betadex and Corneal-Ulcer

alpha-Toxin mediates extreme corneal damage during Staphylococcus aureus keratitis. Chemical inhibition of this toxin was sought to provide relief from toxin-mediated pathology.. Inhibition of alpha-toxin by phosphate-buffered saline (PBS), 0.1% methyl-beta-cyclodextrin (CD), or CD plus cholesterol (0.1%, CD-cholesterol) was assayed by hemolysis of rabbit erythrocytes. Pathologic changes in rabbit corneas injected with 12 hemolytic units of alpha-toxin suspended in PBS, 1% CD, or 1% CD-cholesterol were compared over time. Rabbit corneas injected with 10(2) colony forming units (CFU) of S. aureus were treated from 7 to 13 hours postinfection (PI) with a total of 15 drops of CD-cholesterol, CD, or PBS. Slit lamp examination (SLE) and measurement of erosions were performed at 13 hours PI and bacteria were quantified at 14 hours PI.. Toxin-mediated lysis of erythrocytes was inhibited up to 16,000-fold in the presence of CD-cholesterol compared with CD or PBS. Eyes injected with alpha-toxin mixed with CD-cholesterol had, at 7 hours postinjection, significantly smaller erosions than eyes injected with alpha-toxin in PBS or alpha-toxin mixed with CD (P = 0.0090 and P = 0.0035, respectively). Eyes infected with S. aureus and treated with CD-cholesterol had significantly lower SLE scores than eyes treated with CD or PBS (P or= 0.0648).. CD-cholesterol is a potent inhibitor of alpha-toxin activity in vitro and an effective means to arrest corneal damage during S. aureus keratitis.

Topics: Animals; Bacterial Toxins; beta-Cyclodextrins; Cholesterol; Cornea; Corneal Ulcer; Disease Models, Animal; Drug Therapy, Combination; Erythrocytes; Exotoxins; Eye Infections, Bacterial; Hemolysin Proteins; Hemolysis; Rabbits; Sodium Chloride; Staphylococcal Infections; Staphylococcal Toxoid; Staphylococcus aureus; Virulence; Virulence Factors

Pseudomonas aeruginosa enters corneal epithelial cells in vitro via membrane microdomains or lipid rafts. Bacterial entry, mediated by the cystic fibrosis transmembrane conductance regulator (CFTR), promotes infection and disease. This study was conducted to determine whether P. aeruginosa and CFTR are colocalized to rafts in isogenic corneal cells expressing wild-type (WT) or mutant DeltaF508-CFTR and whether disruption of the rafts both in vitro and in vivo affects the bacterial levels and the course of the disease.. Transformed human corneal epithelial cells from a patient homozygous for DeltaF508-CFTR, and the same cells corrected with WT-CFTR, were exposed to six isolates of P. aeruginosa-three invasive and three cytotoxic strains-in the presence of beta-cyclodextrin (CD), which disrupts rafts. Association and cellular uptake of the invasive strains were measured, as was lactate dehydrogenase release induced by the cytotoxic strains. Scratch-injured mouse eyes were infected with the six P. aeruginosa strains, and the effect of prophylactic or therapeutic administration of CD on bacterial levels and disease was evaluated.. P. aeruginosa and CFTR were colocalized with lipid rafts in cells with WT-CFTR, and CD treatment of these cells disrupted bacterial association, internalization, and cytotoxic effects. Cells expressing DeltaF508-CFTR were marginally affected by CD. Prophylactic and therapeutic topical application of CD ameliorated corneal disease and reduced the bacterial count in the eye.. P. aeruginosa enters human corneal epithelial cells via lipid rafts containing CFTR, and disruption of raft-mediated uptake of this organism by CD protects against disease and reduces bacterial levels in the mouse model of keratitis.

Topics: Animals; Bacterial Adhesion; beta-Cyclodextrins; Blotting, Western; Cell Line, Transformed; Colony Count, Microbial; Corneal Ulcer; Cystic Fibrosis Transmembrane Conductance Regulator; Disease Models, Animal; Epithelium, Corneal; Eye Infections, Bacterial; Fluorescent Antibody Technique; Humans; Membrane Microdomains; Mice; Microscopy, Confocal; Pseudomonas aeruginosa; Pseudomonas Infections; RNA, Small Interfering