Wang J J, Hua Q, Li H J, Zhang D M, Wang Y L
Department of Obstetrics and Gynecology, Zhengzhou Central Hospital Affiliated to Zhengzhou University, Zhengzhou 450007, China.
Zhonghua Yi Xue Za Zhi. 2024 Oct 29;104(40):3757-3764. doi: 10.3760/cma.j.cn112137-20240314-00578.
To investigate the repair effect of human umbilical cord blood mesenchymal stem cells (hUCB-MSC) on endometrium of intrauterine adhesion (IUA) by establishing animal model. Eighteen healthy female New Zealand white rabbits were divided into a control group, an IUA group and an hUCB-MSC transplantation group according to random number table method. The control group underwent laparotomy only. The IUA group underwent IUA modeling surgery using curettage and lipopolysaccharide (LPS) cotton placed in uterine cavity for double injury. The hUCB-MSC transplantation group received the same IUA modeling surgery followed by multipoint injection of hUCB-MSC suspension (2×10 cells/ml, 500 μl) into the bilateral uterine myometrium one week after surgery. Four weeks after the IUA modeling surgery, the rabbits were euthanized and samples were collected. Hematoxylin and eosin (HE) staining and Masson staining were used to assess the number of endometrial glands and the fibrosis rate. RNA sequencing was performed on the endometrium of the IUA group and hUCB-MSC transplantation group.The mRNA and protein expression of the fibrosis markers [transforming growth factor β1 (TGF-β1) and tissue inhibitors of metalloproteinase 1 (TIMP1)] and RNA sequencing-related parameters were detected by quantitative real-time PCR (qRT-PCR) and Western blot. Compared with the control group, the number of glands in IUA group decreased (26.33±1.53 vs 4.33±1.53, <0.001), and the fibrosis rate increased (18.01%±2.21% vs 69.55%±2.42%, <0.001), indicating successful modeling. HE and Masson staining revealed that the number of glands in the hUCB-MSC transplantation group was increased compared with that in IUA group (17.33±2.52 vs 4.33±1.53, <0.001), and the fibrosis rate decreased (69.55%±2.42% vs 41.55%±1.99%,=0.001). Western blot analyses of fibrosis-related proteins (TGF-β1 and TIMP1) showed elevated levels of TGF-β1 (0.91±0.05) and TIMP1 (0.99±0.01) proteins in the IUA group compared to control group (0.61±0.04, 0.68±0.07) (=0.001, 0.015). The hUCB-MSC transplantation group exhibited reduced expression of TGF-β1 (0.69±0.04) and TIMP1 (0.62±0.08) proteins compared to the IUA group (=0.005, 0.014). Western Blot results related to the PI3K/AKT signaling pathway [phosphorylated phosphatidylinositol 3-kinase (p-PI3K), phosphorylated protein kinase B (p-AKT)] showed that the expression of p-PI3K(1.05±0.05) and p-AKT(1.17±0.06) in the IUA group increased compared to the control group (0.78±0.03, 0.85±0.05) (=0.002, 0.002). The expression of p-PI3K (0.74±0.02) and p-AKT (0.93±0.04) proteins in the hUCB-MSC transplantation group decreased compared to the IUA group (=0.003, 0.005). hUCB-MSC can repair the damaged endometrium in rabbit intrauterine adhesion model by increasing the number of endometrial glands and improving its level of fibrosis.
通过建立动物模型，研究人脐带血间充质干细胞（hUCB-MSC）对宫腔粘连（IUA）子宫内膜的修复作用。将18只健康雌性新西兰白兔按随机数字表法分为对照组、IUA组和hUCB-MSC移植组。对照组仅行剖腹手术。IUA组采用刮宫及宫腔放置脂多糖（LPS）棉进行双重损伤建立IUA模型手术。hUCB-MSC移植组在手术后1周接受相同的IUA模型手术，随后将hUCB-MSC悬液（2×10个细胞/ml，500μl）多点注射到双侧子宫肌层。IUA模型手术后4周，对兔子实施安乐死并采集样本。采用苏木精-伊红（HE）染色和Masson染色评估子宫内膜腺体数量和纤维化率。对IUA组和hUCB-MSC移植组的子宫内膜进行RNA测序。通过定量实时聚合酶链反应（qRT-PCR）和蛋白质免疫印迹法检测纤维化标志物[转化生长因子β1（TGF-β1）和基质金属蛋白酶1组织抑制剂（TIMP1）]的mRNA和蛋白表达以及RNA测序相关参数。与对照组相比，IUA组腺体数量减少（26.33±1.53对4.33±1.53，<0.001），纤维化率增加（18.01%±2.21%对69.55%±2.42%，<0.001），表明建模成功。HE和Masson染色显示，hUCB-MSC移植组的腺体数量比IUA组增加（17.33±2.52对4.33±1.53，<0.001），纤维化率降低（69.55%±2.42%对41.55%±1.99%，=0.001）。对纤维化相关蛋白（TGF-β1和TIMP1）进行蛋白质免疫印迹分析显示，与对照组（0.61±0.04，0.68±0.07）相比，IUA组TGF-β1（0.91±0.05）和TIMP1（0.99±0.01）蛋白水平升高（=0.001，0.015）。与IUA组相比，hUCB-MSC移植组TGF-β1（0.69±0.04）和TIMP1（0.62±0.08）蛋白表达降低（=0.005，0.014）。与PI3K/AKT信号通路相关的蛋白质免疫印迹结果[磷酸化磷脂酰肌醇3激酶（p-PI3K）、磷酸化蛋白激酶B（p-AKT）]显示，与对照组（0.78±0.03，0.85±0.05）相比，IUA组p-PI3K（1.05±0.05）和p-AKT（1.17±0.06）表达增加（=0.002，0.002）。与IUA组相比，hUCB-MSC移植组p-PI3K（0.74±0.02）和p-AKT（0.93±0.04）蛋白表达降低（=0.003，0.005）。hUCB-MSC可通过增加子宫内膜腺体数量和改善纤维化程度修复兔宫腔粘连模型中受损的子宫内膜。
