Degradation of glycosaminoglycans by reactive oxygen species derived from stimulated polymorphonuclear leukocytes

The effect of reactive oxygen species (ROS), generated by in vitro stimulation of isolated PMN upon the main GAG components of mineralised and non-mineralised connective tissues was investigated. PMN were isolated from whole blood and the production of the ROS superoxide (O2.-) and hydroxyl radicals (.OH) was stimulated by the addition of phorbol myristyl acetate (PMA) and PMA/FeCl3-EDTA chelate respectively and their production assessed over a 24 h period. The glycosaminoglycans (GAG), hyaluronan, chondroitin 4-sulphate and dermatan sulphate, were exposed to the ROS fluxes, incubated at 37 degrees C for 1 h and 24 h. GAG fragmentation was examined by gel exclusion chromatography and modification to hexuronic acid and hexosamine residues determined. Stimulation of PMN with PMA resulted in a burst of O2.- production for 1 h, which was sustained at a reduced level for 24 h. Fragmentation of GAG was observed for all GAG examined. Modification to the GAG was evident, with hyaluronan being more susceptible to loss of GAG residues than sulphated GAG. Modification of sugar residues increased with the incubation time and loss of the hexuronic acid residues was greater than loss of hexosamine residues. Addition of FeCl3-EDTA chelate, which led to the generation of .OH and was sustained over the 24 h period, demonstrated similar trends of GAG modification although increased degradation and loss of hexosamine and hexuronic acid were observed. GAG chains are constituents of PGs and their modification is likely to affect the function of these macromolecules and be of importance in considering the pathogenesis of inflammatory diseases, including periodontal diseases.