Modulation of equine articular chondrocyte messenger RNA levels following brief exposures to recombinant equine interleukin-1beta.
The effect of recombinant equine IL-1beta (EqIL-1beta) on steady-state mRNA levels of equine articular chondrocytes in high-density monolayer culture was investigated using a customized cDNA array analysis. Total RNA samples isolated from chondrocytes cultured in media alone or with the addition of 1 ng/ml EqIL-1beta for 1-, 3-, and 6-h durations of exposure were reverse transcribed, radiolabeled, and hybridized to a customized 380-target cDNA array. Means of duplicate log base 2 transformed hybridization signals were normalized to equine glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mean signal intensities. Differentially expressed transcripts were identified using a two-stage mixed linear analysis of variance model (Statistical Analysis Software, Cary, NC). A time-dependent pattern was observed in the number of transcripts increased > or =two-fold in response to EqIL-1beta after 1, 3 and 6h (1, 2 and 109 transcripts, respectively). At 6 h of EqIL-1beta stimulation, signal intensities for 88 cDNA targets with purported function in processes related to cell cycle, intracellular signaling, transcription, translation, extracellular matrix turnover, and inflammation, as well as a number of cDNAs lacking homology to previously reported cDNA sequences, were increased >two-fold and were associated with p<0.05. Principal component analysis identified a vector component ( approximately 10% of the total variation) corresponding to a potential EqIL-1beta co-regulation of cell cycle associated gene transcription. These results support and expand our existing comprehension of the complex role of IL-1 in modulated chondrocyte gene expression and suggest the involvement of specific target gene up-regulation and activation of downstream inflammatory cascade mediators. This study adds to the current understanding of the molecular events associated with an IL-1 induced inflammation and pathobiologic processes that may be associated with the development of equine osteoarthritis.
Takafuji
V A
VA
Orthopedic Research Laboratory, VA-MD Regional College of Veterinary Medicine, Blacksburg, VA 24061-0442, USA.
Howard
R D
RD
Ward
D L
DL
Sharova
L V
LV
Crisman
M V
MV
D000818:Animals; D002358:Cartilage, Articular / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000187:drug effects / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D006736:Horses / Q000378:metabolism*; D007375:Interleukin-1 / Q000494:pharmacology / Q000502:physiology*; D020411:Oligonucleotide Array Sequence Analysis / Q000662:veterinary; D012333:RNA, Messenger / Q000378:metabolism*; D011994:Recombinant Proteins / Q000494:pharmacology; D013997:Time Factors
D007375:Interleukin-1; D012333:RNA, Messenger; D011994:Recombinant Proteins