Abstract: Objective To investigate the effect of halofuginone (HF) on proliferation and apoptosis of endometriotic stromal cells ( ESCs) in endometriosis ( EMs) by regulation of macrophage polarization through peroxisome proliferator-activated receptor gamma ( PPARG) -leukemiainhibitory factor ( LIF) axis. Methods Normal endometrial stromal cells ( NESCs) and ectopicendometrial stromal cells ( EESCs) were treated with different concentrations of HF. CCK8 assay was used to detect the cell proliferation activity, and TUNEL assay was used to detect the cell apoptosis. Bioinformatics analysis was performed to identify the intersection of targets of HF, EMs, and macrophage. A co-culture system of macrophages and EESCs was established, cells were then divided into 7 groups: M0 + EESCs group、 M2 + EESCs group、 M2 + EESCsHF group、 M0 + EESCs oe-PPARG group、 M0 + EESCssh-PPARG group、 M0 + EESCsoe-PPARG + oe-LIF group、 M0 + EESCsoe-PPARG + HF group. ELISA was used to detect the concentrations of M1 macrophage markers (iNOS, IL-6) and M2 macrophage markers (Arg-1, TGF-β) in the cell supernatant. CCK8 and TUNEL assay wereused to detect the proliferation and apoptosis of EESCs, respectively. Results HF had no significant effect on the proliferation activity of NESCs (F = 0. 195, P = 0. 959), but significantly inhibited the proliferation of EESCs and promoted cell apoptosis (P< 0. 05). The levels of iNOS and IL-6 in the M2 + EESCs group were decreased when compared with those in the M0 + EESCs group,while the levels of Arg-1 and TGF-β were increased. In addition, the proliferation activity of EESCswas increased and the apoptosis was decreased. HF intervention reversed the results above ( P <0. 05). PPARG was identified as a key target in this study through bioinformatics analysis and literature search. The levels of iNOS and IL-6 in the cell supernatant of the M0 + EESCsoe-PPARG group weredecreased, while the levels of Arg-1 and TGF-β were increased when compared with those in the M0 + EESCs group. The proliferation activity of EESCs were increased and apoptosis were decreased(P< 0. 05), while an opposite results were obtained in the M0 + EESCssh-PPARG group (P < 0. 05).Furthermore, upregulation of LIF expression or HF intervention in the M0 + EESCsoe-PPARG + oe-LIF andM0 + EESCsoe-PPARG + HF groups led to the increase of iNOS and IL-6 levels and proliferation activity ofEESCs, and the decrease of Arg-1 and TGF-β and apoptosis when compared with the M0 + EESCsoe-PPARGgroup (P < 0. 05). Conclusion HF inhibits M2 polarization of macrophages and inhibitsthe proliferation of EESCs while promoting cell apoptosis by regulating the PPARG-LIF signalingpathway.

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