Effects of interleukin-1 on calcium signaling and the increase of filamentous actin in isolated and in situ articular chondrocytes
Effects of interleukin-1 on calcium signaling and the increase of filamentous actin in isolated and in situ articular chondrocytes
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DOI：
10.1002/art.21941
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发表时间：
2006-07-01
影响因子：
--
通讯作者：
Guilak, Farshid
中科院分区：
文献类型：
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作者：
Pritchard, Scott;Guilak, Farshid
Objective.为了确定白细胞介素-1（IL-1）是否启动关节软骨细胞内[Ca 2 +]（i）浓度和丝状肌动蛋白（F-actin）组织的瞬时变化。将软骨外植体内的关节软骨细胞和酶促分离的软骨细胞加载有Ca 2+敏感的荧光指示剂，并在暴露于10 ng/ml IL-1 α期间使用共聚焦荧光比率成像测量[Ca 2 +]（i）。使用Ca 2+动员抑制剂（无Ca 2+培养基、毒胡萝卜素[Ca-ATP酶抑制剂]、U 73122 [磷脂酶C抑制剂]和百日咳毒素[G蛋白抑制剂]）确定[Ca 2 +]增加的机制（i）。使用荧光标记的鬼笔环肽定量细胞F-肌动蛋白。毒素B用于确定小GTP酶Rho家族在F-肌动蛋白重组中的作用。在玻璃上的分离细胞和外植体内的原位软骨细胞中，暴露于IL-1诱导[Ca 2 +]（i）的瞬时峰值，通常随后是一系列衰减振荡。毒胡萝卜素、U 73122和百日咳毒素抑制对IL-1应答的细胞百分比。IL-1以一种被毒素B抑制的方式增加软骨细胞中F-肌动蛋白的含量。分离的和原位软骨细胞均对IL-1产生应答，通过磷脂酶C和三磷酸肌醇途径介导的细胞内Ca 2+释放，[Ca 2 +]（i）瞬时增加。Ca 2+从细胞外空间的流入和G蛋白偶联受体的激活似乎也有助于这些机制。这些结果表明，Ca 2+动员可能是第一个信号事件在。软骨细胞对IL-1的反应。
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 Ca2+-sensitive fluorescence indicators, and [Ca2+](i) was measured using confocal fluorescence ratio imaging during exposure to 10 ng/ml IL-1 alpha. Inhibitors of Ca2+ mobilization (Ca2+-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-I. IL-1 increased F-actin content in chondrocytes in a manner that was inhibited by toxin B.Conclusion. 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.