Differential requirements for IKKα and IKKβ in the differentiation of primary human osteoarthritic chondrocytes

Eleonora Olivotto, Rosa Maria Borzi, Roberta Vitellozzi, Stefania Pagani, Annalisa Facchini, Michela Battistelli, Marianna Penzo, Xiang Li, Flavio Flamigni, Jun Li, Elisabetta Falcieri, Andrea Facchini, Kenneth B. Marcu

Research output: Contribution to journalArticlepeer-review

Abstract

Objective. Osteoarthritic (OA) chondrocytes behave in an intrinsically deregulated manner, characterized by chronic loss of healthy cartilage and inappropriate differentiation to a hypertrophic-like state. IKKα and IKKβ are essential kinases that activate NF-κB transcription factors, which in turn regulate cell differentiation and inflammation. This study was undertaken to investigate the differential roles of each IKK in chondrocyte differentiation and hypertrophy. Methods. Expression of IKKα or IKKβ was ablated in primary human chondrocytes by retrotransduction of specific short-hairpin RNAs. Micromass cultures designed to reproduce chondrogenesis with progression to the terminal hypertrophic stage were established, and anabolism and remodeling of the extracellular matrix (ECM) were investigated in the micromasses using biochemical, immunohistochemical, and ultrastructural techniques. Cellular parameters of hypertrophy (i.e., proliferation, viability, and size) were also analyzed. Results. The processes of ECM remodeling and mineralization, both characteristic of terminally differentiated hypertrophic cells, were defective following the loss of IKKα or IKKβ. Silencing of IKKβ markedly enhanced accumulation of glycosaminoglycan in conjunction with increased SOX9 expression. Ablation of IKKα dramatically enhanced type II collagen deposition independent of SOX9 protein levels but in association with suppressed levels of runt-related transcrition factor 2. Moreover, IKKα-deficient cells retained the phenotype of cells in a pre-hypertrophic-like state, as evidenced by the smaller size and faster proliferation of these cells prior to micromass seeding, along with the enhanced viability of their differentiated micromasses. Conclusion. IKKα and IKKβ exert differential roles in ECM remodeling and endochondral ossification, which are events characteristic of hypertrophic chondrocytes and also complicating factors often found in OA. Because the effects of IKKα were more profound and pleotrophic in nature, our observations suggest that exacerbated IKKα activity may be responsible, at least in part, for the characteristic abnormal phenotypes of OA chondrocytes.

Original languageEnglish
Pages (from-to)227-239
Number of pages13
JournalArthritis and Rheumatism
Volume58
Issue number1
DOIs
Publication statusPublished - Jan 2008

ASJC Scopus subject areas

  • Immunology
  • Rheumatology

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