Avaliação de alternativas ao uso do soro fetal bovino na adaptação de cultivos celulares e obtenção e caracterização das células de córnea bovina (CORBOV

Abstract

Media for cell culture, with different nutritional intakes were created and improved over the years seeking to provide the conditions for in vitro cell proliferation. The serum of animal origin (SA) is commonly added to the medium as the main source of hormones, such as growth factors. The soy hydrolyzate and the medium Virus Production Serum Free Medium (VP-SFM), that do not use any input of animal or human origin were tested as substitutes to fetal bovine serum (FBS) in adaptation of VERO and MDBK cells. In both media, the cells showed the same pattern of cell proliferation compared to the medium supplemented with SA.Bovine corneal cells (CorBov) were obtained from primary culture and maintained for twenty-four passages. The cells retained their morphology and proliferation capacity after recovery from liquid nitrogen (-196ºC) storage. The CorBov cells presented elongated and elliptical core morphology, features present in fibroblasts. In the standard cell proliferation assessment there was a gradual increase in the cell number over 30 days even when the culture medium was not replaced. It was possible to maintain CorBov cells throughout this period with Minimal Essential Medium (MEM) supplemented with one, two, five or ten percent of SA. The CorBov cell population was positively marked by vimentin in Immunocytochemistry assay, demonstrating its mesenchymal origin. Corneal cells were permissible to Bovine Viral Diarrhea Virus (BVDV) and Herpesvirus Type I (BoHV-1) evidenced by the presence of cytopathic effect such as cell lysis and cytoplasmic extensions; and spherical cell structures forming lumps bunch of grapes, respectively for each virus. The BoHV-1 showed the same pattern of virus replication in CorBov when compared to the standard (control) MDBK cells