Abstract:
The (patho)physiological function of the sphingolipids ceramide-1-phosphate (C1P), sphingosine-1-phosphate (S1P), and sphingosylphosphorylcholine (SPC) in articular joints during osteoarthritis (OA) is largely unknown. Therefore, we investigated the influence of these lipids on protein expression by fibroblast-like synoviocytes (FLSs) from OA knees. Cultured human FLSs (n = 7) were treated with 1 of 3 lipid species-C1P, S1P, or SPC-IL-1β, or with vehicle. The expression of individual proteins was determined by tandem mass tag peptide labeling followed by high-resolution electrospray ionization (ESI) mass spectrometry after liquid chromatographic separation (LC-MS/MS/MS). The mRNA levels of selected proteins were analyzed using RT-PCR. The 3sphingolipids were quantified in the SF of 18 OA patients using LC-MS/MS. A total of 4930 proteins were determined using multiplex MS, of which 136, 9, 1, and 0 were regulated both reproducibly and significantly by IL-1β, C1P, S1P, and SPC, respectively. In the presence of IL-1ß, all 3 sphingolipids exerted ancillary effects. Only low SF levels of C1P and SPC were found. In conclusion, the 3 lipid species regulated proteins that have not been described in OA. Our results indicate that charged multivesicular body protein 1b, metal cation symporter ZIP14, glutamine-fructose-6-P transaminase, metallothionein-1F and -2A, ferritin, and prosaposin are particularly interesting proteins due to their potential to affect inflammatory, anabolic, catabolic, and apoptotic mechanisms.
摘要:
鞘脂神经酰胺-1-磷酸(C1P)的(病理)生理功能,鞘氨醇-1-磷酸(S1P),骨性关节炎(OA)期间关节中的鞘氨酰磷酸胆碱(SPC)在很大程度上是未知的。因此,我们研究了这些脂质对OA膝关节成纤维细胞样滑膜细胞(FLSs)蛋白表达的影响。培养的人FLS(n=7)用3种脂质种类C1P中的1种处理,S1P,或SPC-IL-1β,或与车辆。通过串联质量标签肽标记,然后在液相色谱分离(LC-MS/MS/MS)后通过高分辨率电喷雾电离(ESI)质谱测定单个蛋白质的表达。使用RT-PCR分析所选蛋白质的mRNA水平。使用LC-MS/MS对18名OA患者的SF中的3种鞘脂进行定量。使用多重MS测定了总共4930种蛋白质,其中136、9、1和0受到IL-1β的可再现和显着调节,C1P,S1P,和SPC,分别。在IL-1β的存在下,所有3种鞘脂都发挥了辅助作用。仅发现低SF水平的C1P和SPC。总之,3类脂质调节OA中尚未描述的蛋白质。我们的结果表明,带电多囊体蛋白1b,金属阳离子同向转运蛋白ZIP14,谷氨酰胺-果糖-6-P转氨酶,金属硫蛋白-1F和-2A,铁蛋白,和proaposin是特别有趣的蛋白质,因为它们有可能影响炎症,合成代谢,分解代谢,和凋亡机制。