Study on the Anti-Angiogenic Property of Euphorbia hirta Linn. (Tawa-Tawa)and Psidium guajava Linn. (Guava) Crude Leaf Extracts Using Duck (Anaa domesticus) Chorioallantoic Membrane (Cam) Assay

Abstract

INTRODUCTION

Cancer is one of the leading causes of death worldwide and in the Philippines as well. One of the critical events behind cancer metastasis is angiogenesis, which is the formation of new blood vessels. Angiogenesis is the biological process of the creation of new blood vessels from pre-existing capillaries and post-capillary vessels. Prevention of the supply of oxygen and nutrients to tumor cells would be necessary for posing treatment to the disease. Anti-angiogenic chemicals help decrease the proliferation of cancer cells. Thus, the study aimed to determine the difference between the mean percentage inhibition of different treatments and active constituents present in the plants being studied which will pave the way to the discovery of new drugs against cancer.

METHODS

The study made use of Duck (Anaa domesticus) Chorioallantoic Membrane (CAM) Assay in vivo as a tool in the determination of the anti-angiogenic property of Euphorbia hirta Linn. (Tawa-tawa), and Psidium guajava Linn. (Guava), which purportedly exhibit several pharmacological effects. Five treatments replicated five times were prepared: T1: 100% tawa-tawa crude extract, T2: 100% guava crude extract, T3 50% tawa-tawa and 50% guava crude extract, T4: Celecoxib, and T5: distilled water. The CAM at the site of application for angiogenesis was examined. In response to proangiogenic stimuli, the newly formed blood vessels appeared converging toward the disc in a wheel-spoke pattern. Through the help of a qualified scientist, the blood vessel branch point at each area of the quadrant was counted manually. One-way Analysis of Variance (ANOVA) was used to determine if significant differences exist between the variables.

RESULTS

Comparative analysis of the mean percentage inhibition of the different treatments in the CAM treated duck eggs showed that T1, 100% tawa-tawa leaf and stem extract gained the highest percentage inhibition than the other treatments; and T2, 100% Psidium guajava Linn. crude leaf extract gained the lowest percentage of CAM inhibition in the blood vessels formed. Therefore, the individual Euphorbia hirta Linn. crude leaf extract exhibits the highest anti-angiogenic effect when compared to the combination of both Euphorbia hirta Linn. and Psidium guajava Linn. crude leaf extracts, and Celecoxib.

DISCUSSIONS

The inhibition of angiogenesis is done by suppressing the action of the vascular endothelial growth factor of VEGF since it is considered as one of the most important proangiogenic factors. The presence of flavonoids in tawa-tawa leaf extracts contribute to its suppressing property towards angiogenesis (Villagonzalo, 2013). Moreover, flavonoids decrease the thickness of blood vessels and vascular endothelial growth factor (VEGF) levels (Shao et. al, 1998). In conclusion, tawa-tawa crude leaf extract can be an antiangiogenic agent due to the presence of flavonoids, a secondary metabolite.