title: Novel use of a Dektak 150 surface profiler unmasks differences in resorption pit profiles between control and Charcot patient osteoclasts.
abstract: We hypothesized that newly formed osteoclasts from patients with acute Charcot osteoarthropathy can resorb surfaces of bone more extensively compared with controls. Peripheral blood monocytes, isolated from eight Charcot patients and nine controls, were cultured in vitro on 24-well plates and bovine bone discs in duplicate with macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor κβ ligand (RANKL). Osteoclast formation was assessed by tartrate-resistant acid phosphatase staining (TRAcP) at day 17. Resorption was measured at day 21 after toluidine blue staining by two methods: (1) area of resorption at the surface by image analysis (%) and (2) area of resorption under the surface (μm(2)) measured by a Dektak 150 Surface Profiler. Ten 1,000 μm-long scans were performed per disc. Pits were classified as unidented, bidented, and multidented according to their shape. Although the number of newly formed TRAcP positive multinucleated cells (>3 nuclei) was similar in M-CSF + RANKL-treated cultures between controls and Charcot patients, the latter exhibited increased resorbing activity. The area of resorption on the surface by image analysis was significantly greater in Charcot patients compared with controls (21.1 % [14.5-26.2] vs. 40.8 % [35.4-46.0], median [25-75th percentile], p < 0.01), as was the area of resorption under the surface (2.7 x 10(3) μm(2) [1.6 x 10(3)- 3.9 x 10(3)] vs. 8.3 x 10(3) μm (2) [5.6 x 10(3)- 10.6 x 10(3), [corrected] p < 0.01) after profilometry. In Charcot patients pits were deeper and wider and more frequently presented as multidented pits. This application of the Dektak 150 Surface Profiler revealed novel differences in resorption pit profile from osteoclasts derived from Charcot patients compared with controls. Resorption in Charcot patients was mediated by highly aggressive newly formed osteoclasts from monocytes eroding large and deep areas of bone.
authors: Diabetic Foot Clinic, King's College Hospital, Denmark Hill, London, SE5 9RS, UK, petrovanl@yahoo.com.
authors: Nina L
authors: NL
authors: Petrova
authors: Peter K
authors: PK
authors: Petrov
authors: Michael E
authors: ME
authors: Edmonds
authors: Catherine M
authors: CM
authors: Shanahan
mesh_terms: D000328:Adult; D000690:Amyotrophic Lateral Sclerosis / Q000473:pathology*; D000818:Animals; D001862:Bone Resorption; D001842:Bone and Bones; D016022:Case-Control Studies; D002417:Cattle; D003470:Culture Media; D005260:Female; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000166:cytology; D016173:Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D008297:Male; D008875:Middle Aged; D009000:Monocytes / Q000166:cytology; D010010:Osteoclasts / Q000166:cytology*; D053245:RANK Ligand / Q000494:pharmacology; D013499:Surface Properties
chemical_list: D003470:Culture Media; D053245:RANK Ligand; C505688:TNFSF11 protein, human; D016173:Macrophage Colony-Stimulating Factor