Résumé
Em mamíferos a perda das células ciliadas determina perda auditiva neurosensorial permanente, já que estas células encontram-se em diferenciação terminal e seus precursores não mais entram no ciclo celular...
Loss of hair cells in mammals causes permanent sensorineural hearing loss, as these cells are terminally-differentiated and their precursors do not reenter the cell cycle. The aims of this study were to establish primary cell cultures and subcultures of organ of Corti...
Sujets)
Connexines/classification , Oreille , Myosines/classification , Techniques de culture cellulaire , Cycle cellulaire , Cochlée , ÉpithéliumRésumé
Vertebrate gap junctions are aggregates of transmembrane channels which are composed of connexin (Cx) proteins encoded by at least fourteen distinct genes in mammals. Since the same Cx type can be expressed in different tissues and more than one Cx type can be expressed by the same cell, the thorough identification of which connexin is in which cell type and how connexin expression changes after experimental manipulation has become quite laborious. Here we describe an efficient, rapid and simple method by which connexin type(s) can be identified in mammalian tissue and cultured cells using endonuclease cleavage of RT-PCR products generated from "multi primers" (sense primer, degenerate oligonucleotide corresponding to a region of the first extracellular domain; antisense primer, degenerate oligonucleotide complementary to the second extracellular domain) that amplify the cytoplasmic loop regions of all known connexins except Cx36. In addition, we provide sequence information on RT-PCR primers used in our laboratory to screen individual connexins and predictions of extension of the "multi primer" method to several human connexins