Effect of aqueous humor on apoptosis of inflammatory cell types

PURPOSE. TO determine whether aqueous humor promotes cell death in cells in volved in inflammatory responses. METHODS. Multiple immune cell types, most characteristically involved in in flammatory responses, were incubated for 24, 48, and 72 hours in the presen ce or absence of 50% aqueous humor. Promotion of cell death was assayed by staining for an early indicator of apoptosis. The percentage of cells under going apoptosis was measured by flow cytometry. To identify partially the a poptosis-inducing factor, aqueous humor was pretreated with proteinase K to degrade protein. In other experiments, aqueous humor was fractionated by c entrifugation on filters capable of separating molecules above and below 10 kDa or 30 kDa kilodaltons in size. RESULTS. Rabbit aqueous humor promoted apoptosis in a wide variety of immun e cells, including lymphokine-activated natural killer cells, resting T cel ls, an activated T-cell line, RAW 264.7 and J774A0.1 monocyte-macrophage ce ll Lines, and neutrophils. As previously shown, aqueous humor did not promo te apoptosis of murine corneal endothelial cells. Apoptosis was also not in duced in human corneal endothelium, mouse corneal epithelium, or iris/cilia ry body cell lines, instead, aqueous humor partially protected these ocular tissues from starvation-induced cell death. Pretreatment with proteinase K inhibited the apoptosis-inducing activity. Moreover, the apoptosis-inducin g activity segregated with the aqueous humor fraction containing molecules less than than 10 kDa in size. CONCLUSIONS. These data show that aqueous humor contains a factor or factor s that promote death of cells that participate in inflammatory processes. B y contrast, ocular tissues, such as the corneal endothelium and iris/ciliar y body, are impervious to aqueous humor-induced cell death. The aqueous hum or-borne factor(s) may contribute to the immune privilege of the anterior c hamber by purging potential inflammatory cells.