Figure 5.

Mistargeting of lysosomal hydrolases does not result in increased resorptive capacity of MLII osteoclasts.

1. Relative intracellular enzyme activity of lysosomal hydrolases β-hexosaminidase (β-hex) and β-galactosidase (β-gal) in primary cultured osteoclasts from MLII mice compared to WT. Data are shown as mean and SD (n = 3 primary cultures per group). *p (β-hex) = 0.0001, *p (β-gal) = 0.0001 for WT versus MLII by independent Student's t-test.

2. Primary cultures of osteoclasts from MLII and WT mice were incubated with [³⁵S]-methionine for 1 h and either harvested (0 h) or chased for 5 and 20 h, followed by immunoprecipitation of lysosomal proteases CtsZ and CtsK from cell extracts and media (p, precursor posttranslational modified by high-mannose type N-glycans; p′, precursor modified with N-glycans processed to the complex type form, m, mature form).

3. Primary cultured osteoclasts from WT and MLII mice stained for TRAP activity.

4. Number of osteoclasts per visual field (OcN/VF) formed after 6 days of differentiation with 1,25(OH)₂-vitamin D3 alone (−Rankl) (n = 6 primary cultures per group) or for the last 3 days in the additional presence of M-CSF and Rankl (+Rankl) (n = 3 primary cultures per group). Data are shown as mean and SD.

5. Quantification of area covered by resorption pits after 12-days-cultivation of osteoclasts on dentine chips. Scale bars = 50 µm. Data are shown as mean and SD (n = 3 primary cultures per group).

6. Topographic maps of resorption pits visualized by three dimensional stack imaging. Scale bars: 50 µm. Quantification of pit depth (WT, n = 275 pits; MLII, n = 285 pits). All values are mean and SD.