Abstract
As VIP is involved in the intestinal water and electrolyte secretion and a VIP-ergic innervation of the epithelial layer has been demonstrated, we investigated binding, internalization and degredation of VIP by pig jejunum epithelial cells. Enterocytes were isolated by a non-enzymatic dissociation procedure. 125I-VIP binding was time- and temperature dependent and was inhibited dose-depend-ently by VIP(KD=1.5±0.2 nM, mean±SD;n=5) and secretin(half maximal binding 10nM) but not by other peptides. Internalized radioactivity was separated from cell surface-bound radioactivity by washing the cells with isotonic saline, PH2.5.At 37°C the amount of internalized radioactivity increased till 30 min and represented 30% of cell bound radioactivity. At 10°C, all radioactivity bound to the cells was acid dissociable. To investigate degredation of VIP, cells were incubated with VIP(5-20 nmol/1) and the reaction mixture was analysed by HPLC and metabolites were identified by amino acid analysis. After only 30 sec incubation, des(His)VIP(fragment 2-28) was identified as a major metabolite, representing 30% of the substrate In conclusion normal enterocytes internalized VIP but, in view of the rapid cell-surface degradation, it is suggested that VIP is not internalized as intact peptide. As VIP(2-2B) has only 1% of the biological activity of VIP, its action in the gut may be terminated by aminopeptidases.
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