Vitamin K1 prevents diabetic cataract by inhibiting lens aldose reductase 2 (ALR2) activity

R Thiagarajan,K Saraboji,R Ravichandran,M K N Sai Varsha,V Srinivasan

doi:10.1038/s41598-019-51059-2

Abstract

This study investigated the potential of vitamin K1 as a novel lens aldose reductase inhibitor in a streptozotocin-induced diabetic cataract model. A single, intraperitoneal injection of streptozotocin (STZ) (35 mg/kg) resulted in hyperglycemia, activation of lens aldose reductase 2 (ALR2) and accumulation of sorbitol in eye lens which could have contributed to diabetic cataract formation. However, when diabetic rats were treated with vitamin K1 (5 mg/kg, sc, twice a week) it resulted in lowering of blood glucose and inhibition of lens aldose reductase activity because of which there was a corresponding decrease in lens sorbitol accumulation. These results suggest that vitamin K1 is a potent inhibitor of lens aldose reductase enzyme and we made an attempt to understand the nature of this inhibition using crude lens homogenate as well as recombinant human aldose reductase enzyme. Our results from protein docking and spectrofluorimetric analyses clearly show that vitamin K1 is a potent inhibitor of ALR2 and this inhibition is primarily mediated by the blockage of DL-glyceraldehyde binding to ALR2. At the same time docking also suggests that vitamin K1 overlaps at the NADPH binding site of ALR2, which probably shows that vitamin K1 could possibly bind both these sites in the enzyme. Another deduction that we can derive from the experiments performed with pure protein is that ALR2 has three levels of affinity, first for NADPH, second for vitamin K1 and third for the substrate DL-glyceraldehyde. This was evident based on the dose-dependency experiments performed with both NADPH and DL-glyceraldehyde. Overall, our study shows the potential of vitamin K1 as an ALR2 inhibitor which primarily blocks enzyme activity by inhibiting substrate interaction of the enzyme. Further structural studies are needed to fully comprehend the exact nature of binding and inhibition of ALR2 by vitamin K1 that could open up possibilities of its therapeutic application.

Highlights

Aldose reductase inhibition is one of the best strategies to inhibit polyol pathway and its associated complications in diabetes
We would like to emphasise here that in this study animals with blood glucose more than 220 mg/dL were considered to be diabetic/hyperglycemic and this falls into that category, notwithstanding the lower levels when compared to STZ alone rats. These results suggest that vitamin K1 had more beneficial effect in terms of decrease in blood glucose level in diabetic rats, when compared to zopolrestat and is very effective in inhibiting hyperglycemia, which could contribute to its anti-diabetic function
Previous studies have shown that the inhbition of aldose reductase activity could limit the complications arising from diabetes[2,71,72]

Summary

Materials and Methods

Streptozotocin, β-NADH, thiobarbituric acid (TBA), sorbitol dehydrogenase and DL- glyceraldehyde were procured from Sigma Aldrich. Effect of vitamin K1 pre- and co-incubation on control lens ALR2 activity. For co-incubation studies, 100 μl of the control eye lens homogenate was incubated for 5 min with 1.5 mg of vitamin K1 along with reagents for ALR2 measurement as mentioned above. Effect of time of pre-incubation of vitamin K1 on control lens ALR2 activity. 100 μl of control eye lens homogenate was pre-incubated with different concentrations of vitamin K1 (0.0664 fM, 33.20 pM, 0.74 nM, 3.32 mM, and 166 M) and incubated for 5 min. Effect of NADPH concentration on recombinant human aldose reductase activity pre-incubated with vitamin K1. Spectrofluorimetric analysis was performed to analyze the change in the secondary structure of the recombinant human ALR2 due to binding of vitamin K1, DL-glyceraldehyde, NADPH or zopolrestat. One-way analysis of variance (p < 0.05) was followed by Tukey’s post hoc test to evaluate the significant difference between STZ + vit K1, STZ and between STZ + zopolrestat, STZ

Result

C2 C10

Discussion