It is well established that osteoblasts, the key cells involved in bone formation during development and in adult life, secrete a number of glycoproteins harboring autocrine and paracrine functions. Thus, investigating the osteoblastic secretome could yield important information for the pathophysiology of bone. In the present study, we characterized for the first time the secretome of human Hobit osteoblastic cells. We discovered that the secretome comprised 89 protein species including the powerful growth factor progranulin. Recombinant human progranulin (6nM) induced phosphorylation of mitogen-activated protein kinase in both Hobit and osteocytic cells and induced cell proliferation and survival. Notably, risedronate, a nitrogen-containing bisphosphonate widely used in the treatment of osteoporosis, induced the expression and secretion of progranulin in the Hobit secretome. In addition, our proteomic study of the Hobit secretome revealed that risedronate induced the expression of ERp57, HSP60 and HSC70, three proteins already shown to be associated with the prevention of bone loss in osteoporosis. Collectively, our findings unveil novel targets of risedronate-evoked biological effects on osteoblast-like cells and further our understanding of the mechanisms of action of this currently used compound.
Keywords: 2-DE; 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; Bisphosphonates; Differential proteomics; ECL; FBS; FPPS; IPP; LC; MALDI-ToF; MLO-Y4; MS/MS; MTT; Osteoblasts; Osteocytes; PAGE; PMSF; PVDF; Progranulin; Risedronate; SDS; TFA; enhanced chemiluminescence; farnesyl pyrophosphate synthase; fetal bovine serum; isopentyl diphosphate; liquid chromatography; matrix-assisted laser desorption/ionization-time of flight; murine long bone osteocyte-Y4; phenylmethylsulfonyl fluoride; polyacrylamide gel electrophoresis; polyvinylidene fluoride; sodium dodecyl sulfate; tandem mass spectrometry; trifluoroacetic acid; two-dimensional gel electrophoresis.
© 2013.