Catalpol ameliorates LPS-induced endometritis by inhibiting inflammation and TLR4/NF-κB signaling
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Catalpol is the main active ingredient of an extract from Radix rehmanniae, which in a previous study showed a protective effect against various types of tissue injury. However, a protective effect of catalpol on uterine inflammation has not been reported. In this study, to investigate the protective mechanism of catalpol on lipopolysaccharide (LPS)-induced bovine endometrial epithelial cells (bEECs) and mouse endometritis, in vitro and in vivo inflammation models were established. The Toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signaling pathway and its downstream inflammatory factors were detected by enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (qRT-PCR), western blot (WB), and immunofluorescence techniques. The results from ELISA and qRT-PCR showed that catalpol dose-dependently reduced the expression of pro-inflammatory cytokines such as tumor necrosis factor α (TNF-α), interleukin (IL)-1β, and IL-6, and chemokines such as C-X-C motif chemokine ligand 8 (CXCL8) and CXCL5, both in bEECs and in uterine tissue. From the experimental results of WB, qRT-PCR, and immunofluorescence, the expression of TLR4 and the phosphorylation of NF-κB p65 were markedly inhibited by catalpol compared with the LPS group. The inflammatory damage to the mouse uterus caused by LPS was greatly reduced and was accompanied by a decline in myeloperoxidase (MPO) activity. The results of this study suggest that catalpol can exert an anti-inflammatory impact on LPS-induced bEECs and mouse endometritis by inhibiting inflammation and activation of the TLR4/NF-κB signaling pathway.
KeywordsCatalpol Endometritis Inflammation Toll-like receptor 4 (TLR4) Nuclear factor-KB (NF-κB)
梓醇通过抑制炎症反应和 TLR4/NF-κB 信号通路改善脂多糖诱导的小鼠子宫内膜炎
首次证明梓醇对 LPS 刺激的牛子宫内膜上皮细胞炎症和 LPS 诱导的小鼠子宫内膜炎具有保护作用, 其保护机制与抑制 Toll 样受体 4/核因子 κB (TLR4/NF-κB) 炎症信号通路有关。
通过 LPS 的诱导, 分别建立牛子宫内膜上皮细胞体外炎症模型和小鼠子宫内膜体内炎症模型, 设置不同梓醇作用浓度梯度, 采用酶联免疫吸附测定法 (ELISA)、 实时荧光定量聚合酶链式反应 (qRT0PCR)、 蛋白免疫印迹 (western blot) 和免疫荧光技术检测 TLR4/NF-κB 信号通路及其下游炎症因子的表达。
梓醇可以显著抑制 TLR4 和 p65 NF-κB 信号通路的表达, 降低炎性因子肿瘤坏死因子 α (TNF-α)、 白细胞介素 1β (IL-1β) 和白细胞介素 6 (IL-6) 的水平以及趋化因子 CXCL8 和 CXCL5 的表达, 同时降低子宫组织髓过氧化物酶水平。 通过在体内外炎症模型中加入梓醇, 可以显著降低牛子宫内膜上皮细胞的炎症反应, 有效保护小鼠体内子宫内膜的组织损伤。
关键词梓醇 子宫内膜炎 炎症 Toll 样受体 4/核因子 κB (TLR4/NF-κB)
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Hua ZHANG and Zhi-min WU designed and performed this study. Ya-ping YANG, Jing YANG, and Ying-fang GUO assisted in carrying out the research. Aftab SHAUKAT assisted in editing language. Tao ZHANG, Xin-ying ZHU, and Jin-xia QIU assisted in analyzing the data. Gan-zhen DENG and Dong-mei SHI proofread the manuscript. All authors have approved the final manuscript. Therefore, all authors have full access to all the data in the study and take responsibility for the integrity and security of the data.
Compliance with ethics guidelines
Hua ZHANG, Zhi-min WU, Ya-ping YANG, Aftab SHAUKAT, Jing YANG, Ying-fang GUO, Tao ZHANG, Xin-ying ZHU, Jin-xia QIU, Gan-zhen DENG, and Dong-mei SHI declare that they have no conflicts of interest.
All institutional and national guidelines for the care and use of laboratory animals were followed. The animal experiments were carried out according to the guidelines of the Laboratory Animal Research Center of Hubei Province and approved by the Ethical Committee on Animal Research at Huazhong Agricultural University (HZAUMO-2015-12), Wuhan, China.
- Brodzki P, Kostro K, Brodzki A, et al., 2015. Inflammatory cytokines and acute-phase proteins concentrations in the peripheral blood and uterus of cows that developed endometritis during early postpartum. Theriogenology, 84(1): 11–18. https://doi.org/10.1016/j.theriogenology.2015.02.006 CrossRefGoogle Scholar
- Chen BY, Jiang LX, Hao K, et al., 2018. Protection of plasma transfusion against lipopolysaccharide/D-galactosamine-induced fulminant hepatic failure through inhibiting apoptosis of hepatic cells in mice. J Zhejiang Univ-Sci B (Biomed & Biotechnol), 19(6):436–444. https://doi.org/10.1631/jzus.B1700277 CrossRefGoogle Scholar
- Chen Z, Deng S, Yuan DC, et al., 2018. Novel nano-microspheres containing chitosan, hyaluronic acid, and chondroitin sulfate deliver growth and differentiation factor-5 plasmid for osteoarthritis gene therapy. J Zhejiang Univ-Sci B (Biomed & Biotechnol), 19(12): 910–923. https://doi.org/10.1631/jzus.B1800095 CrossRefGoogle Scholar
- Feng XN, Zhang DF, Li XY, et al., 2018. CXCL5, the upregulated chemokine in patients with uterine cervix cancer, in vivo and in vitro contributes to oncogenic potential of Hela uterine cervix cancer cells. Biomed Pharmacother, 107:1496–1504. https://doi.org/10.1016/j.biopha.2018.08.149 CrossRefGoogle Scholar
- Karlsson I, Hagman R, Guo YZ, et al., 2015. Pathogenic Escherichia coli and lipopolysaccharide enhance the expression of IL-8, CXCL5, and CXCL10 in canine endometrial stromal cells. Theriogenology, 84(1): 34–42. https://doi.org/10.1016/j.theriogenology.2015.02.008 CrossRefGoogle Scholar
- Koyama T, Omori R, Koyama K, et al., 2018. Optimization of diagnostic methods and criteria of endometritis for various postpartum days to evaluate infertility in dairy cows. Theriogenology, 119:225–232. https://doi.org/10.1016/j.theriogenology.2018.07.002 CrossRefGoogle Scholar
- Nicholas C, Batra S, Vargo MA, et al., 2007. Apigenin blocks lipopolysaccharide-induced lethality in vivo and proinflammatory cytokines expression by inactivating NF-KB through the suppression of p65 phosphorylation. J Immunol, 179(10):7121–7127. https://doi.org/10.4049/jimmunol.179.10.7121 CrossRefGoogle Scholar
- Socha BM, Lada P, et al., 2018. The influence of experimentally induced endometritis on the PPAR expression profile in the bovine endometrium. Theriogenology, 122: 74–83. https://doi.org/10.1016/j.theriogenology.2018.09.013 CrossRefGoogle Scholar
- Turner ML, Cronin JG, et al., 2014. Epithelial and stromal cells of bovine endometrium have roles in innate immunity and initiate inflammatory responses to bacterial lipopeptides in vitro via Toll-like receptors TLR2, TLR1, and TLR6. Endocrinology, 155(4): 1453–1465. https://doi.org/10.1210/en.2013-1822 CrossRefGoogle Scholar
- van Schyndel SJ, Pascottini OB, et al., 2018. Comparison of cow-side diagnostic techniques for subclinical endometritis in dairy cows. Theriogenology, 120:117–122. https://doi.org/10.1016/j.theriogenology.2018.08.001 CrossRefGoogle Scholar
- Wagener K, Gabler C, Drillich M, 2017. A review of the ongoing discussion about definition, diagnosis and pathomecha-nism of subclinical endometritis in dairy cows. Theriogenology, 94:21–30. https://doi.org/10.1016/j.theriogenology.2017.02.005 CrossRefGoogle Scholar
- Ye GY, Wang KY, Gui QD, et al., 2018. Ureaplasma urealyticum-derived lipid-associated membrane proteins introduce IL-6, IL-8, and TNF-α cytokines into human amniotic epithelial cells via Toll-like receptor 2}. J Zhejiang Univ-Sci B (Biomed & Biotechnol), 19(8):654–661. https://doi.org/10.1631/jzus.B1800005 CrossRefGoogle Scholar