Modulation of tumor necrosis factor related apoptosis-inducing ligand (trail) receptors in a human osteoclast model in vitro
We have previously shown that osteoclasts (OCLs) from multiple myeloma (MM) specimens vary from healthy OCLs in their expression of the TRAIL receptors. TRAIL (TNF-Related Apoptosis-Inducing Ligand), a member of the TNF superfamily, has been shown to induce apoptosis in cells by binding receptors DR4 and DR5, but not DcR1 and DcR2, its decoy receptors, which lack the necessary internal death domain. The observed modulation of these receptors may confer a resistance to apoptosis in the MM environment, and could be related to the cytokine pattern that primarily involves the resorption promoting Receptor Activator of NF-[kappa]B Ligand (RANKL) and Macrophage Inflammatory Protein 1 (MIP-1[alpha]). The aim of our study was to determine which cytokines present in the disease might be responsible for this modulation. In long term cultures of OCL precursors from cord blood in the presence of M-CSF and RANKL, multinucleated cells (MNCs) that express OCL markers form, and can resorb bone. Through immunocytochemistry we showed that these MNCs can express all four TRAIL receptors. By stimulating with various cytokines (RANKL, MIP-1[alpha], Transforming Factor [bêta] (TGF[bêta]), osteoprotegerin (OPG), TRAIL), and parathyroid hormone (PTH) in OCL cultures, we were able to observe receptor modulation at the mRNA level using real time PCR, the protein level using Western blot analysis, and cell surface expression via immunocytochemistry. To determine if these changes translated to a difference in resistance to apoptosis, cells treated with [with] apoptosis-inducing levels of TRAIL after 5 days of stimulation with the selected cytokines were evaluated via TUNEL to quantify apoptosis. While no correlation has yet been established between the observed receptor modification and apoptosis induction, sample size is a factor, and further tests will be performed. Our results suggest the possibility that TRAIL receptor modification is induced by multiple cytokines present in bone diseases, capable of altering both the susceptibility and resistance pathways in osteoclasts. By potentially prolonging the lifespan of the OCL, these regulatory influences may ultimately be contributory factors to the augmentation of resorption in the micro-environment of bone resorptive diseases like multiple myeloma, Paget's disease of bone, or osteoporosis.