Effects and Safety of Subtilisin NAT (Nattokinase) as a Novel Enzyme for Pharmacological Vitrectomy

Abstract

Abstract: : Purpose: Subtilisin NAT (nattokinase) is a serine protease that is produced by Bacillus subtilis (natto). Our study was conducted in an attempt to investigate the effects and safety of intravitreal injection of this enzyme, for induction of posterior vitreous detachment (PVD). Methods: Subtilisin NAT was purified from NSK–SD obtained from Japan Bio Science Laboratory Co., Ltd (JBSL; Oita, Japan) and injected into the vitreous in rabbit eyes. Fellow eyes were injected with balanced salt solution (BSS) as control experiments. Fibrin degradation assay was conducted to compare the enzymatic activities of subtilisin NAT in comparison with plasmin. B–mode ultrasonography and electroretinography (ERG) was performed for elucidation of the time–course of enzymatic activities and retinal functional changes after injection. Scanning electron microscopy (SEM) was used for the observation of vitreoretinal surface. Results: Enzymatic activity of plasmin of 0.5 units, which is applied for clinical use, is equivalent to 13.8 fibrin degradation units (FU). Thus, in our experiments, 0.1 ml of enzymatic solution containing subtilisin NAT of 13.8 FU was injected into the vitreous in animal eyes. B–mode ultrasonography demonstrated the occurrence of PVD after intravitreal injection of subtilisin NAT. ERG analysis showed no significant changes in electrophysiological retinal function even after the use of subtilisin NAT. Also, SEM showed smooth retinal surfaces in the experimental eyes in a dose–dependent manner, indicating the occurrence of PVD, but not in control eyes. Conclusions: Our results suggested that subtilisin NAT is a useful enzyme for pharmacological vitrectomy because of its safe and effective PVD–inducing activity.

Keywords: enzymes/enzyme inhibitors • vitreoretinal surgery • pharmacology