pubmed.ncbi.nlm.nih.gov

Adipose-derived exosomal miR-421 targets CBX7 and promotes metastatic potential in ovarian cancer cells - PubMed

  • ️Sun Jan 01 2023

Adipose-derived exosomal miR-421 targets CBX7 and promotes metastatic potential in ovarian cancer cells

Yi Zhang et al. J Ovarian Res. 2023.

Abstract

Background: Chromobox protein homolog 7 (CBX7), a member of the Polycomb repressor complex, is a potent epigenetic regulator and gene silencer. Our group has previously reported that CBX7 functions as a tumor suppressor in ovarian cancer cells and its loss accelerated formation of carcinomatosis and drove tumor progression in an ovarian cancer mouse model. The goal of this study is to identify specific signaling pathways in the ovarian tumor microenvironment that down-regulate CBX7. Given that adipocytes are an integral component of the peritoneal cavity and the ovarian tumor microenvironment, we hypothesize that the adipose microenvironment is an important regulator of CBX7 expression.

Results: Using conditioned media from human omental explants, we found that adipose-derived exosomes mediate CBX7 downregulation and enhance migratory potential of human ovarian cancer cells. Further, we identified adipose-derived exosomal miR-421 as a novel regulator of CBX7 expression and the main effector that downregulates CBX7.

Conclusion: In this study, we identified miR-421 as a specific signaling pathway in the ovarian tumor microenvironment that can downregulate CBX7 to induce epigenetic change in OC cells, which can drive disease progression. These findings suggest that targeting exosomal miR-421 may curtail ovarian cancer progression.

© 2023. The Author(s).

PubMed Disclaimer

Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1

Adipose conditioned media (ACM) downregulate CBX7 and enhance migratory capacity in ovarian cancer cells. A panel of human ovarian cancer cells were cultured for 5 days in ACM from omental explants isolated from a female patient with a benign gynecological condition. No treatment control cells (NT) were cultured in growth media. A Levels of CBX7 were determined by Western blot analysis; B Representative microscopic images show acquisition of a fibroblastic spindle-like morphology; C Western blot analysis for EMT markers in OCSC1-F2 human ovarian cancer cells; D top panel, OCSC1-F2 human ovarian cancer cells were cultured in ACM or growth media for 5 days, trypsinized, washed, and placed in trans-well inserts with growth media. Note enhanced migration in cells previously cultured with ACM; bottom panel, quantification of migration (n = 8); *** p < 0.0001. Data are presented as mean ± SEM. Unpaired t-test was used to determine statistical significance

Fig. 2
Fig. 2

Characterization of exosomes derived from adipose conditioned media (ACM). Exosomes were isolated from ACM by differential ultracentrifugation. A Representative particle size distribution (ave = 137 nm) of ACM-derived exosomes analyzed by Nanoparticle Tracking Analysis; B Representative transmission electron microscopy image showing the doughnut-shaped morphology of the isolated exosomes; C Representative Western blot analysis of ACM-derived exosomes showing expression of exosomal markers CD63 and CD9 and absence of the endoplasmic reticulum marker, calnexin; Protein lysate of Adipose-Derived Mesenchymal Stem Cells (ADMSC) is used as positive control of calnexin. D Representative images from ExoView analysis showing majority of captured exosomes are positive for the adipocyte marker, CD36, and negative for the macrophage marker, CD11b; E Quantification of D and showing data from 6 individual patients. Data are presented as mean ± SEM (n = 6). Unpaired t-test was used to determine statistical significance. Data shown are for exosomes 654 and 652. Similar results were observed with other exosomal preparations

Fig. 3
Fig. 3

Adipose-derived exosomes are internalized by human ovarian cancer cells via endocytosis and induce down-regulation of CBX7. A Labelled ACM-derived exosomes were internalized by mCherry-labelled OCSC1-F2 cells; B Human R182 ovarian cancer cells were treated for 48 h with adipose-derived exosomes (5 × 108 exosomes/ml) isolated from patients in Table 1. top panel, levels of CBX7 determined by Western blot analysis. * denotes preparations able to down-regulate CBX7. bottom panel, densitometry quantification of top panel; C OCSC1-F2 human ovarian cancer cells were treated with ACM, exosomes (exo), exosome-depleted ACM (ACM-exo) or exosome with Nystatin and levels of CBX7 determined by Western blot analysis; (D) OVCAR3 human ovarian cancer cells were treated with ACM, exosomes (exo), or exosome-depleted ACM (ACM-exo) and levels of CBX7 determined by Western blot analysis

Fig. 4
Fig. 4

Adipose-derived exosomal miR-421 targets CBX7. A Venn diagram analysis from predicted miRNAs targeting CBX7 from mirTarBase (Table 2) and TargetScanHuman (Table 3) and lists of adipocyte specific miRNA from Adicer KO mice and patients with congenital lipodystrophy (CGL) [64] shows miR-421 as the only common element in the overlap; B OCSC1-F2 human ovarian cancer cells were treated with adipose-derived exosomes (exo) in the presence of anti-miR-421 or anti-miR negative control (-ve con Anti-mir) and effect on CBX7 was determined by Western blot analysis; NT, no treatment control (C) OCSC1-F2 human ovarian cancer cells were treated with miR-421 mimic and the effect on CBX7 was determined by Western blot analysis; Neg, non-specific miRNA; D plasmid design for luciferase reporter system carrying CBX7 3’ UTR predicted to be the binding site for miR-421 (pCBX7) and its mutated version (pCBX7mut). Sequence of human miR-421 (hsa-mir-421) is also shown; E Luciferase reporters were transfected in OCSC1-F2 human OC cells in the presence or absence of miR-421 or non-specific miRNA (Neg) as indicated, and levels of luciferase were measured as surrogate of binding affinity; F miR-421 was quantified in adipose- derived exosomes using qPCR. Note that exosomes successful in downregulating CBX7 (columns with red data points) contain higher levels of mir-421. Data are presented as mean ± SEM (n = 3). Ordinary One way ANOVA with Tukey’s multiple comparison test was used to determine statistical significance

Update of

Similar articles

Cited by

References

    1. Jin MZ, Jin WL. The updated landscape of tumor microenvironment and drug repurposing. Signal Transduct Target Ther. 2020;5(1):166. doi: 10.1038/s41392-020-00280-x. - DOI - PMC - PubMed
    1. Farc O, Cristea V. An overview of the tumor microenvironment, from cells to complex networks (Review) Exp Ther Med. 2021;21(1):96. doi: 10.3892/etm.2020.9528. - DOI - PMC - PubMed
    1. Mbeunkui F, Johann DJ., Jr Cancer and the tumor microenvironment: a review of an essential relationship. Cancer Chemother Pharmacol. 2009;63(4):571–582. doi: 10.1007/s00280-008-0881-9. - DOI - PMC - PubMed
    1. Yao H, He S. Multifaceted role of cancerassociated adipocytes in the tumor microenvironment (Review) Mol Med Rep. 2021;24(6):866. doi: 10.3892/mmr.2021.12506. - DOI - PMC - PubMed
    1. Dumas JF, Brisson L. Interaction between adipose tissue and cancer cells: role for cancer progression. Cancer Metastasis Rev. 2021;40(1):31–46. doi: 10.1007/s10555-020-09934-2. - DOI - PubMed

MeSH terms

Substances