贝洛酸通过 TLR4、MAPK 和 JAK/STAT 信号通路调节输卵管上皮细胞炎症。

Betulonic acid regulates oviduct epithelial cell inflammation through the TLR4, MAPK, and JAK/STAT signalling pathways.

机构信息

Department of Emergency, The First Affiliated Hospital of Hunan Traditional Chinese Medicine College, Zhuzhou, Hunan, China.

Department of TCM Gynaecology, The First Affiliated Hospital of Hunan Traditional Chinese Medicine College, Zhuzhou, Hunan, China.

出版信息

Reprod Fertil Dev. 2023 May;35(8):480-491. doi: 10.1071/RD21380.

DOI:10.1071/RD21380

PMID:37142241

Abstract

CONTEXT

Infertility is a common disease among women of childbearing age and seriously endangers the reproductive health of human beings.

AIMS

We aimed to study the active effect and mechanism of betulonic acid (BTA) on tubal inflammatory infertility.

METHODS

An inflammatory model was established in isolated rat oviduct epithelial cells. Immunofluorescence of cytokeratin 18 was performed in cells. The therapeutic effect of BTA on cells was observed. Subsequently, we added JAK/STAT inhibitor AG490 and MAPK inhibitor U0126 and measured the levels of inflammatory factors via enzyme-linked immunosorbent assay and qRT-PCR. CCK-8 assay was applied to test cell proliferation, whereas flow cytometry was used to measure apoptosis. The levels of TLR4, IκBα, JAK1, JAK2, JAK3, Tyk2, STAT3, p38, ERK and the phosphorylation of p65 were determined by Western blotting.

KEY RESULTS

Betulonic acid inhibited the activation of TLR4 and NF-κB signalling pathways, and significantly downregulated IL-1β, IL-6, and TNF-α, with high doses being the most effective. Furthermore, high-dose BTA promoted the proliferation of oviduct epithelial cells and inhibited apoptosis. In addition, BTA inhibited the activation of JAK/STAT signalling pathway to perform effectively in oviduct epithelial cells inflammation. The addition of AG490 led to the inhibition of the JAK/STAT signalling pathway. BTA also inhibited the activation of MAPK signalling pathway in oviduct epithelial cells inflammation. Under U0126 treatment, the inhibition of proteins in MAPK pathway by BTA was weakened.

CONCLUSIONS

Therefore, BTA inhibited the TLR, JAK/STAT and MAPK signalling pathways.

IMPLICATIONS

Our study provided a new therapeutic strategy for infertility caused by oviduct inflammation.

摘要

背景

不孕症是育龄妇女的常见病，严重危害人类生殖健康。

目的

研究齐墩果酸（BTA）对输卵管炎性不孕的作用及机制。

方法

建立大鼠离体输卵管上皮细胞炎症模型，细胞免疫荧光法检测细胞角蛋白 18，观察 BTA 对细胞的治疗作用。然后加入 JAK/STAT 抑制剂 AG490 和 MAPK 抑制剂 U0126，通过酶联免疫吸附试验和 qRT-PCR 检测炎症因子水平。CCK-8 法检测细胞增殖，流式细胞术检测细胞凋亡。Western blot 检测 TLR4、IκBα、JAK1、JAK2、JAK3、Tyk2、STAT3、p38、ERK 和 p65 的磷酸化水平。

主要结果

齐墩果酸抑制 TLR4 和 NF-κB 信号通路的激活，显著下调 IL-1β、IL-6 和 TNF-α，高剂量作用最明显。此外，高剂量 BTA 促进输卵管上皮细胞增殖，抑制细胞凋亡。另外，BTA 抑制 JAK/STAT 信号通路的激活，对输卵管上皮细胞炎症有效。加入 AG490 导致 JAK/STAT 信号通路被抑制。BTA 还抑制 MAPK 信号通路在输卵管上皮细胞炎症中的激活。在 U0126 处理下，BTA 对 MAPK 通路蛋白的抑制作用减弱。