Hyaluronic acid of high molecular weight inhibits proliferation and induces cell death in U937 macrophage cells.

Hyaluronic acid (HA), a major glycosaminoglycan component of the extracellular matrix, has regulatory influences on cells and cellular activities. To explore the effects of a high concentration (1 mg/mL) of high molecular weight HA (500-730 kD) on U937 macrophage growth dynamics, three factors that influence overall cellular growth, namely proliferation, apoptosis, and cell death, were examined. Cells were cultured with HA and were analyzed by flow cytometry every 24 hours during a 168-hour period for proliferation and the presence of apoptotic and dead cells. These analyses demonstrated that HA inhibits U937 macrophage proliferation in a time-dependent manner. Through the first 72 hours, cells exhibited slowed proliferation. However, no evidence of cell division arrest or reduced cell viability was observed. Thereafter, HA continued to diminish proliferation, but induced apoptosis. This data is consistent with regulatory influences secondary to HA binding to CD44 and/or RHAMM cell surface receptors, both of which were shown to be expressed on U937 macrophages. This study demonstrates that a high concentration of high molecular weight HA greatly inhibits macrophage population growth by the dual actions of impeding cell proliferation and inducing apoptosis.

Macrophage exposure to particulate titanium induces phosphorylation of the protein tyrosine kinase lyn and the phospholipases Cgamma-1 and Cgamma-2.

A frequent long-term complication of total joint arthroplasty is aseptic loosening, the end result of wear debris production, synovial macrophage activation, inflammatory mediator release, and osteolysis about the implant-bone or cement-bone interface. To elucidate the mechanisms of particle-induced macrophage activation and mediator production, we studied early signal transduction events using J774A.1 macrophages and 3 microm titanium particles. Treating macrophages with herbimycin A or genistein, two inhibitors of protein tyrosine kinases (PTKs), inhibited titanium phagocytosis as well as secretion of tumor necrosis factor-alpha (TNF-alpha) and prostaglandin-E2 (PGE2) in a dose-dependent manner. Both processes therefore depend on a PTK signaling cascade. Specifically, macrophage exposure to titanium-induced phosphorylation of multiple proteins including the Src kinase Lyn and phospholipase Cgamma-1 and Cgamma-2. Phosphorylation peaked within 2 min and returned to baseline within 45 min. Similar but not identical phosphorylation patterns were obtained when cells were stimulated with titanium preincubated with serum or albumin, suggesting distinct signal transduction pathways dependent on particle coating.