Institute of Pathology, University of Regensburg, Franz-Josef-Strauss-Allee 11, 93053 Regensburg, Germany.
Arthritis Res Ther. 2010;12(2):R45. doi: 10.1186/ar2955. Epub 2010 Mar 18.
The repellent factor family of Slit molecules has been described to have repulsive function in the developing nervous system on growing axons expressing the Robo receptors. However, until today no data are available on whether these repellent factors are involved in the regulation of synovial fibroblast (SF) activity in rheumatoid arthritis (RA).
mRNA expression in primary synovial fibroblasts was quantified by quantitative reverse transcription PCR and protein expression was measured by fluorescence activated cell sorting (FACS) analysis. Different functional assays were performed with rheumatoid arthritis synovial fibroblasts (RASF): proliferation, migration and a novel in-vitro cartilage destruction assay.
First, we found increased expression of Robo3 expression in RASF compared to normal SF. Interestingly, analysis of data from a recently published genome-wide association study suggests a contribution of ROBO3 gene polymorphisms to susceptibility of RA. Functional assays performed with RASF revealed induction of migration and cartilage destruction by Robo3 and increased matrix metalloproteinase (MMP)1 and MMP3 expression. Treatment of RASF in early passages with Slit3 led to inhibition of migration whereas RASF in later passages, having reduced Robo3 expression in cell culture, were not inhibited by Slit3 treatment. Here, reduction of Robo3 expression from passage 3 to 10 might reflect an important step in losing repulsive activity of Slit3.
Taken together, our data showed that deregulation of the Robo3 receptor in synovial fibroblasts in RA correlates with aggressiveness of the fibroblasts. Slit3 reduces the migratory activity of synovial cells from patients with RA, potentially by repulsion of the cells in analogy to the neuronal system. Further studies will be necessary to prove Slit activity in vivo.
Slit 分子的驱避因子家族已被描述为在表达 Robo 受体的生长轴突的发育神经系统中具有驱避功能。然而，直到今天，尚无数据表明这些驱避因子是否参与类风湿关节炎（RA）中滑膜成纤维细胞（SF）活性的调节。
通过定量逆转录 PCR 定量检测原代滑膜成纤维细胞中的 mRNA 表达，并通过荧光激活细胞分选（FACS）分析测量蛋白表达。对类风湿关节炎滑膜成纤维细胞（RASF）进行了不同的功能测定：增殖、迁移和新型体外软骨破坏测定。
首先，我们发现 RASF 中 Robo3 表达增加，而正常 SF 中则没有。有趣的是，对最近发表的全基因组关联研究数据的分析表明，ROBO3 基因多态性对 RA 的易感性有贡献。对 RASF 进行的功能测定显示，Robo3 诱导迁移和软骨破坏，并增加基质金属蛋白酶（MMP）1 和 MMP3 的表达。早期传代的 RASF 用 Slit3 处理会抑制迁移，而在细胞培养中 Robo3 表达降低的晚期传代的 RASF 则不受 Slit3 处理的抑制。这里，从第 3 代到第 10 代 Robo3 表达的减少可能反映了 Slit3 失去驱避活性的重要步骤。
总之，我们的数据表明，RA 中滑膜成纤维细胞中 Robo3 受体的失调与成纤维细胞的侵袭性相关。Slit3 通过类比神经元系统对细胞的排斥作用，减少 RA 患者滑膜细胞的迁移活性。进一步的研究将有必要证明 Slit 在体内的活性。
