Dose–Dependent in vitro Inhibition of Corneal Matrix Metalloproteinase by Pothomorphe umbellata Ethanolic Roots Extract After Alkali Injury

Abstract

Purpose: : To evaluate the dose–dependent ability of Pothomorphe umbellata ethanolic crude extract to inhibit matrix metalloproteinases (MMP) in normal cornea and in cornea after alkali injury in rabbits.

Material and Methods: : Corneas of albino rabbits were injured with 1M NaOH for 20 seconds. After 48 hours of injury corneas (6 eyes) were excised, homogenized and analyzed for matrix metalloproteinase MMP–9 (92kDa), pro–MMP–2 (72kDa) and MMP–2 (67kDa) activity by gelatinase zymography. The activity was also measured in normal corneas. After eletrophoresis separation (20 µg/mL protein) the gels were incubated with 50, 100 or 250 µg/mL of lyophilized hydro alcoholic (1:1) root crude extract of P.umbellata containing 7% de 4– nerolidylcathecol. The activity of the enzymes was compared with that of untreated gel.

Results: : After 48 hours of injury the activity of all MMPs were increased when compared with control eyes. When the gels were incubated with P.umbellata extract the activity of all MMPs decreased in a dose–dependent manner. MMP–9 activity decreased by approximately 39% after incubation with 50 µg/mL and was completely abolished with 100 and 250 µg/mL of the extract. The activity of pro–MMP–2 decreased by 56% and MMP–2 decreased by 38% after incubation with 50 µg/mL. The activity of pro–MMP–2 was almost completely abolished after incubation with 250 µg/mL of the extract. For MMP–2 the incubation with 100 or 250 µg/mL of ethanolic roots crude extract of P.umbellata promoted a 10 fold decrease in the activity.

Conclusions: : The results suggest that Pothomorphe umbellata root crude extract can be useful as an alternative therapy to control MMPs activity after corneal injury.

Keywords: cornea: basic science • neovascularization • extracellular matrix