BM osteoclast progenitors are increased in SCD and produce increased amounts of active catK. (A) Flow cytometry identification of OPCs in the BM by gating for cells, live cells, CD11b^–, CD45R/B220^–, CD3^–, and finally CD117⁺/CD115⁺ cells. (B) Quantification of OPC percentage of live cells in the BM of AA, AS, and SS mice (n = 5-8 mice per group). Data are expressed as mean ± standard deviation. Statistical significance, *P < .05, determined by one-way analysis of variance with Tukey’s post hoc test. (C) Representative images of BM cells from AA and SS mice treated with either 10 ng/mL M-CSF or 30 ng/mL M-CSF + 100 ng/mL RANKL for 21 days. Osteoclasts were determined as tartrate-resistant acid phosphatase–positive (pink) with at least 3 nuclei (blue). (D) Quantified number of osteoclasts per well (n = 4 mice and 8 wells per group). (E) Representative cathepsin zymograms of AA and SS osteoclasts. Active protein appears as white bands. Densitometry quantification of 200 kDa, 110 kDa, 60 kDa, and 50 kDa bands of active cathepsins in AA and SS osteoclasts (n = 4 mice per group). (F) Representative western blots of catK protein in AA and SS osteoclasts. Recombinant catK was used as a positive control. Densitometry quantification of 60 kDa, 55 kDa, 37 kDa, and 25 kDa bands of catK protein in AA and SS osteoclasts (n = 4-5 mice per group). AA = black, SS = red. Data are expressed as mean ± standard deviation. Statistical significance, *P < .05, determined by Welch’s t test. AU, arbitrary unit.