Tags

Type your tag names separated by a space and hit enter

Immunocytochemical localization and biochemical characterization of large proteoglycans in developing rat bone.
J Oral Sci 1998; 40(2):77-87JO

Abstract

This study used biochemical and light and electron microscopic immunohistochemical methods to localize and characterize large hyaluronate-binding proteoglycans in the developing mandible of fetal rats at embryonic day 15 (Day 15) to Day 18 using a monoclonal antibody (MAb) 5D5. This antibody is derived from bovine sclera and specifically recognizes the core protein of large proteoglycan such as versican, neurocan and brevican, but not that of aggrecan. At the light microscopic level, MAb 5D5 moderately stained the extracellular matrices among osteoblasts at the centers of ossification in Day 15 mandible specimens. Weaker staining was observed in osteoblasts, whereas Meckel's cartilage lacked staining. Ultrastructural immunocytochemistry showed the presence of immunogold particles over unmineralized matrices among osteoblasts and their intracellular organelles. In Day 16 to 18 specimens, bone nodules were recognized in LR gold sections before immunostaining, but, after immunostaining, consistently appeared devoid of mineral crystals and were seen as a demineralized structure that had an electron dense periphery within which fine filamentous and granular material were present. The appearance of these structures was created by the demineralization of thin sections on grids during immunostaining. Specific immunogold staining was clearly seen over the demineralized structures corresponding to bone nodules. The majority of immunogold particles tended to localize inside of the structures. Bone proteins were extracted from fresh, Day 18 specimens with a three-step technique: 4 M guanidine HCl (GdnCl,-extract), aqueous EDTA without GdnCl (E-extract), followed by GdnCl. Western blot analysis of SDS-polyacrylamide gel electrophoresis after chondroitinase ABC digestion, showed that G1-extract gave a 5D5 reactive band greater than 400 kDa, whereas E-extract produced two major reactive populations of small molecular size with core proteins approximately 63 and 74 kDa. These results indicate that the large proteoglycan having smaller molecular weight is preferentially localized to bone nodules and may correlate with bone matrix mineralization.

Authors+Show Affiliations

Department of Anatomy, Nihon University School of Dentistry, Tokyo, Japan.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

9680765

Citation

Lee, I, et al. "Immunocytochemical Localization and Biochemical Characterization of Large Proteoglycans in Developing Rat Bone." Journal of Oral Science, vol. 40, no. 2, 1998, pp. 77-87.
Lee I, Ono Y, Lee A, et al. Immunocytochemical localization and biochemical characterization of large proteoglycans in developing rat bone. J Oral Sci. 1998;40(2):77-87.
Lee, I., Ono, Y., Lee, A., Omiya, K., Moriya, Y., & Takagi, M. (1998). Immunocytochemical localization and biochemical characterization of large proteoglycans in developing rat bone. Journal of Oral Science, 40(2), pp. 77-87.
Lee I, et al. Immunocytochemical Localization and Biochemical Characterization of Large Proteoglycans in Developing Rat Bone. J Oral Sci. 1998;40(2):77-87. PubMed PMID: 9680765.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Immunocytochemical localization and biochemical characterization of large proteoglycans in developing rat bone. AU - Lee,I, AU - Ono,Y, AU - Lee,A, AU - Omiya,K, AU - Moriya,Y, AU - Takagi,M, PY - 1998/7/29/pubmed PY - 1998/7/29/medline PY - 1998/7/29/entrez SP - 77 EP - 87 JF - Journal of oral science JO - J Oral Sci VL - 40 IS - 2 N2 - This study used biochemical and light and electron microscopic immunohistochemical methods to localize and characterize large hyaluronate-binding proteoglycans in the developing mandible of fetal rats at embryonic day 15 (Day 15) to Day 18 using a monoclonal antibody (MAb) 5D5. This antibody is derived from bovine sclera and specifically recognizes the core protein of large proteoglycan such as versican, neurocan and brevican, but not that of aggrecan. At the light microscopic level, MAb 5D5 moderately stained the extracellular matrices among osteoblasts at the centers of ossification in Day 15 mandible specimens. Weaker staining was observed in osteoblasts, whereas Meckel's cartilage lacked staining. Ultrastructural immunocytochemistry showed the presence of immunogold particles over unmineralized matrices among osteoblasts and their intracellular organelles. In Day 16 to 18 specimens, bone nodules were recognized in LR gold sections before immunostaining, but, after immunostaining, consistently appeared devoid of mineral crystals and were seen as a demineralized structure that had an electron dense periphery within which fine filamentous and granular material were present. The appearance of these structures was created by the demineralization of thin sections on grids during immunostaining. Specific immunogold staining was clearly seen over the demineralized structures corresponding to bone nodules. The majority of immunogold particles tended to localize inside of the structures. Bone proteins were extracted from fresh, Day 18 specimens with a three-step technique: 4 M guanidine HCl (GdnCl,-extract), aqueous EDTA without GdnCl (E-extract), followed by GdnCl. Western blot analysis of SDS-polyacrylamide gel electrophoresis after chondroitinase ABC digestion, showed that G1-extract gave a 5D5 reactive band greater than 400 kDa, whereas E-extract produced two major reactive populations of small molecular size with core proteins approximately 63 and 74 kDa. These results indicate that the large proteoglycan having smaller molecular weight is preferentially localized to bone nodules and may correlate with bone matrix mineralization. SN - 1343-4934 UR - https://www.unboundmedicine.com/medline/citation/9680765/Immunocytochemical_localization_and_biochemical_characterization_of_large_proteoglycans_in_developing_rat_bone_ L2 - https://www.lens.org/lens/search?q=citation_id:9680765 DB - PRIME DP - Unbound Medicine ER -