Evidence for secretion of high molecular weight mucins by canine tracheal epithelial cells in primary culture: Effects of select secretagogues on mucin secretion

Abstract

The purpose of this investigation was to provide evidence for the secretion of high molecular weight mucins, CTM-A and CTM-B, in primary culture of canine tracheal epithelial (CTE) cells. The cells were isolated from tracheas of mongrel dogs by pronase treatment. Primary cultures of the epithelial cells were established using ICN collagen inserts in Dulbecco's modified Eagle's/F12 medium supplemented with growth factors and could be maintained for up to 23 days. The evidence for the mucin secretion in culture medium and their localization in the cells was established by a) positive immunocytochemical staining using specific antibodies developed against purified native as well as deglycosylated CTM-A and CTM-B; b) incorporation of labeled amino acids, followed by electrophoresis and autoradiography detection of glycoconjugates purified from the culture medium; c) comparison of the amino acid compositions of mucin purified from canine tracheal pouch secretions and that purified from the culture medium; and d) Western blot analyses using specific polyclonal antibodies directed against deglycosylated CTM-A and CTM-B. Immunoaffinity purified secreted labeled glycoconjugates were resistant to hyaluronidase treatment. The effects of cyclic AMP (1 x 10(-5) M), dibutyryl cyclic AMP (1 x 10(-5) M), 8-bromocyclic AMP (1 x 10(-5) M), and prostaglandin E1 (1 x 10(-6) M) on mucin secretion by CTE cells were also investigated. Secretion of mucins by CTE cells in culture was considerably more enhanced by 8-bromocyclic AMP than that observed for other secretagogues used in this study.

Effects of In Vitro Production on Horse Embryo Morphology, Cytoskeletal Characteristics, and Blastocyst Capsule Formation

“…In this latter respect, it is likely that the absolute production of capsular glycoproteins by IVP embryos was relatively low, due to their low cell numbers, and it is also possible that the ICSIderived hole in the ZP would have hindered the accumulation of the glycoproteins. In addition, there are comparable reports of reduced secretion of other high-molecularweight mucins in culture, such that they fail to assemble into a mucous layer [64].…”