Protective effect of the total flavonoids from Clinopodium chinense against LPS-induced mice endometritis by inhibiting NLRP3 inflammasome-mediated pyroptosis

子宫内膜炎 免疫印迹 上睑下垂 脂多糖 髓过氧化物酶 体内 药理学 TLR4型 生物 染色 传统医学 化学 男科 分子生物学 医学 炎症 免疫学 生物化学 炎症体 病理 怀孕 遗传学 生物技术 基因
作者
Lili Li,Jiajia Qi,Tao Hong,Lele Wang,Lu Wang,Ning Wang,Qi Huang
出处
期刊:Journal of Ethnopharmacology [Elsevier BV]
卷期号:312: 116489-116489 被引量:16
标识
DOI:10.1016/j.jep.2023.116489
摘要

Clinopodium chinense (Benth.) O. Kuntze (C. chinense) is a Chinese herbal medicine used in treating gynecological hemorrhagic diseases for hundreds of years. Flavonoids are one kind of the major components in C. chinense. The flavonoids of C. chinense (TFC) have a vital role in treating endometritis but the underlying therapeutic mechanisms of TFC against endometritis have been rarely reported.To elucidate the therapeutic effect and possible mechanisms of TFC against lipopolysaccharide (LPS)-induced endometritis in vivo and LPS-induced primary mouse endometrial epithelial cells (MEECs) injury in vitro.The holistic phytochemicals of the TFC and TFC-contained serum were screened and identified using UPLC-Q-TOF-MS. The model of endometritis was established by intrauterine injection of LPS (5 mg/mL) into female BALB/c mice, and the model mice were treated with TFC for 7 days. The value of MPO was measured by Myeloperoxidase assay kit, the pathological changes in the endometrium were evaluated using H&E staining and transmission electron microscope (TEM), the secretions of IL-18, IL-1β and TNF-α were determined by ELISA kits, the mRNA expressions of IL-18, IL-1β and TNF-α were determined by RT-PCR assay, and the protein levels of TLR4, IKBα, p-IKBα, p65, p-p65, caspase-1, ASC, NLRP3 and GSDMD were measured by Western blot. Subsequently, MEECs were isolated from the uterus of pregnant female mice, injured by LPS for 24 h and incubated with the TFC-contained serum. Finally, cell viability, LDH release, hoechst 33342/PI staining, immunofluorescence staining, scanning electron microscope observation, ELISA assay, RT-PCR detection and Western blot analysis were carried out to further validate the therapeutic effect and the underlying mechanisms of TFC.A total of 6 compounds in the plasma of mice after being intragastric administrated of TFC were identified. The results in vivo showed that TFC significantly reduced MPO value and alleviated pathological injury of the endometrium. Furthermore, TFC significantly decreased the serum IL-18, IL-1β and TNF-α levels, and the mRNA levels of IL-18, IL-1β and TNF-α. TFC also inhibited the expressions of TLR4, p-IKBα, p-p65, caspase-1, ASC, NLRP3 and GSDMD. Besides, compared with the model group in MEECs cells, TFC-contained serum prevented pyroptosis, decreased the levels of IL-18 and IL-1β, and inhibited the mRNA expressions of IL-18, IL-1β and GSDMD. TFC-contained serum also reversed the activation of NLRP3 inflammasome caused by nigericin, and restrainted the translocation of NF-κB into nuclear.TFC protects mice endometritis from the injury of LPS via suppressing the activation of NLRP3 inflammasome and pyroptosis, the underlying mechanisms of which were related to restraining the TLR4/NF-κB/NLRP3 pathway activation.
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