PCR detection of cytokines in normal human and pagetic osteoblast-like cells

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Abstract
We investigated the expression of cytokine transcripts in osteoblast-like cells derived from explants of pagetic and normal bone. A reverse transcription-linked PCR was used that allowed the simultaneous analysis of a range of cytokines. Normal osteoblast-like cells were found to contain the transcripts for IL-1β, IL-6, and TGF-β1. For the first time we detected in bone cells the two other mammalian isoforms of TGF-β, β2, and β3. Furthermore, we have also identified mRNA for IL-3 and the novel chemotactic factor, IL-8. Using this sensitive technique it was not possible to detect mRNA for IL-1α, IL-2, IL-4, IL-5, IL-7, TNF-α, or interferon-γ. The human osteosarcoma cell line Saos-2 also showed a similar pattern of expression of these cytokines to primary osteoblast-like cells, with the exception that TNF-α was also identified. Cells isolated from pagetic bone showed essentially the same profile of cytokine expression as normal bone except that TNF-α was also detected in two of four samples. The cytokine profile of successive populations of cells harvested from one explant culture at 9, 22, and 57 days showed a consistent pattern of cytokine expression, demonstrating the phenotypic stability of the osteoblast-like cells in long-term cultures.
Funding Information
  • Wellcome Trust
  • Anatomical Society of Great Britain
  • North West Cancer Research Fund

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