摘要:

Our goal was to determine if the major endogenous vitreous matrix metalloproteinase (MMP-2) could digest known collage-nous components of the vitreous body. Matrix metalloproteinase-2 and its associated inhibitors were isolated from porcine vitreous by affinity column chromatography. The inhibitors were inactivated by chemical modification with dithiothreitol and iodoaceta-mide. The latent MMP-2 was then activated with the organo-mercurial, p-aminophenyl mercuric acetate (APMA). Bovine vitreous fibrillar collagens (types II, V/XI and IX) were isolated by pepsin extraction and differential salt precipitation. Intact type IX collagen was purified by selective salt precipitation followed by ion exchange and size exclusion chromatography. These isolated collagens were incubated for 6 to 24 h with different concentrations of activated MMP-2, and the extent of collagen degradation was analyzed. Activated MMP-2 was also introduced into freshly isolated vitreous gels and the degree of liquefaction was determined. Our results showed that the activated MMP-2 has no apparent effect upon type II collagen but can degrade type V/XI collagen and type IX collagen fragments (COL2 and COL2 + COL3). In addition, when the type IX collagen was in the intact helical form, MMP-2 appeared to selectively digest a3 (IX) chains. This suggested that vitreous MMP-2 preferentially cleaved certain vitreous collagen chains into large fragments rather than small peptides. MMP-2 also disrupted the vitreous gelin vitro, releasing proteins but not hexuronic acid or sulfated glycosaminoglycans into the liquefied supernatant. We conclude that MMP-2 activity should be considered as a potential mechanism of vitreous liquefaction that is seen in aging and various pathological states.

ISSN:0271-3683    

DOI:10.3109/02713689608995835

出版商:Taylor&Francis    

年代:1996

数据来源: Taylor

摘要:

The objective of this study was to determine whether Na+-glucose cotransport contributed to the residual short-circuit current (Isc) in the pigmented rabbit conjunctiva not accounted for by CI- secretion. The Iscof the pigmented rabbit conjunctiva was measured following exposure of its mucosal or serosal side to varying concentrations of D-glucose or phloridzin in a glutathione-bicarbonated Ringer's solution (GBR). Addition of D-glucose to the mucosal side of the conjunctiva bathed in glucose-free GBR elevated Iscup to 26±6% in a dose-dependent manner (K0.5= 1.2 mM), while mucosal (but not serosal) addition of phloridzin reduced the Isc(IC50= 0.05 mM) of the conjunctiva bathed in regular GBR. In a mucosal Na+-free medium, neither 20 mM D-glucose nor 0.5 mM phloridzin was effective. In a mucosal glucose-containing medium, deletion of Nafreduced Iscup to 27±4%. It was, however, restored to its initial value upon the addition of 17.5–141 mM Na+to a medium containing 5 mM D-glucose. Hill plot analysis of I., elevation at 141 mM Na+in the presence of varying D-glucose concentrations in the mucosal bathing fluid yielded a Hill coefficient of 0.86. There is, therefore, evidence for the apical localization of phloridzin-sensitive Na+-coupled D-glucose transport that exhibits apparent 1:1 stoichiometry, being responsible for the maximal 26% increase in the Isc.

ISSN:0271-3683    

DOI:10.3109/02713689608995836

出版商:Taylor&Francis    

年代:1996

数据来源: Taylor