Ex vivo analysis of leukocyte hydrogen peroxide production using a bi-plate model in mice

Abstract

Implantation of artificial materials is followed by inflammation and wound healing, where phagocytic cells play an important role. The mechanisms whereby the implant surface may elicit and modulate leukocyte functions in vivo are not understood, partly due to the technical difficulties of examining the local inflammatory events in vicinity of the material-tissue interface with conventional biochemical and immunological techniques. In the present study a newly developed biplate implant was inserted subcutaneously in the mouse. Leukocytes from the local inflammatory exudate and leukocytes associated to the surface of the implants were retrieved after 1 and 6 days and separately assayed with respect to hydrogen peroxide (H2O2) production ex vivo. Implantation caused a local accumulation of predominantly mononuclear cells in the surrounding subcutaneous tissue. The H2O2 production was found to be low in both the subcutaneous exudate and the implant-associated leukocytes, irrespective of implant material and implantation times. However, ex vivo-stimulation with phorbol myristate acetate (PMA) caused an enhanced H2O2 production. These observations show that biplate implants do not maximally activate the oxidative metabolism of the recruited leukocytes. The exudate leukocytes were more responsive to PMA stimulation in comparison with implant-associated leukocytes, indicating that properties of the implant surface and possibly surface-associated proteins could modify the responsiveness of the phagocytic cells at the implant site. Our results suggest that the present biplate model may be suitable for further studies on local production of oxygen metabolites and function of leukocytes at implanted biomaterials.