This study was conducted to define the underlying molecular mechanism of tripartite motif (TRIM) 59-induced invasion of ectopic endometrial stromal cells in endometriosis. Primary endometriosis ectopic endometrial stromal cells and normal endometrial cells were isolated and purified. Western blot was used to detect the expression of TRIM59, protein phosphatase Mg²⁺/Mn²⁺-dependent 1A (PPM1A), smad2/3, and phosphorylated (p)-smad2/3. Lentiviral vector-mediated TRIM59 interference and overexpression were established. Cell Counting Kit-8 assay was used to detect cell proliferation, and the Transwell migration assay was used to detect cell invasion. Matrix metalloproteinase (MMP-2), MMP9, smad2/3, and p-smad2/3 expressions were also detected using Western blot analysis; degradation of PPM1A was verified to be through ubiquitination. We found that TRIM59 expression levels in the endometriosis group was significantly higher compared with the normal group (P < 0.05), whereas the expression levels of PPM1A in the endometriosis group were significantly lower (P < 0.05). Endometriosis did not alter smad2/3 (P > 0.05) expression. However, after activating smad2/3 by phosphorylation, the expression of p-smad2/3 in the endometriosis group was significantly higher compared with the normal group (P < 0.05). The content of PPM1A in the TRIM59 overexpression group was significantly lower than that in the control group (P < 0.001), whereas the content of PPM1A in the siTRIM59 group was significantly higher than that in the control group (P < 0.001). In addition, there were no significant differences in the mRNA levels of PPM1A among the five groups, indicating that TRIM59 affects the expression of PPM1A at the posttranslational level (P < 0.05). Overexpression of TRIM59 significantly promoted the ubiquitination of PPM1A. We conclude that TRIM59 inhibits PPM1A through ubiquitination and activates the transforming growth factor-β/Smad pathway to promote the invasion of ectopic endometrial stromal cells in endometriosis.

中文翻译：

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TRIM59通过泛素化抑制PPM1A，并激活TGF-β/ Smad信号传导，促进子宫内膜异位症中异位子宫内膜基质细胞的侵袭。

进行这项研究来确定三方基序（TRIM）59诱导子宫内膜异位症异位子宫内膜间质细胞入侵的潜在分子机制。分离并纯化原发性子宫内膜异位症异位子宫内膜间质细胞和正常子宫内膜细胞。用Western blot检测TRIM59，蛋白磷酸酶Mg ²⁺ / Mn ²⁺的表达。-依赖性1A（PPM1A），smad2 / 3和磷酸化（p）-smad2 / 3。建立了慢病毒载体介导的TRIM59干扰和过表达。使用Cell Counting Kit-8检测法检测细胞增殖，并使用Transwell迁移检测法检测细胞侵袭。还使用蛋白质印迹分析检测了基质金属蛋白酶（MMP-2），MMP9，smad2 / 3和p-smad2 / 3的表达。证实PPM1A的降解是通过泛素化。我们发现，子宫内膜异位症组中TRIM59的表达水平明显高于正常组（P <0.05），而子宫内膜异位症组中PPM1A的表达水平则显着较低（P <0.05）。子宫内膜异位症并未改变smad2 / 3（P> 0.05）的表达式。然而，子宫内膜异位症组通过磷酸化激活smad2 / 3后，p-smad2 / 3的表达明显高于正常组（P <0.05）。TRIM59过表达组的PPM1A含量显着低于对照组（P <0.001），而siTRIM59组的PPM1A含量显着高于对照组（P <0.001）。此外，五组中PPM1A的mRNA水平没有显着差异，表明TRIM59在翻译后水平上影响PPM1A的表达（P<0.05）。TRIM59的过表达显着促进了PPM1A的泛素化。我们得出的结论是，TRIM59通过泛素化抑制PPM1A，并激活转化生长因子-β/ Smad途径，以促进子宫内膜异位症中异位子宫内膜基质细胞的侵袭。