title: Time-dependent cellular and transcriptional changes in the osteoblast lineage associated with sclerostin antibody treatment in ovariectomized rats.
abstract: Inhibition of sclerostin with sclerostin antibody (Scl-Ab) has been shown to stimulate bone formation, decrease bone resorption, and increase bone mass in both animals and humans. To obtain insight into the temporal cellular and transcriptional changes in the osteoblast (OB) lineage associated with long-term Scl-Ab treatment, stereological and transcriptional analyses of the OB lineage were performed on lumbar vertebrae from aged ovariectomized rats. Animals were administered Scl-Ab 3 or 50mg/kg/wk or vehicle (VEH) for up to 26weeks (d183), followed by a treatment-free period (TFP). At 50mg/kg/wk, bone volume (BV/total volume [TV]) increased through d183 and declined during the TFP. Bone formation rate (BFR/bone surface [BS]) and total OB number increased through d29, then progressively declined, coincident with a decrease in total osteoprogenitor (OP) numbers from d29 through d183. Analysis of differentially expressed genes (DEGs) from microarray analysis of mRNA isolated from laser capture microdissection samples enriched for OB, lining cells, and osteocytes (OCy) revealed modules of genes that correlated with BFR/BS, BV/TV, and osteoblastic surface (Ob.S)/BS. Expression change of canonical Wnt target genes was similar in all three cell types at d8, including upregulation of Twist1 and Wisp1. At d29, the pattern of Wnt target gene expression changed in the OCy, with Twist1 returning to VEH level, sustained upregulation of Wisp1, and upregulation of several other Wnt targets that continued into the TFP. Predicted activation of pathways recognized to integrate with and regulate canonical Wnt signaling were also activated at d29 in the OCy. The most significantly affected pathways represented transcription factor signaling known to inhibit cell cycle progression (notably p53) and mitogenesis (notably c-Myc). These changes occurred at the time of peak BFR/BS and continued as BFR/BS declined during treatment, then trended toward VEH level in the TFP. Concurrent with this transcriptional switch was a reduction in OP numbers, an effect that would ultimately limit bone formation. This study confirms that the initial transcriptional response in response to Scl-Ab is activation of canonical Wnt signaling and the data demonstrate that there is induction of additional regulatory pathways in OCy with long-term treatment. The interactions between Wnt and p53/c-Myc signaling may be key in limiting OP populations, thus contributing to self-regulation of bone formation with continued Scl-Ab administration.
authors.affiliation: Department of Comparative Biology and Safety Sciences, Amgen Inc., Thousand Oaks, CA, USA.
authors.forename: Scott
authors.initials: S
authors.lastname: Taylor
authors.affiliation: Department of CardioMetabolic Disorders, Amgen Inc., Thousand Oaks, CA, USA.
authors.forename: Michael S
authors.initials: MS
authors.lastname: Ominsky
authors.affiliation: Department of Comparative Biology and Safety Sciences, Amgen Inc., Thousand Oaks, CA, USA.
authors.forename: Rong
authors.initials: R
authors.lastname: Hu
authors.affiliation: Department of Comparative Biology and Safety Sciences, Amgen Inc., Thousand Oaks, CA, USA.
authors.forename: Efrain
authors.initials: E
authors.lastname: Pacheco
authors.affiliation: Department of Comparative Biology and Safety Sciences, Amgen Inc., Thousand Oaks, CA, USA.
authors.forename: Yudong D
authors.initials: YD
authors.lastname: He
authors.affiliation: WIL Research Laboratories, Hillsborough, NC, USA.
authors.forename: Danielle L
authors.initials: DL
authors.lastname: Brown
authors.affiliation: Department of Physiological Sciences, University of Florida, Gainesville, FL, USA.
authors.forename: J Ignacio
authors.initials: JI
authors.lastname: Aguirre
authors.affiliation: Department of Physiological Sciences, University of Florida, Gainesville, FL, USA.
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Wronski
authors.affiliation: Department of Comparative Biology and Safety Sciences, Amgen Inc., Thousand Oaks, CA, USA.
authors.forename: Sabina
authors.initials: S
authors.lastname: Buntich
authors.affiliation: Department of Comparative Biology and Safety Sciences, Amgen Inc., Thousand Oaks, CA, USA.
authors.forename: Cynthia A
authors.initials: CA
authors.lastname: Afshari
authors.affiliation: Department of Comparative Biology and Safety Sciences, Amgen Inc., Thousand Oaks, CA, USA.
authors.forename: Ian
authors.initials: I
authors.lastname: Pyrah
authors.affiliation: Department of Comparative Biology and Safety Sciences, Amgen Inc., Thousand Oaks, CA, USA.
authors.forename: Paul
authors.initials: P
authors.lastname: Nioi
authors.affiliation: Department of Comparative Biology and Safety Sciences, Amgen Inc., Thousand Oaks, CA, USA. Electronic address: rboyce@amgen.com.
authors.forename: Rogely Waite
authors.initials: RW
authors.lastname: Boyce
mesh_terms: D000818:Animals; D000906:Antibodies / Q000494:pharmacology*; D019485:Bone Morphogenetic Proteins / Q000276:immunology*; D002452:Cell Count; D019070:Cell Lineage / Q000187:drug effects*; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D005819:Genetic Markers / Q000276:immunology*; D008954:Models, Biological; D009929:Organ Size / Q000187:drug effects; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D010011:Osteocytes / Q000187:drug effects / Q000378:metabolism; D010012:Osteogenesis / Q000187:drug effects; D010052:Ovariectomy; D010641:Phenotype; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D013234:Stem Cells / Q000187:drug effects / Q000378:metabolism; D013997:Time Factors; D014158:Transcription, Genetic / Q000187:drug effects*
chemical_list: D000906:Antibodies; D019485:Bone Morphogenetic Proteins; D005819:Genetic Markers; C486031:Sost protein, rat
keywords: Anabolics; Bone; Cell signaling; Osteoporosis; Therapeutics; Transcription factor; Wnt signaling