The Affiliated Hospital of Medical School, Ningbo University, Ningbo, China.
Ningbo University, Ningbo, China.
J Clin Lab Anal. 2020 Nov;34(11):e23482. doi: 10.1002/jcla.23482. Epub 2020 Aug 5.
Endometriosis (EMS) is a prevalent gynecological condition characterized by the growth of endometrial tissue outside the uterine cavity. This study aimed to clarify the targeted therapeutic effect of sunitinib in an endometriosis in vitro experiment.
Primary culture of ectopic endometrial cells and normal endometrial cells. Six tumor targeting drugs were selected to screen. MTT was used to determine the IC50, flow cytometry, and DAPI staining of the targeted drugs, in order to determine the apoptosis. The differential proteins after seeding were analyzed by protein spectrum, the correlation between the specific protein and cell apoptosis was determined by small molecule interference, and the expression of each related protein was detected by Western blot. Immunohistochemistry and ELISA were used to detect the expression of p-PDGFR and p-STAT1 in clinical samples, and the correlation between p-STAT1 expression and ectopic focal size was analyzed by SPSS 19.
Through the drug screening, it was found that sunitinib has a significant inhibitory effect on ectopic endometrial cells. It was determined that the IC50 of sunitinib on ectopic stromal endometrial cells was 3.32 μM, while the IC50 on normal endometrium was 7.9 μM. Meanwhile, the flow cytometry and DAPI nuclear dye that took out sunitinib had an inhibition effect on the ectopic endometrium at a concentration of 4 μM. Protein spectrum analysis was conducted on ectopic intimal cells after sunitinib treatment, and it was found that STAT1 is specifically expressed in ectopic endometrial cells. In vitro, and through fludarabine interference, it was revealed that sunitinib specifically inhibited the phosphorylation site Tyr751 of PDGFR, while the expression of STAT1, p-STAT1, and caspase-3 was significantly upregulated, and the expression of STAT1 and p-STAT1 was positively correlated with the expression of caspase-3. Finally, the expression of p-PDGFR and p-STAT1 in ectopic foal tissues was both higher than that in normal endometrium, and p-STAT1 expression was positively with ectopic focal size.
The in vitro experiments revealed that sunitinib could upregulate the expression of STAT1 by inhibiting the phosphorylation site Tyr751 of PDGFR, thereby specifically inducing the apoptosis of the primary heterotopic mesenchymal endometrium.
子宫内膜异位症（EMS）是一种常见的妇科疾病，其特征是子宫内膜组织在子宫腔外生长。本研究旨在阐明舒尼替尼在子宫内膜异位症的体外实验中的靶向治疗效果。
原代培养异位子宫内膜细胞和正常子宫内膜细胞。选择六种肿瘤靶向药物进行筛选。采用 MTT 法测定 IC50，流式细胞术和 DAPI 染色检测靶向药物的凋亡作用。通过蛋白谱分析，分析接种后差异蛋白，通过小分子干扰确定特定蛋白与细胞凋亡的相关性，通过 Western blot 检测各相关蛋白的表达。采用免疫组化和 ELISA 检测临床样本中 p-PDGFR 和 p-STAT1 的表达，采用 SPSS 19 分析 p-STAT1 表达与异位灶大小的相关性。
通过药物筛选发现舒尼替尼对异位子宫内膜细胞有明显的抑制作用。确定舒尼替尼对异位基质子宫内膜细胞的 IC50 为 3.32μM，而对正常子宫内膜的 IC50 为 7.9μM。同时，在 4μM 浓度下，取出舒尼替尼对异位子宫内膜也有抑制作用。对舒尼替尼处理后的异位内膜细胞进行蛋白谱分析，发现 STAT1 特异性表达于异位子宫内膜细胞。在体外，并通过氟达拉滨干扰，发现舒尼替尼特异性抑制 PDGFR 的 Tyr751 磷酸化位点，而 STAT1、p-STAT1 和 caspase-3 的表达明显上调，且 STAT1 和 p-STAT1 的表达与 caspase-3 的表达呈正相关。最后，异位病灶组织中 p-PDGFR 和 p-STAT1 的表达均高于正常子宫内膜，p-STAT1 表达与异位灶大小呈正相关。
体外实验表明，舒尼替尼通过抑制 PDGFR 的 Tyr751 磷酸化位点，上调 STAT1 的表达，从而特异性诱导原代异位间充质子宫内膜细胞凋亡。
