Effects of interleukin-6 and interleukin-1β on expression of growth differentiation factor-5 and Wnt signaling pathway genes in equine chondrocytes.
OBJECTIVE
To determine the effects of interleukin (IL)-6 and IL-1β stimulation on expression of growth differentiation factor (GDF)-5 and Wnt signaling pathway genes in equine chondrocytes.


METHODS
Macroscopically normal articular cartilage samples from 6 horses and osteochondral fragments (OCFs) from 3 horses.


METHODS
Chondrocyte pellets were prepared and cultured without stimulation or following stimulation with IL-6 or IL-1β for 1, 2, 12, and 48 hours; expression of GDF-5 was determined with a quantitative real-time PCR assay. Expression of genes in various signaling pathways was determined with microarrays for pellets stimulated for 1 and 2 hours. Immunohistochemical analysis was used to detect GDF-5, glycogen synthase kinase 3β (GSK-3β), and β-catenin proteins in macroscopically normal cartilage samples and OCFs.


RESULTS
Chondrocytes stimulated with IL-6 had significantly higher GDF-5 expression within 2 hours versus unstimulated chondrocytes. Microarray analysis of Wnt signaling pathway genes indicated expression of GSK-3β and coiled-coil domain containing 88C increased after 1 hour and expression of β-catenin decreased after 2 hours of IL-6 stimulation. Results of immunohistochemical detection of proteins were similar to microarray analysis results. Chondrocytes in macroscopically normal articular cartilage and OCFs had immunostaining for GDF-5.


CONCLUSIONS
Results indicated IL-6 stimulation decreased chondrocyte expression of the canonical Wnt signaling pathway transactivator β-catenin, induced expression of inhibitors of the Wnt pathway, and increased expression of GDF-5. This suggested IL-6 may inhibit the Wnt signaling pathway with subsequent upregulation of GDF-5 expression. Anabolic extracellular matrix metabolism in OCFs may be attributable to GDF-5 expression. This information could be useful for development of cartilage repair methods.
Department of Biomedical Sciences and Veterinary Public Health, Division of Pathology, Pharmacology and Toxicology, Swedish University of Agricultural Sciences, 750 07 Uppsala, Sweden.
Emilia
E
Svala
Anna I
AI
Thorfve
Cecilia
C
Ley
Helena K Barreto
HK
Henriksson
Jane M
JM
Synnergren
Anders H
AH
Lindahl
Stina
S
Ekman
Eva S R
ES
Skiöldebrand
D000818:Animals; D002358:Cartilage, Articular / Q000166:cytology / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D038362:Glycogen Synthase Kinase 3 / Q000235:genetics / Q000378:metabolism; D055428:Growth Differentiation Factor 5 / Q000235:genetics / Q000378:metabolism*; D006736:Horses / Q000378:metabolism*; D053583:Interleukin-1beta / Q000494:pharmacology*; D015850:Interleukin-6 / Q000494:pharmacology*; D060449:Wnt Signaling Pathway / Q000187:drug effects* / Q000235:genetics; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism
D055428:Growth Differentiation Factor 5; D053583:Interleukin-1beta; D015850:Interleukin-6; D051176:beta Catenin; D038362:Glycogen Synthase Kinase 3