Activin Regulation of Osteoclast Formation, Motility, Function, and Survival

The process of bone resorption is regulated at multiple levels. Current bone-active therapies target bone resorption, whereas targets such as the formation of osteoclasts (OC) and their motility, fusion and lifespan have been largely unexplored. We and others have shown that RANKL and Activin A (ActA) are potent stimulators of murine bone marrow osteoclastogenesis. However, ActA treatment does not induce osteoclastogenesis in purified populations of murine bone marrow macrophages (BMM), but inhibits RANKLNinduced OC development. We hypothesized that these factors differentially regulate the migration of both OC precursors and mature OC. We determined the effects of ActA and RANKL on BMM motility and bone resorption using time-lapse video microscopy. BMMs were plated onto culture dishes with 25 ng/ml mCSF alone for 24hr, prior to the addition of 100 ng/mL RANKL or 50 ng/mL ActA, alone or together, for up to 4d. Other BMMs were cultured on bone slices for 5d in the presence of mCSF+RANKL to stimulate OC differentiation prior to treatment with RANKL or ActA, alone or together for an additional 1N2d. The 4hr movement of ten OC precursors were tracked and analyzed with NIH ImageJ. Parameters calculated for each cell included cumulative track length, migration rate, maximum instantaneous velocity, range of velocity, and percent total displacement. BMMs migrated at a constant velocity and distance from d1N4. RANKL significantly increased parameters on d2 and d3, an effect that was lost by d4. In contrast, ActA treatment decreased all parameters on d1N4. Surprisingly, when added with RANKL, ActA treatment completely blocked all RANKL stimulated OC precursor motility on plastic, as well as mature RANKLNstimulated OC motility on bone slices. ActA treatment of mature OC (d5) also suppressed RANKLN stimulated bone resorption, measured by pit formation. The suppression was associated with a decrease in OC lifespan, since ActA increased activated Caspase 3, and caused a decrease in actin ring formation in mature OC on bone slices. Collectively these data indicate a dual modulatory function of ActA on OCs. ActA indirectly stimulates murine whole bone marrow osteoclastogenesis, but it also directly inhibits RANKLNinduced murine OC differentiation, motility, and resorption and enhances OC apoptosis.

Year	Entry
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Year

Entry

1983

Jee WS, Wronski TJ, Morey ER, Kimmel DB. Effects of spaceflight on trabecular bone in rats. Am J Physiol Regul Integr Comp Physiol. 1983;244(3):R310-R314.

2005

Bellido T, Ali AA, Gubrij I, Plotkin LI, Fu Q, O’Brien CA, Manolagas SC, Jilka RL. Chronic elevation of parathyroid hormone in mice reduces expression of sclerostin by osteocytes: a novel mechanism for hormonal control of osteoblastogenesis. Endocrinology. November 2005;146(11):4577-4583.

2004

van Bezooijen RL, Roelen BAJ, Visser A, van der Wee-Pals L, de Wilt E, Karperien M, Hamersma H, Papapoulos SE, ten Dijke P, Löwik CWGM. Sclerostin is an osteocyte-expressed negative regulator of bone formation, but not a classical BMP antagonist. J Exp Med. March 15, 2004;199(6):805-814.

1998

Lacey DL, Timms E, Tan H-L, Kelley MJ, Dunstan CR, Burgess T, Elliott R, Colombero A, Elliott G, Scully S, Hsu H, Sullivan J, Hawkins N, Davy E, Capparelli C, Eli A, Qian Y-X, Kaufman S, Sarosi I, Shalhoub V, Senaldi G, Guo J, Delaney J, Boyle WJ. Osteoprotegerin ligand is a cytokine that regulates osteoclast differentiation and activation. Cell. April 17, 1998;93(2):165-176.

2001

Toyosawa S, Shintani S, Fujiwara T, Ooshima T, Sato A, Ijuhin N, Komori T. Dentin matrix protein 1 is predominantly expressed in chicken and rat osteocytes but not in osteoblasts. J Bone Miner Res. November 2001;16(11):2017-2026.