Purchase this article with an account.
H. Alizadeh, M. Hurt, S. Neelam, J.Y. Niederkorn; Characterization of the Mannose-Stimulated Acanthamoeba Castellanii Cytolytic Protein . Invest. Ophthalmol. Vis. Sci. 2003;44(13):713.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Purpose: Acanthamoeba trophozoites express a mannose binding receptor, which facilitates adhesion of the trophozoites to mannosylated proteins on corneal epithelial cells. Upon binding, Acanthamoeba castellanii trophozoites secrete a ~133 kDa protein that is cytolytic to corneal epithelium. We examined whether the protein was cytolytic to a range of ocular and non-ocular cells, how it killed corneal epithelium, and whether it could play a role in degrading types I and IV collagen. Methods: A. castellanii trophozoites were grown in PYG or PYG containing 100 mM methyl α-D-mannopyranoside. The cytolytic protein (133-kDa) was purified by FPLC size exclusion and ionic exchange and tested for cytopathic effects (CPE) and collagenolytic activity in vitro. Collagenolytic activity was measured by ELISA. A chicken antiserum against the 133-kDa protein was used to neutralize CPE and collagenolytic activity in vitro. Results: The 133-kDa protein was highly cytolytic to the corneal epithelial and stromal cells. Non-ocular cells such as human smooth muscle, liver, and small intestinal epithelium were resistant. Annexin V and caspase-3 analysis revealed that corneal cell death was mediated by apoptosis (P<0.001). The 133-kDa protein was successful at degrading both types I and IV collagen in vitro (P<0.001). The anti-133-kDa antiserum specifically prevented collagenolytic activity and neutralized CPE of the corneal epithelial cells in vitro. Conclusion: The results suggest that mannose induces A. castellanii to secrete a protein that may aid in the desquamation of the corneal epithelium, the penetration of Bowman’s membrane, and the dissolution of the stromal matrix.
This PDF is available to Subscribers Only