoquímica , Masculino , Metaloendopeptidases/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Distribuição TecidualRESUMO
PURPOSE: To verify the capability of rabbit and rat ciliary body to synthesize and secrete ceruloplasmin. METHODS: Isolated ciliary body (CB) was cultured in the presence of [35S]-methionine, and the incubation medium was processed for immunoprecipitation. Total RNA from CB was processed for RT-PCR, and the amplification products were sequenced. Also, sections of CB were immunostained for the localization of ceruloplasmin. RESULTS: A labeled peptide, having a molecular weight of about 135 kDa, the expected size of ceruloplasmin, was immunopurified in the incubation media from both animal species. The RT-PCR and sequencing experiments detected the presence of ceruloplasmin mRNA in rat samples. Both layers of rabbit and rat ciliary epithelium (CE) exhibited ceruloplasmin reactivity after immunohistochemical processing. CONCLUSIONS: Taken altogether, these results indicate the CB, particularly its epithelium, as one of the possible sources of the ocular intrinsic ceruloplasmin.
Assuntos
Ceruloplasmina/genética , Ceruloplasmina/metabolismo , Corpo Ciliar/enzimologia , Animais , Técnicas Imunoenzimáticas , Imunoprecipitação , Masculino , Técnicas de Cultura de Órgãos , RNA Mensageiro/metabolismo , Coelhos , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transferrina/genéticaRESUMO
It has been shown that the vitreous contains several intrinsic glycoproteins whose origin remains to be clarified. Isolated ciliary epithelium (CE) was assayed to verify its role in the synthesis and secretion of transferrin for the vitreous body. It was cultured in the presence of [35S]methionine and the incubation medium was processed for immunoprecipitation. Total RNA from CE was processed for RT-PCR and the amplification products were sequenced. Also, whole preparations of isolated CE were processed for immunolocalization of transferrin. From the incubation assays, a labeled peptide of about 80 kDa was immunopurified that is the expected size of transferrin. The RT-PCR and sequencing experiments detected the presence of transferrin mRNA. Both layers of the CE exhibited transferrin reactivity, following immunohistochemical processing. Taken altogether, these results indicate the CE as one of the possible sources of vitreous intrinsic transferrin.