Department of Gynecology, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei Province, PR China.
Department of Clinical Laboratory, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei Province, PR China.
Reprod Biol. 2024 Jun;24(2):100852. doi: 10.1016/j.repbio.2023.100852. Epub 2024 Feb 14.
Intrauterine adhesions (IUA) refers to endometrial fibrosis caused by irreversible damage of the endometrial basal layer. As the key regulators in tissue repair, regeneration, and fibrosis, macrophages play an essential role in endometrial regeneration and repair during the normal menstrual cycle. However, the mechanism of macrophages involved in IUA remains unclear.
In the late stages of proliferation, the endometrium was collected to make paraffin sections. HE and Masson staining were used to observing endometrial morphology and endometrial fibrosis. Immunohistochemistry and Western blotting were used to detect the expression level of fibrosis indexes COL1A1 and α-SMA. The macrophage infiltration was evaluated by immunohistochemistry for the expression levels of CD 206 and CD163. Next, we cultured the primary human endometrial stromal cells (HESCs), and then an IUA cell model was established with 10 ng/ml TGF-β1 for 72 h. THP 1 cells were differentiated by 100 ng/ml PMA into macrophages for 48 h, then macrophages were polarized to M2 macrophages by 20 ng/ml IL-4 for 24 h. The culture supernatants (M(IL-4) -S) of M2 macrophages were applied to the IUA cell model. The expression of fibrosis markers was then assessed using immunofluorescence and Western blotting.
The results show that Patients with IUA have fewer endometrial glands and significantly increased fibrosis levels. Moreover, the infiltration of CD206-positive (M2) macrophages was significantly reduced in IUA patients, and negatively correlated with the expression of endometrial fibrosis indexes α-SMA and COL1A1. In addition, the primary HESCs treated with 10 ng/ml TGF-β1 for 72 h were found to have significantly increased levels of fibrosis indexes. Furthermore, supernatants from IL4-induced M2 macrophages inhibit the TGF-β1-induced fibrosis of HESCs.
M2 macrophages may negatively regulate the expression of COL1A1 and α-SMA, inhibiting the TGF-β1-induced fibrosis of HESCs. Our study suggests that targeting macrophage phenotypes and promoting the polarization of macrophages to M2 may become a novel strategy for the clinical treatment of IUA.
宫腔粘连（IUA）是指子宫内膜基底层不可逆损伤引起的纤维化。巨噬细胞作为组织修复、再生和纤维化的关键调节因子，在正常月经周期中子宫内膜的再生和修复中发挥着重要作用。然而，巨噬细胞在 IUA 中的作用机制尚不清楚。
在增殖晚期，收集子宫内膜制作石蜡切片。HE 和 Masson 染色观察子宫内膜形态和子宫内膜纤维化。免疫组织化学和 Western blot 检测纤维化指标 COL1A1 和 α-SMA 的表达水平。免疫组织化学检测 CD206 和 CD163 的表达水平评估巨噬细胞浸润。然后，培养原代人子宫内膜基质细胞（HESCs），用 10ng/ml TGF-β1 培养 72h 建立 IUA 细胞模型。用 100ng/ml PMA 将 THP-1 细胞分化为巨噬细胞 48h，然后用 20ng/ml IL-4 将巨噬细胞极化为 M2 巨噬细胞 24h。将 M2 巨噬细胞的培养上清液（M(IL-4)-S）应用于 IUA 细胞模型。然后用免疫荧光和 Western blot 评估纤维化标志物的表达。
结果表明，IUA 患者的子宫内膜腺体较少，纤维化程度显著增加。此外，IUA 患者子宫内膜中 CD206 阳性（M2）巨噬细胞浸润明显减少，与子宫内膜纤维化指标 α-SMA 和 COL1A1 的表达呈负相关。此外，用 10ng/ml TGF-β1 处理 72h 的原代 HESCs 发现纤维化指标水平显著升高。此外，IL4 诱导的 M2 巨噬细胞的上清液抑制 TGF-β1 诱导的 HESCs 纤维化。
M2 巨噬细胞可能负调控 COL1A1 和 α-SMA 的表达，抑制 TGF-β1 诱导的 HESCs 纤维化。我们的研究表明，靶向巨噬细胞表型并促进巨噬细胞向 M2 极化可能成为治疗 IUA 的一种新策略。
