Nature and distribution of chondroitin sulphate and dermatan sulphate proteoglycans in rabbit alveolar bone |
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Authors: | Minoru Takagi Masao Maeno Toshihiro Yamada Kunihiko Miyashita and Kichibee Otsuka |
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Institution: | (1) Department of Anatomy, Nihon University School of Dentistry, 1-8-13, Kanda-Surugadai, Chiyoda-ku, 101 Tokyo, Japan;(2) Department of Biochemistry, Nihon University School of Dentistry, 1-8-13, Kanda-Surugadai, Chiyoda-ku, 101 Tokyo, Japan |
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Abstract: | Summary The type and distribution of mineral binding and collagenous matrix-associated chondroitin sulphate and dermatan sulphate
proteoglycans in rabbit alveolar bone were studied biochemically and immunocytochemically, using three monoclonal antibodies
(mAb 2B6, 3B3, and 1B5). The antibodies specifically recognize oligosaccharide stubs that remain attached to the core protein
after enzymatic digestion of proteoglycans and identify epitopes in chondroitin 4-sulphate and dermatan sulphate; chondroitin
6-sulphate and unsulphated chondroitin; and unsulphated chondroitin, respectively. In addition, mAb 2B6 detects chondroitin
4-sulphate with chondroitinase ACII pre-treatment, and dermatan sulphate with chondroitinase B pre-treatment. Bone proteins
were extracted from fresh specimens with a three-step extraction procedure: 4m guanidine HCl (G-1 extract), 0.4m EDTA (E-extract), followed by guanidine HCl (G-2 extract), to characterize mineral binding and collagenous matrix associated
proteoglycans in E- and G2-extracts, respectively. Biochemical results using Western blot analysis of SDS-polyacrylamide gel
electrophoresis of E- and G2-extracts demonstrated that mineral binding proteoglycans contain chondroitin 4-sulphate, chondroitin
6-sulphate, and dermatan sulphate, whereas collagenous matrix associated proteoglycans showed a predominance of dermatan sulphate
with a trace of chondroitin 4-sulphate and no detectable chondroitin 6-sulphate or unsulphated chondroitin. 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 chondroitin 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 distributed over the remainder of the bone matrix. |
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