School of Pharmacy, Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, PR China; Engineering Research Center of Shanghai Colleges for TCM New Drug Discovery, Shanghai, 201203, PR China.
School of Pharmacy, Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, PR China; Engineering Research Center of Shanghai Colleges for TCM New Drug Discovery, Shanghai, 201203, PR China.
J Ethnopharmacol. 2022 Sep 15;295:115396. doi: 10.1016/j.jep.2022.115396. Epub 2022 May 19.
Leonurus japonicus Houtt., also known as motherwort, is a traditional Chinese medicine that was first identified in Shennong Bencao Jing, the first and essential pharmacy monograph in China. L. japonicus has been regarded as a good gynecological medicine since ancient times. It has been widely used in clinical settings for treatment of gynecological diseases and postnatal rehabilitation with good efficacy and low adverse effects.
The main purpose of this study was to determine the angiogenic and wound healing effects of total alkaloid fraction from L. japonicus Houtt. (TALH) in vivo and in vitro. In addition, the main bioactive components of total alkaloids were to be identified and analyzed in this study.
First, the UHPLC/Q-TOF-MS method was used to identify and quantify the major components in the TALH extract. The wound healing activity was evaluated in vivo using a rat full-thickness cutaneous wound model. Histological study of wound healing in rat model was performed via immunohistochemistry and immunofluorescence. Cell proliferation was determined by MTT assay. Wound healing and transwell assays were used for detection of cell migration. The effect on tube formation was determined by tube formation assay in HUVECs. Western blot and RT-PCR were used to detect the expressions of relative proteins and genes respectively. Knock-down of SRC by siRNA was done to verify the crucial role of SRC in promotion of angiogenesis induced by TALH.
Seven characteristic peaks were recognized in the UHPLC/Q-TOF-MS spectrum, while four of the main components were quantified. The wound model in rats showed that treatment of TALH promoted wound healing by stimulating cellular proliferation and collagen deposition. In vitro experiments showed that co-treatment of TALH and VEGF increased cell proliferation, migration and tube formation in HUVECs. Mechanistic studies suggested that the co-treatment increased gene expressions of SRC, MEK1/2 and ERK1/2, as well as the phosphorylation levels of these proteins. Furthermore, the effect of co-treatment was attenuated after SRC knockdown, suggesting that SRC plays an important role in angiogenesis and wound healing induced by TALH and VEGF co-treatment.
Our results showed that TALH was one of the main active components of L. japonicus that promoted angiogenesis and wound healing by regulating the SRC/MEK/ERK pathway. Our study provided scientific basis for better clinical application of L. japonicas.
益母草，又名益母，是一种传统中药，最早在中国第一部药学专著《神农本草经》中被发现。自古以来，益母草一直被视为妇科良药。它在临床上被广泛用于治疗妇科疾病和产后康复，疗效好，不良反应低。
本研究的主要目的是确定益母草总生物碱（TALH）在体内和体外的促血管生成和伤口愈合作用。此外，本研究还对总生物碱的主要生物活性成分进行了鉴定和分析。
首先，采用 UHPLC/Q-TOF-MS 法鉴定和定量 TALH 提取物中的主要成分。采用大鼠全层皮肤创面模型评价促伤口愈合活性。通过免疫组织化学和免疫荧光法对大鼠模型的伤口愈合进行组织学研究。采用 MTT 法测定细胞增殖。采用划痕和 Transwell 实验检测细胞迁移。采用 HUVECs 管形成实验检测管形成。采用 Western blot 和 RT-PCR 分别检测相对蛋白和基因的表达。通过 siRNA 敲低 SRC 验证 SRC 在 TALH 诱导的血管生成中的关键作用。
UHPLC/Q-TOF-MS 图谱中识别出 7 个特征峰，同时定量了 4 种主要成分。大鼠创面模型表明，TALH 治疗通过刺激细胞增殖和胶原沉积促进创面愈合。体外实验表明，TALH 与 VEGF 共同处理可增加 HUVEC 细胞的增殖、迁移和管形成。机制研究表明，共同处理增加了 SRC、MEK1/2 和 ERK1/2 的基因表达以及这些蛋白的磷酸化水平。此外，SRC 敲低后共同处理的效果减弱，提示 SRC 在 TALH 和 VEGF 共同处理诱导的血管生成和伤口愈合中发挥重要作用。
我们的研究结果表明，TALH 是益母草的主要活性成分之一，通过调节 SRC/MEK/ERK 通路促进血管生成和伤口愈合。本研究为更好地临床应用益母草提供了科学依据。
