Differential Vitreous Dye Diffusion Following Microplasmin or Plasmin Pre-Treatment

Abstract

Purpose: Plasmin and microplasmin are related enzymes that differ mainly in size. The differential effect of plasmin and microplasmin on vitreous structure, protein degradation, and dye diffusion through porcine vitreous was evaluated.

Methods: The enzymatic effect was examined using a number of approaches on fresh porcine eyes: (1) structural integrity of vitreous after a 2-hr incubation using the electron microscope (EM); (2) effect on soluble proteins within the vitreous using gel electrophoresis after incubation at various time points over a 24-hr period; (3) fluorescein dye diffusion within the vitreous cavity measured over a 1-hr period following a 2-hr incubation. The chosen enzymatic activities for plasmin 0.5 IU and microplasmin 125 µg were within the clinical range, and were chosen for equipotence. A saline control was also used in all experiments.

Results: Significant structural changes were seen with both microplasmin and plasmin when examined by EM. Gel electrophoresis showed that microplasmin and plasmin digested the same proteins, mainly molecular weights above 50 kDa. The enzymatic effect was noticeable earlier in microplasmin-treated eyes and was more significant by the end of the incubation period. Differential fluorescein diffusion rates were seen between normal saline, plasmin, and microplasmin within the vitreous cavity. The greatest diffusion rate was seen with microplasmin and was statistically significantly higher than plasmin.

Conclusion: Microplasmin and plasmin have a similar enzymatic effect on vitreous. However, an equipotent amount of microplasmin appears to have a more extended effect on vitreous gel. This may, in part, be related to its smaller size allowing it to diffuse more readily through the vitreous matrix.

KEYWORDS::

• Fluorescein diffusion
• Gel electrophoresis
• Microplasmin
• Pharmacologic vitreolysis
• Plasmin

Declaration of interest: ThromboGenics NV, Leuven, Belgium, provided the microplasmin used for the experiments as well as some technical assistance in producing the gels. They did not influence in any way the content of the article. The authors report no conflict of interest. The authors alone are responsible for the content and writing of the paper.