title: High density micromass cultures of a human chondrocyte cell line: a reliable assay system to reveal the modulatory functions of pharmacological agents.
abstract: Osteoarthritis is a highly prevalent and disabling disease for which we do not have a cure. The identification of suitable molecular targets is hindered by the lack of standardized, reproducible and convenient screening assays. Following extensive comparisons of a number of chondrocytic cell lines, culture conditions, and readouts, we have optimized an assay utilizing C-28/I2, a chondrocytic cell line cultured in high-density micromasses. Utilizing molecules with known effects on cartilage (e.g. IL-1β, TGFβ1, BMP-2), we have exploited this improved protocol to (i) evoke responses characteristic of primary chondrocytes; (ii) assess the pharmacodynamics of gene over-expression using non-viral expression vectors; (iii) establish the response profiles of known pharmacological treatments; and (iv) investigate their mechanisms of action. These data indicate that we have established a medium-throughput methodology for studying chondrocyte-specific cellular and molecular responses (from gene expression to rapid quantitative measurement of sulfated glycosaminoglycans by Alcian blue staining) that may enable the discovery of novel therapeutics for pharmacological modulation of chondrocyte activation in osteoarthritis.
authors.affiliation: William Harvey Research Institute, Barts and The London School of Medicine, Queen Mary University of London, London, United Kingdom. k.v.greco@qmul.ac.uk
authors.forename: K V
authors.initials: KV
authors.lastname: Greco
authors.forename: A J
authors.initials: AJ
authors.lastname: Iqbal
authors.forename: L
authors.initials: L
authors.lastname: Rattazzi
authors.forename: G
authors.initials: G
authors.lastname: Nalesso
authors.forename: N
authors.initials: N
authors.lastname: Moradi-Bidhendi
authors.forename: A R
authors.initials: AR
authors.lastname: Moore
authors.forename: M B
authors.initials: MB
authors.lastname: Goldring
authors.forename: F
authors.initials: F
authors.lastname: Dell'Accio
authors.forename: M
authors.initials: M
authors.lastname: Perretti
mesh_terms: D000423:Alcian Blue; D000893:Anti-Inflammatory Agents / Q000494:pharmacology*; D002358:Cartilage, Articular / Q000166:cytology; D002460:Cell Line; D049109:Cell Proliferation; D019902:Chondrocytes / Q000166:cytology* / Q000187:drug effects* / Q000502:physiology; D005786:Gene Expression Regulation / Q000502:physiology; D006801:Humans; D053583:Interleukin-1beta / Q000494:pharmacology; D009288:Naproxen / Q000494:pharmacology*; D011239:Prednisolone / Q000494:pharmacology*
chemical_list: D000893:Anti-Inflammatory Agents; D053583:Interleukin-1beta; D009288:Naproxen; D011239:Prednisolone; D000423:Alcian Blue