Phagocytosis and biodegradation of short-chain poly[(R)-3-hydroxybutyric acid] particles in macrophage cell line

The biocompatibility a nd the degradation mechanisms of block co-polyurethanes containing crystallizable telechelic poly [(R)-3-hydroxybutyric acid] (PHB) segments have found recently growing interest for the possible biomedical applications of these new materials. The random hydrolytic cleavage of the amorphous part of these polymers might result, in vivo, in the production of small crystalline particles of low molecular weight PHB that could then undergo phagocytosis and biodegradation inside phagosomes. To test this possibility, a fluorescent-labelled PHB segment was synthesized, precipitated in the form of crystalline particles, and used for an in vitro investigation of its interaction with macrophage cell line. Light and fluorescence microscopy performed in the present study clearly show that the fluorescent particles are well internalized in phagosomes already after 1 h of incubation. The number of phagocytized particles decreases notably after 8 days of incubation. A quantitative determination of the time dependence of the phagocytosis was obtained through laser cytometry and fluorescence-activated cell sorting (FAGS). Fluorescence spectroscopy and high-performance liquid chromatography (HPLC) analysis of extracts of cell supernatants revealed the presence of supposed degradation products of PHB after 8 days of incubation, suggesting that macrophages could degrade low molecular weight PHB.