6-(7-Nitro-2,1,3-benzoxadiazol-4-ylthio) Hexanol Inhibits Proliferation and Induces Apoptosis of Endometriosis by Regulating Glutathione S-Transferase Mu Class 4

Endometriosis is a chronic disease associated with a disrupted oxidative balance and chronic inflammation. In this study, we investigated the role of glutathione S-transferase Mu class 4 (GSTM4) in endometriosis and determined whether 6-(7-nitro-2,1,3-benzoxadiazol-4-ylthio) hexanol (NBDHEX) regulates GSTM4 expression to affect cellular functions and oxidative stress. GSTM4 expression was detected by immunohistochemistry in endometrium from 15 endometriosis patients and 15 healthy controls. Western blotting was used to detect the expression of GSTM4, proliferating cell nuclear antigen (PCNA), matrix metalloproteinase-9 (MMP-9), Survivin, B-cell lymphoma-extra-large (Bcl-XL), Bax, kelch-like ECH-associated protein 1 (Keap1), and nuclear factor-erythroid 2-related factor 2 (Nrf2) in primary endometrial stromal cells with endometriosis (EESC) and normal endometrial stromal cells (NESC). The effects of NBDHEX on cell proliferation, migration, and invasion were evaluated using Cell Counting Kit-8 (CCK8) and Transwell assays. Apoptosis was detected by flow cytometry. The expression of GSTM4 was significantly increased in endometrium from endometriosis patients. Upon NBDHEX treatment, ESC exhibited reduced proliferation, migration and invasion abilities, and increased apoptosis. NBDHEX decreased the expression of endometriosis prognostic markers (PCNA and MMP-9) and anti-apoptotic proteins (Survivin and Bcl-xl), while it increased the expression of the apoptotic protein Bax. It had no effect on Keap1 expression, and it decreased the expression of Nrf2. The effect of siRNA-mediated knockdown of GSTM4 was similar to that of suppressing GSTM4 expression with NBDHEX treatment. These results indicate that GSTM4 is highly expressed in endometriosis and its expression is inhibited by NBDHEX. Decreased expression of GSTM4 inhibits cell growth, migration, and invasion, and negatively regulates Nrf2 to affect oxidative stress–induced apoptosis. Our results suggest that GSTM4 may play a role in ameliorating the progression of endometriosis. NBDHEX may have therapeutic potential in the treatment of endometriosis. Similar content being viewed by others Data Availability All data generated or analyzed during this study will be provided on request.

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The authors wish to thank the patients for providing clinical samples, and all investigators in this study. We thank Liwen Bianji (Edanz) (https://www.liwenbianji.cn) for editing the language of a draft of this manuscript. Funding This work was supported by the National Natural Science Foundation of China (82071617) and the Basic Research Program of Shanxi Province(202203021212106). Author information Authors and Affiliations Contributions Lei Yan and Lei Cheng contributed to conception and design of the study. Wei Liu and Yanbo Du performed the experiments. Xiaoqiang Liu and Wei Liu performed the statistical analysis. Wei Liu and Lei Yan wrote the first draft of the manuscript. Lei Cheng and Jinlong Ma contributed to manuscript revision. All authors approved the submitted version. Corresponding author Ethics declarations Ethics Approval The study was reviewed and approved by the Reproductive Hospital Affiliated to Shandong University (registration number 2020–14). Informed consent was obtained from each patient. Competing Interests The authors declare no competing interests. Additional information Publisher's Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Rights and permissions Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law. About this article Cite this article Liu, W., Cheng, L., Du, Y. et al. 6-(7-Nitro-2,1,3-benzoxadiazol-4-ylthio) Hexanol Inhibits Proliferation and Induces Apoptosis of Endometriosis by Regulating Glutathione S-Transferase Mu Class 4. Reprod. Sci. 30, 2945–2961 (2023). https://doi.org/10.1007/s43032-023-01207-x Received: Accepted: Published: Version of record: Issue date: DOI: https://doi.org/10.1007/s43032-023-01207-x