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Reveromycin A‐Induced Apoptosis in Osteoclasts Is Not Accompanied by Necrosis
Author(s) -
Mead Brittany,
Morgan Heather,
MannKnowlton Alyssa,
Tedeschi Laura,
Sloan Chris,
Lang Spenser,
Hines Cory,
Gragg Megan,
Stofer Jonathan,
Riemann Kaitlin,
Derr Tyler,
Heller Emily,
Collins David,
Landis Paul,
Linthan,
Jones Daniel
Publication year - 2015
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.25125
Subject(s) - rankl , osteoclast , apoptosis , necrosis , microbiology and biotechnology , chemistry , acid phosphatase , lactate dehydrogenase , programmed cell death , activator (genetics) , receptor , medicine , biology , biochemistry , enzyme
Reveromycin A (RM‐A), a small natural product isolated from Streptomyces bacteria, is a potential osteoporosis therapeutic in that it specifically induces apoptosis in osteoclasts but not osteoblasts. The purpose of the study presented here was to further elucidate the intracellular mechanisms of RM‐A death effects in mature osteoclasts. A specific clone of RAW264.7 murine macrophages that was previously characterized for its ability to acquire an osteoclast nature on differentiation was differentiated in the presence of receptor activator of nuclear factor kappa B ligand (RANKL). Subsequent staining was performed for tartrate‐resistant acid phosphatase to confirm their osteoclast character. These osteoclasts were treated with ten micromolar RM‐A for 2, 4, 6, 24, and 48 h at a pH of 5.5. Peak apoptosis induction occurred at 4–6 h as measured by caspase 3 activity. Lactate dehydrogenase release assay revealed no significant RM‐A‐induced necrosis. Western blot analysis of cytoplasmic extracts demonstrated activation of caspase 9 (2.3‐fold at 2 h and 2.6‐fold at 4 h, each P < 0.05) and no significant changes in Bcl‐X L . In nuclear extracts, NFκB levels significantly increased on differentiation with RANKL but then remained constant through RM‐A treatment. Over the extended time course studied, RM‐A‐induced apoptosis in osteoclasts was not accompanied by necrosis, suggesting that RM‐A would likely have limited effects on immediate, neighboring bone cell types. This specific cell death profile is promising for potential clinical investigations of RM‐A as a bone antiresorptive. J. Cell. Biochem. 116: 1646–1657, 2015. © 2015 Wiley Periodicals, Inc.