CHEMISTRY OF COLLAGEN CROSS-LINKS - GLUCOSE-MEDIATED COVALENT CROSS-LINKING OF TYPE-IV COLLAGEN IN LENS CAPSULES

The incubation of lens capsules with glucose in vitro resulted in chan ges in the mechanical and thermal properties of type-IV collagen consi stent with increased cross-linking. Differential scanning calorimetry (d.s.c.) of fresh lens capsules showed two major peaks at melting temp eratures T(m) 1 and T(m) 2 at approx. 54-degrees-C and 90-degrees-C, w hich can be attributed to the denaturation of the triple helix and 7S domains respectively. Glycosylation of lens capsules in vitro for 24 w eeks caused an increase in T(m) 1 from 54-degrees-C to 61-degrees-C, w hile non-glycosylated, control incubated capsules increased to a T(m) 1 of 57-degrees-C. The higher temperature required to denature the typ e-IV collagen after incubation in vitro suggested increased intermolec ular cross-linking. Glycosylated lens capsules were more brittle than fresh samples, breaking at a maximum strain of 36.8 +/- 1.8% compared with 75.6 +/- 6.3% for the fresh samples. The stress at maximum strain (or 'strength') was dramatically reduced from 12.0 to 4.7 N . mm . mg -I after glycosylation in vitro. The increased constraints within the system leading to loss of strength and increased brittleness suggested not only the presence of more cross-links but a difference in the loc ation of these cross-links compared with the natural lysyl-aldehyde-de rived cross-links. The chemical nature of the fluorescent glucose-deri ved cross-link following glycosylation was determined as pentosidine, at a concentration of 1 pentosidine molecule per 600 collagen molecule s after 24 weeks incubation. Pentosidine was also determined in the le ns capsules obtained from uncontrolled diabetics at a level of about 1 per 100 collagen molecules. The concentration of these pentosidine cr oss-links is far too small to account for the observed changes in the thermal and mechanical properties following incubation in viro, clearl y indicating that another as yet undefined, but apparently more import ant cross-linking mechanism mediated by glucose is taking place.