M. Takagi et al., NATURE AND DISTRIBUTION OF CHONDROITIN SULFATE AND DERMATAN SULFATE PROTEOGLYCANS IN RABBIT ALVEOLAR BONE, Histochemical Journal, 28(5), 1996, pp. 341-351
The type and distribution of mineral binding and collagenous matrix-as
sociated chondroitin sulphate and dermatan sulphate proteoglycans in r
abbit alveolar bone were studied biochemically and immunocytochemicall
y, using three monoclonal antibodies (mAb 2B6, 3B3, and 1B5). The anti
bodies specifically recognize oligosaccharide stubs that remain attach
ed to the core protein after enzymatic digestion of proteoglycans and
identify epitopes in chondroitin 4-sulphate and dermatan sulphate; cho
ndroitin 6-sulphate and unsulphated chondroitin; and unsulphated chond
roitin, respectively. In addition, mAb 2B6 detects chondroitin 4-sulph
ate with chondroitinase ACII pre-treatment, and dermatan sulphate with
chondroitinase B pre-treatment. Bone proteins were extracted from fre
sh specimens with a three-step extraction procedure: 4 M guanidine HCl
(G-1 extract), 0.4 M EDTA (E-extract), followed by guanidine HCl (G-2
extract), to characterize mineral binding and collagenous matrix asso
ciated proteoglycans in E- and G2-extracts, respectively Biochemical r
esults using Western blot analysis of SDS-polyacrylamide gel electroph
oresis of E- and G2-extracts demonstrated that mineral binding proteog
lycans contain chondroitin 4-sulphate, chondroitin 6-sulphate, and der
matan sulphate, whereas collagenous matrix associated proteoglycans sh
owed a predominance of dermatan sulphate with a trace of chondroitin 4
-sulphate and no detectable chondroitin 6-sulphate or unsulphated chon
droitin. Immunocytochemistry showed that staining associated with the
mineral phase was limited to the walls of osteocytic lacunae and bone
canaliculi, whereas staining associated with the matrix phase was seen
on and between collagen fibrils in the remainder of the bone matrix.
These results indicate that mineral binding proteoglycans having chond
roitin 4-sulphate, dermatan sulphate, and chondroitin 6-sulphate were
localized preferentially in the walls of the lacunocanalicular system,
whereas collagenous associated dermatan sulphate proteoglycans were d
istributed over the remainder of the bone matrix.