Abstract: Regeneration of lost human tissue is well known, the research on regenerative medicine has gained scientific advancements. The growing understanding of biological concepts in the regeneration of ocular tissues as well as experiments on stem cells result in shift in the treatment of corneal diseases. Stem cells have been successfully isolated from variety of human tissues including mesenchymal tissues. Initial evidence from several studies has documented the likely breakthrough that stem cells offer for various life-threatening diseases that have so, far defeated modern medical care. So, this research aimed to study the role of stem cells in treatment of Acanthamoebic corneal ulcer. Test experiment included Acanthamoeba-infected female albino rats started local treatment by chlorhexidine 0.02% after 2 weeks post infection and Mesenchymal Stem Cells (MSCs) injected. Female rats were subdivided into 4 groups. Group (Gp) 1:15 rat infected and injected with MSCs intravenously at 2 weeks post-infection. Gp 2:15 rat infected and intravenously injected with MSCs at 3 weeks post-infection. Gp 3:15 rat infected and injected locally subconjunctival at 2 weeks post-infection. Gp 4:15 rat infected and locally subconjunctival injected at 3 weeks post-infection with MSCs. Control groups were composed of age and weight-matched control female rats, subdivided into: Gp 5-8. Gp 5 was composed of 10 Acanthamoeba-infected and non-injected (non-treated negative control) rats. Each 10 of them was sacrificed on dates corresponding to sacrifice dates of each test groups (at one week post treatment). Gp 6 is composed of 10 rats infected and intravenously injected by PBS (vehicle). Each were sacrificed on dates corresponding to sacrifice dates of Gp 1 and 2 (at one week post treatment). Gp 7:10 rats infected and locally subconjunctival injected by PBS (vehicle). Each 10 was sacrificed on dates corresponding to sacrifice dates of Gp 3 and 4 (at one week post treatment). Gp 8:10 Acanthamoeba-infected and locally treated by chlorhexidine 0.02% positive control rats which are infected and not injected with MSCs. Each 10 of them was sacrificed on dates similar to sacrifice dates of each test group (at one week post treatment). Rates were sacrificed on the planned dates then corneal specimens and blood samples were collected. There were significant decrease in the level of Vascular Endothelial Growth Factor (VEGF) in group 1, 3 and highly significant decrease in the level of Nitric Oxide (NO) in group 1-3 in comparison to positive control group 8. There were 20% engraftments with MSC in group 1. Treatment of Acanthamoeba corneal ulcer with chlorhexidine 0.02% and MSCs accelerated regeneration of the corneal epithelium and restored the antioxidant protective mechanism.