G1-G2 aggrecan product that can be generated by M-calpain on truncation at Ala709-Ala710 is present abundantly in human articular cartilage.
To elucidate the specific function of m-calpain in the metabolism of aggrecan in human articular cartilage, the prevalence and localization of a large glycosaminoglycan-bearing aggrecan product generated by m-calpain in human osteoarthritis (OA) cartilage were investigated. Extracts of human OA articular cartilage were analysed by immunostaining using new polyclonal anti-VPGVA antiserum that detects the COOH terminal neoepitope IVTQVVPGVA(709) generated by m-calpain-related cleavage within the keratan sulphate rich region of human aggrecan. Immunoblotting analyses of aggrecan populations in guanidine hydrochloride-extracts showed that OA cartilages contained anti-VPGVA positive aggrecan products with the COOH terminal neoepitope ... VPGVA(709), resulting from truncation between the Ala(709)-Ala(710) m-calpain-related cleavage site. This aggrecan product consisted of two NH(2) terminal globular domain (G1 and G2) and KS side chains. Immunohistochemical staining showed that anti-VPGVA positive staining was localized within chondrocytes and spread to the surrounding interterritorial matrix. Confocal microscopic analysis showed subcellular colocalization of anti-VPGVA and anti m-calpain. These results indicate that the aggrecan product with the COOH terminal neoepitope VPGVA(709) is synthesized regularly by intracellular processing in chondrocytes, and is present abundantly as a limited form of aggrecan. M-calpain is the major candidate of the proteinase to generate this aggrecan product during the intracellular aggrecan processing.
Maehara
Hideaki
H
Department of Orthopaedic Surgery, Gifu University Graduate School of Medicine, 1-1 Yanagido, Gifu, Japan.
Suzuki
Kiichi
K
Sasaki
Tomohiro
T
Oshita
Hidefumi
H
Wada
Eriko
E
Inoue
Toshiyuki
T
Shimizu
Katsuji
K
D053674:Aggrecans / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D002154:Calpain / Q000378:metabolism*; D002358:Cartilage, Articular / Q000378:metabolism*; D019902:Chondrocytes / Q000378:metabolism; D000939:Epitopes; D006801:Humans; D007632:Keratan Sulfate / Q000378:metabolism; D010003:Osteoarthritis / Q000378:metabolism; D013552:Swine
D053674:Aggrecans; D000939:Epitopes; D007632:Keratan Sulfate; D002154:Calpain; C449000:m-calpain