Department of Oral Biology, University Clinic of Dentistry, Medical University of Vienna, Sensengasse 2a, 1090 Vienna, Austria.
Department of Periodontology, School of Dental Medicine, University of Bern, Freiburgstrasse 7, 3010 Bern, Switzerland.
Int J Mol Sci. 2022 Aug 29;23(17):9778. doi: 10.3390/ijms23179778.
Enamel matrix derivative (EMD) prepared from extracted porcine fetal tooth material can support the regrow of periodontal tissues. Previous findings suggest that EMD has anti-inflammatory properties and TGF-β activity in vitro. However, the anti-inflammatory activity of EMD is mediated via TGF-β has not been considered. To this aim, we first established a bioassay to confirm the anti-inflammatory activity of EMD. The bioassay was based on the RAW 264.7 macrophage cell line and proven with primary macrophages where EMD significantly reduced the forced expression of IL-6. We then confirmed the presence of TGF-β1 in EMD by immunoassay and by provoking the Smad2/3 nuclear translocation in RAW 264.7 macrophages. Next, we took advantage of the TGF-β receptor type I kinase-inhibitor SB431542 to block the respective signalling pathway. SB431542 reversed the anti-inflammatory activity of EMD and TGF-β in a bioassay when IL-6 and CXCL2 expression was driven by the LPS stimulation of RAW 264.7 macrophages. This central observation was supported by showing that SB431542 reversed the anti-inflammatory activity of EMD using IL-1β and TNF-α-stimulated ST2 bone marrow stromal cells. Together, these findings implicate that the TGF-β activity mediates at least part of the anti-inflammatory activity of EMD in vitro.
从提取的猪胎儿牙材料中制备的釉基质衍生物 (EMD) 可以支持牙周组织的再生。先前的研究结果表明，EMD 在体外具有抗炎特性和 TGF-β 活性。然而，尚未考虑 EMD 的抗炎活性是通过 TGF-β 介导的。为此，我们首先建立了一种生物测定法来确认 EMD 的抗炎活性。该生物测定法基于 RAW 264.7 巨噬细胞系，并通过原代巨噬细胞进行了验证，其中 EMD 显著降低了 IL-6 的强制表达。然后，我们通过免疫测定法和在 RAW 264.7 巨噬细胞中引发 Smad2/3 核易位来证实 EMD 中存在 TGF-β1。接下来，我们利用 TGF-β 受体 I 型激酶抑制剂 SB431542 阻断相应的信号通路。当 LPS 刺激 RAW 264.7 巨噬细胞驱动 IL-6 和 CXCL2 表达时，SB431542 在生物测定中逆转了 EMD 和 TGF-β 的抗炎活性。这一主要观察结果得到了支持，表明 SB431542 逆转了使用 IL-1β 和 TNF-α 刺激的 ST2 骨髓基质细胞中 EMD 的抗炎活性。总之，这些发现表明 TGF-β 活性至少部分介导了 EMD 在体外的抗炎活性。
