Dose-dependent in vitro inhibition of rabbit corneal matrix metalloproteinases by an extract of Pothomorphe umbellata after alkali injury

Abstract

The in vitro ability of Pothomorphe umbellata ethanolic crude extract to inhibit matrix metalloproteinase (MMP) in normal cornea and in cornea after alkali injury was demonstrated. Corneas of albino rabbits were injured with 1 N NaOH for 20 s. After 48 h the corneas were excised, homogenized and analyzed for MMP-9 (92 kDa), pro-MMP-2 (72 kDa) and MMP-2 (67 kDa) activity by gelatin zymography. The activity was also measured in untreated corneas. After electrophoresis of 20 µg protein, gels were incubated with 50, 100, or 250 µg/mL lyophilized hydroethanolic (1:1) root crude extract of P. umbellata standardized for 4-nerolidylcatechol (7.09%). The activity of the enzymes was compared with that of untreated gel. At 48 h after injury, the activity of all MMPs was increased compared with untreated eyes. When the gels were incubated with P. umbellata extract the activity of MMP-2, pro-MMP-2 and MMP-9 decreased in a dose-dependent manner. MMP-9 activity decreased by approximately 50% after incubation with 50 µg/mL and was completely abolished at 100 and 250 µg/mL of the extract. After incubation with 50 µg/mL the activity of pro-MMP-2 and MMP-2 also decreased by 50%. 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 the extract of P. umbellata promoted a 10-fold decrease in activity. In conclusion, P. umbellata root crude extract can be useful as an alternative therapy to control MMP activity after corneal injury.

Activity of matrix metalloproteinases 2 and 9 in cultured rabbit corneal epithelium cells stimulated by tumor necrosis factor alpha

“…MMP-2, together with MMP-9, degrades basement membrane components such as collagen IV, and is now extensively studied in the eye tissue (Sivak and Fini, 2002). Through numerous studies of MMP-2 and MMP-9, many investigators have shown that in corneal lesions such as keratitis (Yang et al, 2003;Xue et al, 2003), corneal alkali burn (Zhang et al, 2002;Barros et al, 2007), and corneal transplant rejection (Stuart et al, 2004), the expression levels and activities of MMP-2 and MMP-9 are increased, and they are considered the main factors contributing to these pathologies.…”

Piper umbellatum L.: A comparative cross-cultural analysis of its medicinal uses and an ethnopharmacological evaluation

“…In vitro inhibition of chloroquine-sensitive F32 strain of P. falciparum (70% inhibition at 40 g/ml) Bidla et al (2004) Ethanolic extract of the root In vitro dose-related reduction of MMP-2, pro-MMP-2 and MMP-9 in the cornea of the rabbit (dose 50, 100, 250 g/ml) (matrix metalloproteinases, MMP, are related to the failure of the cornea to re-epithelialize after injury) Barros et al (2007) Ethanolic extract of the root in gel containing 0.1% 4-nerolidylcatechol…”

Aqueous extract of Centella asiatica promotes corneal epithelium wound healing in vitro

“…The important steps of corneal epithelial wound healing are cell proliferation from the healthy area and cell migration to cover the damaged area, and form epithelial intercellular junctions to restore corneal epithelial integrity (Imanishi et al, 2000;Zelenka and Arpitha, 2008). Herbal components from Pothomorphe umbellata ethanolic crude extract and Chinese herbal medicine component, emodin have been reported to possess therapeutic value for corneal wound healing (Barros et al, 2007;Kitano et al, 2007). In addition, exogenous application of natural anti-oxidant such as ascorbic acid and vitamin E analog (Trolox) shown to improve the corneal wound healing activity in animal model by re-epithelialization in the wounded area (Hallberg et al, 1996).…”