Study on the Inhibitory Effects of Ephedra Aconite Asarum Decoction on LPS-Induced Dendritic Cells

Hindawi Evidence-Based Complementary and Alternative Medicine Volume 2017, Article ID 3272649, 9 pages https://doi.org/10.1155/2017/3272649 Research Article Study on the Inhibitory Effects of Ephedra Aconite Asarum Decoction on LPS-Induced Dendritic Cells Wenting Ji, Jinghong Hu, Xue Yu, Lanlan Zhang, Min Liu, and Qingguo Wang Beijing University of Chinese Medicine, Beijing 100029, China Correspondence should be addressed to Min Liu; and Qingguo Wang; Received 3 July 2017; Revised 4 October 2017; Accepted 9 October 2017; Published 26 November 2017 Academic Editor: Juraj Majtan Copyright © 2017 Wenting Ji et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Dendritic cells (DCs) can secrete cytokines stimulated by lipopolysaccharide (LPS), which leads to not just acute inflammatory responses but also Th1 polarization. Furtherly, chronic inflammation or autoimmune diseases could be triggered. As a classic Traditional Chinese Medicine formula, Ephedra Aconite Asarum Decoction with the main ingredients of ephedrine and hypaconitine can show effect on anti-inflammation and immunoregulation. But it remains unclear whether Ephedra Aconite Asarum Decoction controls DCs. In this study, we investigated the effects of Ephedra Aconite Asarum Decoction on LPS-induced bone marrow-derived DCs (BMDCs) in vitro. We found that Ephedra Aconite Asarum Decoction lowered surface costimulators on DCs and reduced the expression of Th1 type cytokines. Yet it is slightly beneficial for shifting to Th2. Our work reveals that the Ephedra Aconite Asarum Decoction can regulate Th1 inflammation through intervening DCs. 1. Introduction LPS is a bacterial wall component of the gram-negative pathogens and a major component of endotoxin. It can cause tissue infiltration of inflammatory cells, activation of macrophages, endothelial cells and DCs, and release of proinflammatory cytokines such as IL-12, IL-1, and IFN-𝛾, leading to acute inflammation [1–3]. Meanwhile, cytokines including IFN-𝛾 and IL-12 can also promote differentiation of T cells to the Th1 direction [4, 5], while overproduction of Th1 cytokines (such as IL-12 and IFN-𝛾) will lead to chronic inflammation, organ-specific autoimmune responses, and contact dermatitis [6]. DCs are antigen presenting cells (APC) with the strongest antigen presentation ability, which activate T cells and regulate T cell differentiation [7]. DCs are at the center of initiating, regulating, and maintaining immune responses [8]. The function of DCs is closely related to its maturation status. Cell surface expression of CD83, CD80, MHCII, and other costimulators significantly increases upon DCs maturation [9]. In general, the increase of CD83 can be used as a reliable indicator of DCs maturation [10]. Mature DCs play a powerful antigen presenting role to activate T cells and to secrete a variety of cytokines that affect T helper (Th) cell differentiation. Thus, DCs are the key to the pathogenesis of many autoimmune diseases and may also be a main therapeutic target for these diseases [11]. Signal transducers and activators of transcription (STAT) are a family of cytoplasmic proteins with signal transduction and transcriptional activation functions. STAT1, STAT2, STAT3, STAT4, STAT5, and STAT6 are the main members of the STAT family [12, 13]. Except the limited distribution pattern of STAT4, the rest of the STAT proteins are widely distributed among different cell types and tissues. Among them, STAT6 is closely related to Th2 differentiation of the CD4+ T cells [14, 15]. STAT6 can activate GATA-3 through the Il-4-STAT6 pathway and promote T cell differentiation towards Th2 cells. T-bet and GATA-3 are the transcription factors of Th1 and Th2 differentiation, respectively, and they guide the specific differentiation of T cells. In addition, GATA-3 is an upstream molecule to STAT6, so Tbet and GATA-3 together promote preferential differentiation of T cell to Th2 cells. Ephedra Aconite Asarum Decoction was made of three herbs including Ephedra, Aconite, and Asarum. It has the advantages of precise ingredient compatibility and comprehensive efficacy for the targeted treatment of Shaoyin exogenous syndromes based on its effects in supporting, dispelling, 2 Evidence-Based Complementary and Alternative Medicine and invigorating Yang as described in the treatise on Febrile Diseases [16–20]. Ohhashi from Japan claimed that this formula is a “special cure” for AR (Allergic Rhinitis) [21], and there are many case reports and clinical observations from China that also show similar efficacies [22, 23]. Modern pharmacological studies found that Ephedra Aconite Asarum Decoction had broad indications, as it is commonly used in the clinic for the treatment of AR [24], cold [25], arrhythmia [26], asthma [27], and other diseases and showed great efficacy. The major active ingredients of Ephedra Aconite Asarum Decoction include ephedrine, aconite total alkaloids, and Asarum volatile oil [28, 29] among others. Studies have shown that ephedrine, aconite total alkaloids, and Asarum volatile oil had anti-inflammatory and immunomodulatory effects [30, 31]. In this study, we investigated the impact of Ephedra Aconite Asarum Decoction on LPS-induced DCs to explore the effects of this composite formula in LPS-induced inflammatory model. into 96-well plate at 100 𝜇l per well, add 10x series dilution of 1 g/ml Ephedra Aconite Asarum Decoction water extracts into 96-well plate, make hexplicate of each concentration of water extract, place the cell culture plate in 37∘ C, 5% CO2 , saturated humidity incubator for 24 hours, then add 10 𝜇l Cell Counting Kit-8 (CCK-8) reagent to each well, read on microplate reader every half hour, and record the value; next, adjust concentration of the water extract and repeat the experiment until an appropriate concentration of Ephedra Aconite Asarum Decoction was chosen for subsequent experiments. 2. Methods 2.1. Animals and Materials. Twenty SPF male C57BL/6 mice of 6 weeks old (20 ± 5 g) were purchased from SPF (Beijing) Biotechnology Co., Ltd. All mice were maintained under specific pathogen-free conditions. 2.2. Preparation of Ephedra Aconite Asarum Decoction Water Extract. Ephedra Aconite Asarum Decoction was prepared as follows (usual dose for adults): Ephedra 6 g, Aconite 9 g, and Asarum 3 g (this dose is the commonly used clinical dose), boil with the right drug to water ratio in ultrapure water for 1 hour until final drug concentration reaches 1 g/ml. Centrifuge at high speed for 2 times, 12,000 rpb/min, 10 min each, collect supernatant, filter with 0.22 𝜇m filter for sterilization, aliquot, and store at −20∘ C for future use. 2.3. Isolation and Culture of DCs from Mouse Bone Marrow. Methods for DCs isolation and culture from C57BL/6J mice which were raised in the Animal Laborator (...truncated)