Desenvolvimento de sistemas celulares de peixe adequados ao estudo da mineraliz...

• Author(s): Marques, C. L. ; Rafael, M. S. ; Tiago, Daniel M. ; Cancela, Leonor ; Laizé, Vincent
• Date: 2009
• Persistent ID: http://hdl.handle.net/10400.1/4289
• Origin: Sapientia - Universidade do Algarve
• Subject(s): Sistemas celulares in vitro; Mineralização; Tecidos calcificados; Dourada (Sparus aurata L.)
• Document Type: Article
• Language: Portuguese

Description

Os peixes foram recentemente reconhecidos como organismos modelo apropriados para o estudo da biologia de vertebrados, particularmente de processos relacionados com a mineralização tecidular e o desenvolvimento do esqueleto. Apesar de existirem alguns estudos in vivo, a identificação de mecanismos associados à calcificação em peixes tem sido prejudicada pelo facto de não existirem sistemas celulares para estudos in vitro. Este artigo descreve um protocolo simples e de baixo custo adequado ao desenvolvimento de culturas celulares mineralogénicas, derivadas de tecidos calcificados de peixes. Fish have been recently recognized as a suitable model organism to study vertebrate biology, in particular physiological processes such as tissue mineralization and skeletal development. Despite various studies in vivo, identification of mechanisms associated with calcification in fish has been largely hampered by the lack of suitable in vitro cell systems. We describe here a simple and inexpensive protocol to develop mineralogenic cell cultures from fish calcified tissues. This protocol was successfully used to develop the first fish cell lines capable of mineralizing their extracellular matrix (from the vertebra of gilthead seabream Sparus aurata) and, more recently, applied to develop additional fish mineralogenic cell cultures from different fishes. We also describe a variety of biochemical and histological methods to characterize extracellular matrix during in vitro mineralization, in particular mineral deposition, and a protocol to enhance DNA transfer into fish bone-derived cells. Finally, we present recent expression data obtained using these cell lines to further demonstrate their suitability to study mechanisms of in vitro mineralization.