Novel use of a Dektak 150 surface profiler unmasks differences in resorption pit profiles between control and Charcot patient osteoclasts

TY - JOUR

T1 - Novel use of a Dektak 150 surface profiler unmasks differences in resorption pit profiles between control and Charcot patient osteoclasts

AU - Petrova, Nina L.

AU - Petrov, Peter K.

AU - Edmonds, Michael E.

AU - Shanahan, Catherine M.

PY - 2014/4

Y1 - 2014/4

N2 - 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 (μm2) 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 μm2 [1.6-3.9] vs. 8.3 μm 2 [5.6-10.6], 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.

AB - 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 (μm2) 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 μm2 [1.6-3.9] vs. 8.3 μm 2 [5.6-10.6], 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.

KW - Charcot osteoarthropathy

KW - Osteoclasts

KW - Resorption pits

KW - Surface profilometry

UR - http://www.scopus.com/inward/record.url?scp=84896491130&partnerID=8YFLogxK

U2 - 10.1007/s00223-013-9820-9

DO - 10.1007/s00223-013-9820-9

M3 - Article

C2 - 24322885

AN - SCOPUS:84896491130

SN - 0171-967X

VL - 94

SP - 403

EP - 411

JO - Calcified Tissue International

JF - Calcified Tissue International

IS - 4

ER -