Effects of interleukin-1 on calcium signaling and the increase of filamentous actin in isolated and in situ articular chondrocytes.
OBJECTIVE
To determine whether interleukin-1 (IL-1) initiates transient changes in the intracellular concentration of [Ca2+]i and the organization of filamentous actin (F-actin) in articular chondrocytes.


METHODS
Articular chondrocytes within cartilage explants and enzymatically isolated chondrocytes were loaded with Ca(2+)-sensitive fluorescence indicators, and [Ca2+]i was measured using confocal fluorescence ratio imaging during exposure to 10 ng/ml IL-1alpha. Inhibitors of Ca2+ mobilization (Ca(2+)-free medium, thapsigargin [inhibitor of Ca-ATPases], U73122 [inhibitor of phospholipase C], and pertussis toxin [inhibitor of G proteins]) were used to determine the mechanisms of increased [Ca2+]i. Cellular F-actin was quantified using fluorescently labeled phalloidin. Toxin B was used to determine the role of the Rho family of small GTPases in F-actin reorganization.


RESULTS
In isolated cells on glass and in in situ chondrocytes within explants, exposure to IL-1 induced a transient peak in [Ca2+]i that was generally followed by a series of decaying oscillations. Thapsigargin, U73122, and pertussis toxin inhibited the percentage of cells responding to IL-1. IL-1 increased F-actin content in chondrocytes in a manner that was inhibited by toxin B.


CONCLUSIONS
Both isolated and in situ chondrocytes respond to IL-1 with transient increases in [Ca2+]i via intracellular Ca2+ release mediated by the phospholipase C and inositol trisphosphate pathways. The influx of Ca2+ from the extracellular space and the activation of G protein-coupled receptors also appear to contribute to these mechanisms. These findings suggest that Ca2+ mobilization may be one of the first signaling events in the response of chondrocytes to IL-1.
Pritchard
Scott
S
Duke University Medical Center, 375 Medical Sciences Research Building, Research Drive, Durham, NC 27710, USA.
Guilak
Farshid
F
D000199:Actins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001426:Bacterial Proteins / Q000494:pharmacology; D001427:Bacterial Toxins / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects* / Q000502:physiology*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002358:Cartilage, Articular / Q000737:chemistry / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D019902:Chondrocytes / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D004963:Estrenes / Q000494:pharmacology; D005260:Female; D020558:GTP Phosphohydrolases / Q000502:physiology; D007375:Interleukin-1 / Q000494:pharmacology* / Q000502:physiology; D010003:Osteoarthritis / Q000503:physiopathology; D037342:Pertussis Toxin / Q000494:pharmacology; D011760:Pyrrolidinones / Q000494:pharmacology; D013552:Swine; D019284:Thapsigargin / Q000494:pharmacology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors
D000199:Actins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004963:Estrenes; D007375:Interleukin-1; D011760:Pyrrolidinones; C031815:toxB protein, Clostridium difficile; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D019284:Thapsigargin; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D020558:GTP Phosphohydrolases; D000252:Calcium-Transporting ATPases