Polymorphonuclear cell apoptosis in exudates generated by polymers.

Abstract

Flow cytometry was used to quantify apoptotic and necrotic polymorphonuclear (PMN) cells in an exudate generated by biomaterials, and the results were compared with determinations of spontaneous apoptosis and necrosis in PMN cells from the bloodstream. The exudate formed inside cylindrical tubes subcutaneously implanted in the dorsal region of rats was collected over a 1-week period. A rapid and simple staining procedure based on the spectral properties of the bisbenzemide Hoechst 33342 was used to identify apoptotic PMN cells. Quantification of permeabilized PMN cells stained by propidium iodide was possible in the same unfixed specimens. The percentages of apoptotic and permeabilized PMN cells in peripheral rat blood were low (1.8 +/-0 0.5% and 1.7 +/- 0.7%, respectively), similar to results found in humans. In exudates generated by polyvinyl chloride (PVC), the percentages of apoptotic and permeabilized PMN cells were higher than in the blood. The percentage of PMN cells undergoing apoptosis progressively increased with time and reached a maximum at day 2 (27% +/- 6%). The percentage of permeabilized cells progressively increased with time and was much higher than the percentage of apoptotic cells on days 4 and 8. Apoptosis and necrosis of PMN cells at day 2 were inhibited when tubes were filled with 10% serum. Selective inhibition of apoptosis with a caspase inhibitor in vivo indicated that apoptosis and necrosis are two separate pathways leading to the death of PMN cells in the exudate. At day 2, polyurethane (PU) was associated with a lower rate of apoptosis than PVC or a random copolymer of trimethylene carbonate (TMC) and epsiloncaprolactone (ECL). Apoptosis was interpreted as an organized cell removal process that limits inflammation. Apoptosis was the natural route of PMN cell death at the early stage of inflammation.