Stradner Martin H, Gruber Gerald, Angerer Hannes, Huber Verena, Setznagl Daniela, Kremser Marie-Luise, Moazedi-Fürst Florentine C, Windhager Reinhard, Graninger Winfried B
University of California at San Diego, La Jolla, California; Medical University of Graz, Graz, Austria.
Arthritis Rheum. 2013 Aug;65(8):2113-22. doi: 10.1002/art.37989.
The lipid mediator sphingosine 1-phosphate (S1P) is found in the synovial fluid of osteoarthritis (OA) patients. S1P protects bovine cartilage by counteracting the effects of interleukin-1β (IL-1β). This study was undertaken to examine the interaction of S1P and IL-1β in human OA chondrocytes.
Human cartilage was obtained from patients undergoing total knee joint replacement. Chondrocytes were cultured in monolayer and treated with IL-1β and S1P. Expression of S1P receptor subtypes and genes involved in cartilage degradation was evaluated using real-time polymerase chain reaction, immunohistochemistry, and Western blotting. S1P signaling was evaluated using inhibitors of S1P receptors and small interfering RNA (siRNA) knockdown of the S1P2 receptor. Phosphorylation of MAP kinases and NF-κB in response to IL-1β and S1P was detected by Western blotting.
S1P2 was identified as the most prevalent S1P receptor subtype in human OA cartilage and chondrocytes in vitro. S1P reduced expression of inducible nitric oxide synthase (iNOS) in IL-1β-treated chondrocytes. Reduction of ADAMTS-4 and matrix metalloproteinase 13 expression by S1P correlated with S1P2 expression. Pharmacologic inhibition of the S1P2 receptor, but not the S1P1 and S1P3 receptors, abrogated the inhibition of iNOS expression. Similar results were observed using siRNA knockdown. S1P signaling inhibited IL-1β-induced phosphorylation of p38 MAPK.
In human chondrocytes, S1P reduces the induction of catabolic genes in the presence of IL-1β. Activation of the S1P2 receptor counteracts the detrimental phosphorylation of p38 MAPK by IL-1β.
脂质介质1-磷酸鞘氨醇（S1P）存在于骨关节炎（OA）患者的滑液中。S1P通过抵消白细胞介素-1β（IL-1β）的作用来保护牛软骨。本研究旨在探讨S1P与IL-1β在人OA软骨细胞中的相互作用。
从接受全膝关节置换术的患者获取人软骨。将软骨细胞进行单层培养，并用IL-1β和S1P处理。使用实时聚合酶链反应、免疫组织化学和蛋白质印迹法评估S1P受体亚型及参与软骨降解的基因的表达。使用S1P受体抑制剂和S1P2受体的小干扰RNA（siRNA）敲低来评估S1P信号传导。通过蛋白质印迹法检测IL-1β和S1P刺激下丝裂原活化蛋白激酶（MAP激酶）和核因子κB（NF-κB）的磷酸化。
S1P2被确定为体外人OA软骨和软骨细胞中最普遍的S1P受体亚型。S1P降低了IL-1β处理的软骨细胞中诱导型一氧化氮合酶（iNOS）的表达。S1P对含血小板解聚蛋白样金属蛋白酶4（ADAMTS-4）和基质金属蛋白酶13表达的降低与S1P2的表达相关。S1P2受体的药理学抑制而非S1P1和S1P3受体的抑制消除了对iNOS表达的抑制。使用siRNA敲低观察到类似结果。S1P信号传导抑制了IL-1β诱导的p38 MAPK磷酸化。
在人软骨细胞中，S1P在存在IL-1β的情况下减少分解代谢基因的诱导。S1P2受体的激活抵消了IL-1β对p38 MAPK的有害磷酸化作用。
