Properties of two hydrolases: beta-glucuronidase and N-acetyl-beta-hexosaminidase were studied in lysosomal fractions of media-intima from arterial wall. These enzymatic activities change significantly with aging. In the arterial wall, the decrease in activities of beta-glucuronidase and N-acetyl-beta-hexosaminidase between young and old rats was highly significant while there no notable change in the activities of acid phosphatase. These data are in agreement with the metabolic slow-down and the modifications of the glycosaminoglycan distribution in the media-intima of arterial wall with aging.
We recently reported that cultured gland serous cells release chondroitin sulfate proteoglycans (CSPGs) in response to beta-adrenergic agonists. In this study, we analyzed this regulatory pathway and other cellular mechanisms responsible for CSPG secretion. We show the following. 1) Isoproterenol increased CSPG secretion in a concentration-dependent manner, with maximal stimulation (50%) obtained at 10(-5) M; at this concentration, the beta-agonist also stimulated protein kinase A (PKA) by 50%, whereas it increased cellular adenosine 3',5'-cyclic monophosphate (cAMP) content by 300%. 2) Phenylephrine (10(-5) M), 4 beta-phorbol 12 beta-myristate 13 alpha-acetate (1.6 x 10(-7) M), and A23187 (10(-6) M) also stimulated CSPG secretion; this stimulation was concomitant with protein kinase C (PKC) translocation from cytosol to membrane, was blocked by sphingosine (2 x 10(-5) M), and was additive with that elicited by isoproterenol. 3) All PKC activators potentiated the isoproterenol-induced increased in cAMP accumulation without modifying the activation of PKA elicited by the beta-agonist. Our results indicate that although the signaling pathways triggered by alpha- and beta-adrenergic agonists converge at the level of adenylate cyclase in tracheal serous cells, PKA and PKC independently regulate CSPG secretion.
In comparison to skin fibroblasts from normal subjects, those from patients with cystic fibrosis (CF): (1) bound [20-3H] phorbol 12,13-dibutyrate (PDBu) with a higher affinity (Kd=25.8 vs 12.8 nM respectively) but expressed a similar number of total phorbol ester binding sites (about 2.5 pmol PDBu bound/mg of protein); (2) exhibited a faster and higher response to 4?-phorbol 12?-myristate 13?-acetate (PMA) for the stimulation of [35S]-labelled glycoconjutate release, but were equally sensitive to the synergistic effect of A23187 on this process; and (3) secreted glycoconjugates with similar [35S]-sulfate and [14C]-leucine to [14C]-glucosamine labelling ratios. Taken together, these results provide further evidence for abnormal protein kinase C (PKC) regulation of macromolecule secretion in CF disease.
