Comparison in localization between cystatin C and cathepsin K in osteoclasts and other cells in mouse tibia epiphysis by immunolight and immunoelectron microscopy

We compared the distribution of a cysteine proteinase inhibitor, cystatin C , with that of cathepsin K in osteoclasts of the mouse tibia by immunolight and immunoelectron microscopy. Light microscopically, strong immunoreactiv ity for cystatin C was found extracellularly along the resorption lacuna an d intracellularly in the organelles of osteoclasts. In serial sections, var ious patterns of cystatin C and cathepsin K localization were seen, specifi cally: (1) some resorption lacuna were positive for both cystatin C and cat hepsin K; (2) others were positive for either cystatin C or cathepsin K, bu t not both; and (3) some lacuna were negative for both. In osteoclasts, the localization of cystatin C was similar to that of cathepsin K. Furthermore , cystatin C immunoreactivity was detected in preosteoclasts and osteoblast s, whereas cathepsin K was seen only in preosteoclasts. Electron microscopi cally, cystatin C immunoreactive products were found in the rough endoplasm ic reticulum (ER), Golgi apparatus, vesicles, granules, and vacuoles of ost eoclasts. These cystatin C-positive vesicles had fused or were in the proce ss of fusion with the ampullar vacuoles (extracellular spaces) containing c ystatin C-positive, fragmented, fibril-like structures. The extracellular c ystatin C was deposited on and between the cytoplasmic processes of ruffled borders, and on and between type I collagen fibrils. In the basolateral re gion of osteoclasts, cystatin C-positive vesicles and granules also fused w ith vacuoles that contained cystatin C-positive or negative fibril-like str uctures. These results indicate that osteoclasts not only synthesize and se crete cathepsin K from the ruffled border into the bone resorption lacunae, but also a cysteine proteinase inhibitor, cystatin C. Therefore, it is sug gested that cystatin C regulates the degradation of bone matrix by cathepsi n K, both extracellularly and intracellularly. (C) 2001 by Elsevier Science Inc. All rights reserved.