Acanthamoeba binds to extracellular matrix proteins in vitro.
OBJECTIVE
To identify host-tissue amoeba interactions that may be important in the pathogenesis of Acanthamoeba keratitis, the ability of the opportunistic pathogen Acanthamoeba polyphaga to bind various components of the extracellular matrix (collagen type IV, laminin, or fibronectin) was examined in vitro.


METHODS
A polyphaga, isolated from a case of human amoebic keratitis, was used in the studies. In the experiments, 96-well plates were coated with 0-, 5-, 10-, 20-, or 50-micrograms/ml solutions of the basal lamina proteins laminin or collagen type IV, the extracellular matrix protein fibronectin, or casein (control). Amoeba were metabolically labeled with 35[S]-methionine, and 1x 10(4) labeled amoeba in phosphate buffered saline (PBS) were seeded per well and allowed to bind for 20 min. After washing with PBS, bound amoeba were solubilized with 1% sodium dodecyl sulphate (SDS) and scintillation counting was used to determine the number of bound amoeba.


RESULTS
Counts from casein and protein-free controls were not significantly different from each other (P > 0.05). There was a significant increase in the binding of 35[S]-labeled A. polyphaga to collagen IV, laminin, and fibronectin over controls (P < 0.0001) and the binding was concentration-dependent. The rank order of binding was collagen > or = laminin >> fibronectin. Alpha-methyl-mannopyranoside, but not fucose, inhibited binding of labeled A. polyphaga to collagen IV, laminin, and fibronectin in a concentration-dependent manner.


CONCLUSIONS
In summary, the binding assays show that Acanthamoeba bind preferentially to collagen, laminin, and fibronectin, in that order, and that the adherence process is inhibited by mannose.
Gordon
V R
VR
Department of Veternary Pathobiology, Purdue University, West Lafayette, Indiana.
Asem
E K
EK
Vodkin
M H
MH
McLaughlin
G L
GL
D000048:Acanthamoeba / Q000378:metabolism*; D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D002448:Cell Adhesion / Q000187:drug effects; D003094:Collagen / Q000378:metabolism*; D005109:Extracellular Matrix / Q000187:drug effects; D005353:Fibronectins / Q000378:metabolism*; D005643:Fucose / Q000494:pharmacology; D007797:Laminin / Q000378:metabolism*; D008766:Methylmannosides / Q000494:pharmacology
D005353:Fibronectins; D007797:Laminin; D008766:Methylmannosides; D005643:Fucose; D003094:Collagen