Polarization and secretion of cathepsin K precede tartrate-resistant acid phosphatase secretion to the ruffled border area during the activation of matrix-resorbing clasts

Abstract

The activation sequence of clasts (the designation clast was used because ultrastructurally in this tissue, it is not always possible to differentiate between chondroclasts sitting on cartilage and osteoclasts sitting on bone matrix) was studied in vivo using the healing of low-phosphate, vitamin D-deficiency rickets as a model system. Thus, the bones of 7-week-old rachitic animals were analyzed with a combination of morphological, biochemical, and molecular biological methods at 48 and 72 h, respectively, after change to normal food. A quantitative ultrastructural analysis showed that the number of clast profiles exhibiting the characteristic polarized features of actively resorbing cells, i.e., ruffled borders and clear zones, had reached normal levels after 48 h. By combining the data with quantitative analyses by the immunogold technique, we demonstrated that cathepsin K secretion was coupled to ruffled border formation in clasts irrespective of whether the number of polarized clasts was low (in rickets) or high (in healing). In contrast, the levels of tartrate-resistant acid phosphatase (TRAP) both between ruffles and in the outside matrix adjoining the ruffled border were low in polarized clasts both in rickets and at the early (48 h) healing time-point, but were increased at the latest (72 h) healing time-point. Interestingly, expression of TRAP and the cathepsin K at the mRNA level, as well as protein expression and the activity of TRAP, were not different during the healing sequence. Although the two enzymes are confined to the same clast populations, their secretion during the resorption process is apparently differentially regulated: cathepsin K secretion is coupled to ruffled border formation in clasts, whereas TRAP is secreted at a later stage during the resorption sequence, suggesting a role for secreted TRAP as a modulator of resorptive activity.

Glucocorticoids maintain human osteoclasts in the active mode of their resorption cycle

“…Since the common belief is that TRACP is a marker of OC activity, it may be argued that a prolongation of the resorption phase should lead to an increased instead of the decreased release of TRACP shown in this study. However, there are data indicating that TRACP is rather involved at the end of the resorption phase of the resorption cycle, (26,27) and in this case it makes sense that a prolongation of the resorption phase results in the observations made in this study, that is, a decreased TRACP release, which goes along with the increased levels of the resorption product CTX. Noteworthy, the generation of a high proportion of trenches in the presence of GCs is also supported by pictures from another study performed with human OCs generated from peripheral blood mononucleated cells, (13) but this report does not comment on the morphology of the excavations and mentions only the absence of effect of GCs on total resorption area.…”

Increased Tartrate-Resistant Acid Phosphatase Expression in Osteoblasts and Osteocytes in Experimental Osteoporosis in Rats

“…Moreover, Ovx-D rats presented a tendency toward increased TRAP mRNA expression in osteocytes, questioning the hypothesis of endocytosis being the mechanism enhancing TRAP protein expression and enzyme activity in osteoblasts and osteocytes in these rats. To further address this question, rats healing from nutritionally induced low-phosphate and vitamin D-deficiency rickets (experimental rickets) were analyzed as a model of increased osteoclast activity [23]. However, such rats did not show any differences in the level of TRAP protein expression or enzyme activity either in osteoblasts or in osteocytes, making it less likely that osteoblasts and osteocytes endocytose osteoclastic TRAP.…”

Osteoclast Responses to Lipopolysaccharide, Parathyroid Hormone and Bisphosphonates in Neonatal Murine Calvaria Analyzed by Laser Scanning Confocal Microscopy

“…The part of the plasma membrane enclosed by the actin enlarges into the highly convoluted ruffled border through which protons and liposome proteases such as cathepsin K, a cysteine protease highly active in digesting native collagen fibers, are transported (Vaananen et al 2000). Formation of the ruffled border and the intravesicular transport of degradation products are modulated by TRACP (Hollberg et al 2002), whereas the attachment of osteoclasts to the bone surface involves ligation of osteoclast integrins with extracellular matrix proteins within the bone matrix.…”