TET3-overexpressing macrophages promote endometriosis
Haining Lv, … , Hugh S. Taylor, Yingqun Huang
Haining Lv, … , Hugh S. Taylor, Yingqun Huang
Published August 14, 2024
Citation Information: J Clin Invest. 2024;134(21):e181839. https://doi.org/10.1172/JCI181839.
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Research Article Inflammation Reproductive biology

TET3-overexpressing macrophages promote endometriosis

Abstract

Endometriosis is a debilitating, chronic inflammatory disease affecting approximately 10% of reproductive-age women worldwide with no cure. While macrophages have been intrinsically linked to the pathophysiology of endometriosis, targeting them therapeutically has been extremely challenging due to their high heterogeneity and because these disease-associated macrophages (DAMs) can be either pathogenic or protective. Here, we report identification of pathogenic macrophages characterized by TET3 overexpression in human endometriosis lesions. We show that factors from the disease microenvironment upregulated TET3 expression, transforming macrophages into pathogenic DAMs. TET3 overexpression stimulated proinflammatory cytokine production via a feedback mechanism involving inhibition of let-7 miRNA expression. Remarkably, these cells relied on TET3 overexpression for survival and hence were vulnerable to TET3 knockdown. We demonstrated that Bobcat339, a synthetic cytosine derivative, triggered TET3 degradation in both human and mouse macrophages. This degradation was dependent on a von Hippel-Lindau (VHL) E3 ubiquitin ligase whose expression was also upregulated in TET3-overexpressing macrophages. Furthermore, depleting TET3-overexpressing macrophages either through myeloid-specific Tet3 ablation or using Bobcat339 strongly inhibited endometriosis progression in mice. Our results defined TET3-overexpressing macrophages as key pathogenic contributors to and attractive therapeutic targets for endometriosis. Our findings may also be applicable to other chronic inflammatory diseases where DAMs have important roles.

Authors

Haining Lv, Beibei Liu, Yangyang Dai, Feng Li, Stefania Bellone, Yuping Zhou, Ramanaiah Mamillapalli, Dejian Zhao, Muthukumaran Venkatachalapathy, Yali Hu, Gordon G. Carmichael, Da Li, Hugh S. Taylor, Yingqun Huang

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Figure 5

TET3 knockdown reduces macrophage expression of IL-1β and IL-6.

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TET3 knockdown reduces macrophage expression of IL-1β and IL-6. (A) qRT-...
(A) qRT-PCR of Il1b and Il6 mRNAs of cultured PMs isolated from Mye-Tet3–KO mice or WT controls and treated with 10 ng/mL LPS plus 20 ng/mL IFN-γ. RNAs were isolated after 6 hours of LPS/IFN-γ stimulation. Uns, unstimulated. n = 3 mice per genotype. (B) ELISA analysis (after 6 hours of LPS/IFN-γ stimulation) of IL-1β and IL-6 of cultured PMs treated as in A. n = 3 mice per genotype. (C) Human MDMs primed with 10 ng/mL of TGF-β1 were transfected with NT siRNA or TET3 siRNA. After 48 hours of transfection, cells were stimulated with 10 ng/mL LPS plus 20 ng/mL IFN-γ for 8 hours, followed by RNA extraction and qRT-PCR of IL-1B and IL-6 mRNAs. n = 3 biological replicates. (D) ELISA analysis (after 8 hours of LPS/IFN-γ stimulation) of IL-1β and IL-6 of MDMs following treatment as in C. (E) Human MDMs were infected with Ad-GFP or Ad-TET3. The next day, proteins were extracted, followed by Western blot analysis. The Ad-TET3–infected cells showed approximately 5-fold TET3 overexpression as compared with Ad-GFP–infected cells. (F) Human MDMs were infected with Ad-GFP or Ad-TET3. The next day, cells were stimulated with 10 ng/mL LPS plus 20 ng/mL IFN-γ for 8 hours, followed by RNA extraction and qRT-PCR of IL-1B and IL-6 mRNAs. n = 3 biological replicates. (G) ELISA analysis (after 8 hours of LPS/IFN-γ stimulation) of IL-1β and IL-6 of MDMs following treatment as in F. All data are represented as mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001, 2-tailed Student’s t test.