Z. Lin, M. B. , N. J. Pavlos, B. Sc. (Hons. ), Ph. D. , M. A. Cake, B

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Presentation transcript:

Evidence that human cartilage and chondrocytes do not express calcitonin receptor Z. Lin, M.B., N.J. Pavlos, B.Sc. (Hons.), Ph.D., M.A. Cake, B.VMS. (Hons.), Ph.D., D.J. Wood, B.Sc., M.B.B.S., M.S., F.R.C.S., F.R.A.C.S., J. Xu, M.D., Ph.D., M.H. Zheng, Ph.D., D.M., A.R.C.P.A., F.R.C.Path Osteoarthritis and Cartilage Volume 16, Issue 4, Pages 450-457 (April 2008) DOI: 10.1016/j.joca.2007.08.003 Copyright © 2007 Osteoarthritis Research Society International Terms and Conditions

Fig. 1 The mRNA expression of CTR in human cartilage and chondrocytes. (A) Primary human chondrocytes (passage 0) cultured on monolayer with spherical phenotype. (B) The gene expression of CTR in five individual human cartilage tissue samples and cultured chondrocytes. Articular cartilage samples were taken from OA patients undergoing total joint replacement. Chondrocytes were isolated from human cartilage and cultured in vitro. RNA extracted from human GCT served as positive control. CTR (200bp) is expressed in human GCT, but not in any of the human cartilage tissue or isolated chondrocytes. Housekeeping gene 18S served as internal control. Osteoarthritis and Cartilage 2008 16, 450-457DOI: (10.1016/j.joca.2007.08.003) Copyright © 2007 Osteoarthritis Research Society International Terms and Conditions

Fig. 2 Western blot analysis of CTR in human giant cell tumor and primary chondrocytes. A band representing CTR appears at ∼56.9kDa only in human GCT tissue. Thirty micrograms of GCT and chondrocyte samples were resolved by SDS-PAGE and transferred overnight onto nitrocellulose before being probed for CTR with specific antibodies. Alpha-tubulin served as an internal loading control. Osteoarthritis and Cartilage 2008 16, 450-457DOI: (10.1016/j.joca.2007.08.003) Copyright © 2007 Osteoarthritis Research Society International Terms and Conditions

Fig. 3 Immunodetection of CTR in human GCT and articular cartilage. (A) Immunostained osteoclast in human GCT imprint, hematoxylin counterstain, immunostaining is predominantly on the osteoclast plasma membrane (original magnification, 400×); (B) immuno- and TRAP-staining of osteoclasts in human GCT imprint (original magnification, 400×); (C) negative control without primary antibodies in a human GCT imprint, hematoxylin counterstain (original magnification, 400×); (D) full thickness of human articular cartilage section with subchondral bone, immunostained with anti-CTR antibody and counterstained with hematoxylin (original magnification, 40×); (E) chondrocytes in the same section show no specific staining, hematoxylin counterstain (original magnification, 400×); (F) osteoclasts (as indicated by arrow) in the subchondral bone region stained positive for CTR (original magnification, 400×). Osteoarthritis and Cartilage 2008 16, 450-457DOI: (10.1016/j.joca.2007.08.003) Copyright © 2007 Osteoarthritis Research Society International Terms and Conditions

Fig. 4 The mRNA expression of CRLR in human GCT, human cartilage and chondrocytes. CRLR is abundantly expressed in human GCT and cultured chondrocytes (passage 1), but absent in human cartilage tissue. Osteoarthritis and Cartilage 2008 16, 450-457DOI: (10.1016/j.joca.2007.08.003) Copyright © 2007 Osteoarthritis Research Society International Terms and Conditions

Fig. 5 cAMP response of chondrocytes to sCT. Human chondrocytes (passage 1) isolated from OA cartilage were cultured on monolayer and stimulated with sCT (10−7M, 10−8M, and 10−9M) or forskolin (100μM) for 30min. The cAMP content from culture supernatant and cell lysis were both measured. sCT does not change the cAMP content in chondrocyte compared to vehicle treatment (IBMX/100μM), however, forskolin significantly elevates the cAMP level (P<0.001). Osteoarthritis and Cartilage 2008 16, 450-457DOI: (10.1016/j.joca.2007.08.003) Copyright © 2007 Osteoarthritis Research Society International Terms and Conditions

Fig. 6 The expression profiles of chondrocytic genes (aggrecan, Type II collagen, MMP-1, MMP-3, MMP-13) in sCT treated human chondrocytes (passage 1) by Real-time PCR. Mean Ct value of target genes was normalized to it housekeeping gene 18S. Statistic significances were calculated by Student's t test. Results are shown as mean±sem. P<0.05 is considered statistically significant. Osteoarthritis and Cartilage 2008 16, 450-457DOI: (10.1016/j.joca.2007.08.003) Copyright © 2007 Osteoarthritis Research Society International Terms and Conditions