Department of Chinese Medicine and Mitochondrial Research Unit, Chang Gung Memorial Hospital-Kaohsiung Medical Center, Chang Gung University College of Medicine, Kaohsiung, Taiwan.
Division of General Surgery, Ministry of Health and Welfare Pingtung Hospital, Pingtung, Taiwan.
J Ethnopharmacol. 2017 Jan 20;196:213-224. doi: 10.1016/j.jep.2016.12.019. Epub 2016 Dec 16.
Davallia bilabiata Hosokawa (D. bilabiata), also called GuSuiBu, is popularly used as a substitute for Drynaria fortunei J. Sm for rheumatoid and degenerative arthritis in traditional Chinese medicine. Little is known about the underlying mechanisms of anti-angiogenesis responsible for arthritis in D. bilabiata which needs to be elucidated.
The present study is intended to investigate the anti-angiogenic effect of D. bilabiata associated with the modulation of matrix metalloproteinases (MMPs) and down regulation of vascular endothelial growth factor (VEGF) ligand/receptors both in vivo and in vitro.
We investigated the potential anti-angiogenic effect of D. bilabiata by the in vivo neovascularization of chick chorioallantoic membranes (CAM) assay, and the in vitro migration and matrix-induced tube formation assay using human umbilical vascular endothelial cells (HUVECs). The expressions of MMP-2, TIMP-2, RECK and VEGF/VEGFR were analyzed by real-time RT-PCR or Western blot method.
One major compound from water extract of D. bilabiata was identified as Epicatechin 3-O-β-D-allopyranoside. D. bilabiata was confirmed to inhibit in vivo angiogenesis by CAM assay. D. bilabiata also exhibited in vitro anti-angiogenic and anti-regrowth effects as demonstrated by tube formation assay, transwell migration assay and wound healing assay. The mRNA expressions of MMP-2, and MMP-14 were decreased. On the contrary, tissue inhibitor of metalloproteinase-2 (TIMP-2), reversion-inducing cysteine-rich protein with kazal motifs (RECK) were increased by D. bilabiata. The extracellular MMP-2 activity was found to be reduced both in vitro and in vivo by D. bilabiata as determined by gelatin zymography. Results from western blot analysis and ELISA further demonstrated the decrease of MMP-2 and increase of TIMP-2 secretion after D. bilabiata treatment. The gene expressions of VEGF-A, -B, -C, -D and VEGFR-1, -2, -3 were all inhibited by D. bilabiata.
We concluded that the anti-angiogenic effect of D. bilabiata was associated with the decreased MMP-2 activity mediated by the upregulation of TIMP-2 and RECK, and the suppression of VEGF/VEGFRs expression.
凤尾蕨（D. bilabiata），也称为骨碎补，在传统中药中被广泛用作类风湿和退行性关节炎的替代品。目前还不太清楚凤尾蕨治疗关节炎的抗血管生成的潜在机制，需要进一步研究。
本研究旨在探讨凤尾蕨的抗血管生成作用及其与基质金属蛋白酶（MMPs）的调节以及血管内皮生长因子（VEGF）配体/受体的下调相关，包括体内和体外实验。
我们通过鸡胚绒毛尿囊膜（CAM）试验研究了凤尾蕨的潜在抗血管生成作用，通过体外迁移和基质诱导的管形成试验研究了人脐静脉内皮细胞（HUVECs）的迁移和基质诱导的管形成试验。通过实时 RT-PCR 或 Western blot 方法分析 MMP-2、TIMP-2、RECK 和 VEGF/VEGFR 的表达。
从凤尾蕨水提取物中鉴定出一种主要化合物为表儿茶素 3-O-β-D-吡喃葡萄糖苷。CAM 试验证实凤尾蕨能抑制体内血管生成。凤尾蕨还表现出体外抗血管生成和抗生长作用，如管形成试验、Transwell 迁移试验和划痕愈合试验所示。MMP-2 和 MMP-14 的 mRNA 表达降低。相反，基质金属蛋白酶抑制剂-2（TIMP-2）和富含半胱氨酸的 Kazal 基序的反转诱导蛋白（RECK）增加。通过明胶酶谱法测定，体外和体内均发现凤尾蕨能降低细胞外 MMP-2 活性。Western blot 分析和 ELISA 结果进一步表明，凤尾蕨处理后 MMP-2 减少，TIMP-2 分泌增加。凤尾蕨还抑制了 VEGF-A、-B、-C、-D 和 VEGFR-1、-2、-3 的基因表达。
我们得出的结论是，凤尾蕨的抗血管生成作用与 MMP-2 活性的降低有关，这是由 TIMP-2 和 RECK 的上调以及 VEGF/VEGFRs 表达的抑制介导的。
