Advertisement

Journal of Assisted Reproduction and Genetics

, Volume 36, Issue 11, pp 2385–2397 | Cite as

SATB2 and NGR1: potential upstream regulatory factors in uterine leiomyomas

  • Shun Sato
  • Ryo Maekawa
  • Isao Tamura
  • Yuichiro Shirafuta
  • Masahiro Shinagawa
  • Hiromi Asada
  • Toshiaki Taketani
  • Hiroshi Tamura
  • Norihiro SuginoEmail author
Reproductive Physiology and Disease
  • 70 Downloads

Abstract

Purpose

We attempted to identify the genes involved in the pathogenesis of uterine leiomyomas, under a hypothesis that the aberrant expression of upstream regulatory genes caused by aberrant DNA methylation is involved in the onset and development of uterine leiomyomas.

Methods

To find such genes, we compared genome-wide mRNA expression and DNA methylation in uterine leiomyomas and adjacent normal myometrium. Analysis of the data by Ingenuity Pathway Analysis software identified SATB2 which is known to be an epigenetic regulator, and NRG1 as candidate upstream regulatory genes. To infer the functions of these genes, human uterine smooth muscle cell lines overexpressing SATB2 or NRG1 genes were established (SATB2 or NRG1 lines), and their transcriptomes and pathways were analyzed.

Results

SATB2 and NRG1 were confirmed to be hypermethylated and upregulated in most uterine leiomyoma specimens (nine to 11 of the 11 cases). Among the established cell lines, morphological changes from spindle-like forms to fibroblast-like forms with elongated protrusions were observed in only the SATB2 line. Pathway analysis revealed that WNT/β-catenin and TGF-β signaling pathways which are related to the pathogenesis of uterine leiomyomas were activated in both SATB2 and NRG1 lines. In addition, signaling of growth factors including VEGF, PDGF, and IGF1, and retinoic acid signaling were activated in the SATB2 and NRG1 lines, respectively.

Conclusions

These results indicate that SATB2 and NRG1 overexpression induced many of the signaling pathways that are considered to be involved in the pathogenesis of uterine leiomyomas, suggesting that these genes have roles as upstream regulatory factors.

Keywords

Uterine leiomyomas Upstream regulatory genes SATB2 NGR1 

Notes

Acknowledgments

We would like to thank Drs. Ikuo Konishi and Noriomi Matsumura (Kyoto University, and Kindai University, respectively) for providing us human immortalized uterine smooth muscle cells (hTERT UtSMCs). This work was supported in part by JSPS KAKENHI Grants 15K10720, 16K11142, 16K20191, 17K11240, 17K11239, and 16K11091 for Scientific Research from the Ministry of Education, Science, and Culture, Japan.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical statement

This study was approved by the Institutional Review Board of Yamaguchi University Graduate School of Medicine. Informed consent was obtained from the patients before the collection of any samples. All of the experiments handling human tissues were performed in accordance with Tenets of the Declaration of Helsinki.

Supplementary material

10815_2019_1582_MOESM1_ESM.pdf (287 kb)
ESM 1 (PDF 287 kb)

References

  1. 1.
    Stewart EA. Uterine fibroids. Lancet. 2001;357:293–8.  https://doi.org/10.1016/S0140-6736(00)03622-9.CrossRefPubMedGoogle Scholar
  2. 2.
    Bajekal N, Li TC. Fibroids, infertility and pregnancy wastage. Hum Reprod Update. 2000;6:614–20.CrossRefGoogle Scholar
  3. 3.
    Borahay MA, Al-Hendy A, Kilic GS, Boehning D. Signaling pathways in leiomyoma: understanding pathobiology and implications for therapy. Mol Med. 2015;21:242–56.  https://doi.org/10.2119/molmed.2014.00053.CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Ishikawa H, Ishi K, Serna VA, Kakazu R, Bulun SE, Kurita T. Progesterone is essential for maintenance and growth of uterine leiomyoma. Endocrinology. 2010;151:2433–42.  https://doi.org/10.1210/en.2009-1225.CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Ciebiera M, Włodarczyk M, Wrzosek M, Męczekalski B, Nowicka G, Łukaszuk K, et al. Role of transforming growth factor b in uterine fibroid biology. Int J Mol Sci. 2017;18:e 2435.  https://doi.org/10.3390/ijms18112435.CrossRefGoogle Scholar
  6. 6.
    Peng L, Wen Y, Han Y, Wei A, Shi G, Mizuguchi M, et al. Expression of insulin-like growth factors (IGFs) and IGF signaling: molecular complexity in uterine leiomyomas. Fertil Steril. 2009;91:2664–75.  https://doi.org/10.1016/j.fertnstert.2007.10.083.CrossRefPubMedGoogle Scholar
  7. 7.
    Ren Y, Yin H, Tian R, Cui L, Zhu Y, Lin W, et al. Different effects of epidermal growth factor on smooth muscle cells derived from human myometrium and from leiomyoma. Fertil Steril. 2011;96:1015–20.  https://doi.org/10.1016/j.fertnstert.2011.07.004.CrossRefPubMedGoogle Scholar
  8. 8.
    Suo G, Jiang Y, Cowan B, Wang JY. Platelet-derived growth factor C is upregulated in human uterine fibroids and regulates uterine smooth muscle cell growth. Biol Reprod. 2009;81:749–58.  https://doi.org/10.1095/biolreprod.109.076869.CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Chang CC, Hsieh YY, Lin WH, Lin CS. Leiomyoma and vascular endothelial growth factor gene polymorphisms: a systematic review. Taiwan J Obstet Gynecol. 2010;49:247–53.  https://doi.org/10.1016/S1028-4559(10)60056-3.CrossRefPubMedGoogle Scholar
  10. 10.
    Helmke BM, Markowski DN, Müller MH, Sommer A, Müller J, Möller C, et al. HMGA proteins regulate the expression of FGF2 in uterine fibroids. Mol Hum Reprod. 2011;17:135–42.  https://doi.org/10.1093/molehr/gaq083.CrossRefPubMedGoogle Scholar
  11. 11.
    Zaitseva M, Vollenhoven BJ, Rogers PA. Retinoids regulate genes involved in retinoic acid synthesis and transport in human myometrial and fibroid smooth muscle cells. Hum Reprod. 2008;23:1076–86.  https://doi.org/10.1093/humrep/den083.CrossRefPubMedGoogle Scholar
  12. 12.
    Faerstein E, Szklo M, Rosenshein N. Risk factors for uterine leiomyoma: a practice-based case-control study. I. African-American heritage, reproductive history, body size, and smoking. Am J Epidemiol. 2001;153:1–10.CrossRefGoogle Scholar
  13. 13.
    Faerstein E, Szklo M, Rosenshein NB. Risk factors for uterine leiomyoma: a practice-based case-control study. II. Atherogenic risk factors and potential sources of uterine irritation. Am J Epidemiol. 2001;153:11–9.CrossRefGoogle Scholar
  14. 14.
    Chiaffarino F, Parazzini F, La Vecchia C, Chatenoud L, Di Cintio E, Marsico S. Diet and uterine myomas. Obstet Gynecol. 1999;94:395–8.PubMedGoogle Scholar
  15. 15.
    Asada H, Yamagata Y, Taketani T, Matsuoka A, Tamura H, Hattori N, et al. Potential link between estrogen receptor-alpha gene hypomethylation and uterine fibroid formation. Mol Hum Reprod. 2008;14:539–45.  https://doi.org/10.1093/molehr/gan045.CrossRefPubMedGoogle Scholar
  16. 16.
    Yamagata Y, Maekawa R, Asada H, Taketani T, Tamura I, Tamura H, et al. Aberrant DNA methylation status in human uterine leiomyoma. Mol Hum Reprod. 2009;15:259–67.  https://doi.org/10.1093/molehr/gap010.CrossRefPubMedGoogle Scholar
  17. 17.
    Maekawa R, Yagi S, Ohgane J, Yamagata Y, Asada H, Tamura I, et al. Disease-dependent differently methylated regions (D-DMRs) of DNA are enriched on the X chromosome in uterine leiomyoma. J Reprod Dev. 2011;57:604–12.CrossRefGoogle Scholar
  18. 18.
    Maekawa R, Sato S, Yamagata Y, Asada H, Tamura I, Lee L, et al. Genome-wide DNA methylation analysis reveals a potential mechanism for the pathogenesis and development of uterine leiomyomas. PLoS One. 2013;8:e66632.  https://doi.org/10.1371/journal.pone.0066632.CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Navarro A, Yin P, Monsivais D, Lin SM, Du P, Wei JJ, et al. Genome-wide DNA methylation indicates silencing of tumor suppressor genes in uterine leiomyoma. PLoS One. 2012;7:e33284.  https://doi.org/10.1371/journal.pone.0033284.CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Miyata T, Sonoda K, Tomikawa J, Tayama C, Okamura K, Maehara K, et al. Genomic, Epigenomic, and transcriptomic profiling towards identifying omics features and specific biomarkers that distinguish uterine leiomyosarcoma and leiomyoma at molecular levels. Sarcoma. 2015;2015:412068.  https://doi.org/10.1155/2015/412068.CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Yang Q, Mas A, Diamond MP, Al-Hendy A. The mechanism and function of epigenetics in uterine leiomyoma development. Reprod Sci. 2016;23:163–75.  https://doi.org/10.1177/1933719115584449.CrossRefPubMedGoogle Scholar
  22. 22.
    Takahashi K, Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell. 2006;126:663–76.  https://doi.org/10.1016/j.cell.2006.07.024.CrossRefPubMedGoogle Scholar
  23. 23.
    Vierbuchen T, Ostermeier A, Pang ZP, Kokubu Y, Südhof TC, Wernig M. Direct conversion of fibroblasts to functional neurons by defined factors. Nature. 2010;463:1035–41.  https://doi.org/10.1038/nature08797.CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Tomasetti C, Marchionni L, Nowak MA, Parmigiani G, Vogelstein B. Only three driver gene mutations are required for the development of lung and colorectal cancers. Proc Natl Acad Sci U S A. 2015;112:118–23.  https://doi.org/10.1073/pnas.1421839112.CrossRefPubMedGoogle Scholar
  25. 25.
    Matsumura N, Mandai M, Miyanishi M, Fukuhara K, Baba T, Higuchi T, et al. Oncogenic property of acrogranin in human uterine leiomyosarcoma: direct evidence of genetic contribution in in vivo tumorigenesis. Clin Cancer Res. 2006;12:1402–11.  https://doi.org/10.1158/1078-0432.CCR-05-2003.CrossRefPubMedGoogle Scholar
  26. 26.
    Malik M, Catherino WH. Development and validation of a three-dimensional in vitro model for uterine leiomyoma and patient-matched myometrium. Fertil Steril. 2012;97:1287–93.  https://doi.org/10.1016/j.fertnstert.2012.02.037.CrossRefPubMedGoogle Scholar
  27. 27.
    Malik M, Britten J, Segars J, Catherino WH. Leiomyoma cells in 3-dimensional cultures demonstrate an attenuated response to fasudil, a rho-kinase inhibitor, when compared to 2-dimensional cultures. Reprod Sci. 2014;21:1126–38.  https://doi.org/10.1177/1933719114545240.CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Sato S, Maekawa R, Yamagata Y, Asada H, Tamura I, Lee L, et al. Potential mechanisms of aberrant DNA hypomethylation on the x chromosome in uterine leiomyomas. J Reprod Dev. 2014;60:47–54.CrossRefGoogle Scholar
  29. 29.
    Sato S, Maekawa R, Yamagata Y, Tamura I, Lee L, Okada M, et al. Identification of uterine leiomyoma-specific marker genes based on DNA methylation and their clinical application. Sci Rep. 2016;6:30652.  https://doi.org/10.1038/srep30652.CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Dobreva G, Chahrour M, Dautzenberg M, Chirivella L, Kanzler B, Fariñas I, et al. SATB2 is a multifunctional determinant of craniofacial patterning and osteoblast differentiation. Cell. 2006;125:971–86.  https://doi.org/10.1016/j.cell.2006.05.012.CrossRefPubMedGoogle Scholar
  31. 31.
    Gyorgy AB, Szemes M, de Juan Romero C, Tarabykin V, Agoston DV. SATB2 interacts with chromatin-remodeling molecules in differentiating cortical neurons. Eur J Neurosci. 2008;27:865–73.  https://doi.org/10.1111/j.1460-9568.2008.06061.x.CrossRefPubMedGoogle Scholar
  32. 32.
    Brocato J, Costa M. SATB1 and 2 in colorectal cancer. Carcinogenesis. 2015;36:186–91.  https://doi.org/10.1093/carcin/bgu322.CrossRefPubMedGoogle Scholar
  33. 33.
    Magnusson K, de Wit M, Brennan DJ, Johnson LB, McGee SF, Lundberg E, et al. SATB2 in combination with cytokeratin 20 identifies over 95% of all colorectal carcinomas. Am J Surg Pathol. 2011;35:937–48.  https://doi.org/10.1097/PAS.0b013e31821c3dae.CrossRefPubMedGoogle Scholar
  34. 34.
    Patani N, Jiang W, Mansel R, Newbold R, Mokbel K. The mRNA expression of SATB1 and SATB2 in human breast cancer. Cancer Cell Int. 2009;9:18.  https://doi.org/10.1186/1475-2867-9-18.CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Yu W, Ma Y, Shankar S, Srivastava RK. Role of SATB2 in human pancreatic cancer: implications in transformation and a promising biomarker. Oncotarget. 2016;7:57783–97.  https://doi.org/10.18632/oncotarget.10860.CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Steinthorsdottir V, Stefansson H, Ghosh S, Birgisdottir B, Bjornsdottir S, Fasquel AC, et al. Multiple novel transcription initiation sites for NRG1. Gene. 2004;342:97–105.  https://doi.org/10.1016/j.gene.2004.07.029.CrossRefPubMedGoogle Scholar
  37. 37.
    Mei L, Nave KA. Neuregulin-ERBB signaling in the nervous system and neuropsychiatric diseases. Neuron. 2014;83:27–49.  https://doi.org/10.1016/j.neuron.2014.06.007.CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Willem M. Proteolytic processing of Neuregulin-1. Brain Res Bull. 2016;126:178–82.  https://doi.org/10.1016/j.brainresbull.2016.07.003.CrossRefPubMedGoogle Scholar
  39. 39.
    Fernandez-Cuesta L, Thomas RK. Molecular pathways: targeting NRG1 fusions in lung cancer. Clin Cancer Res. 2015;21:1989–94.  https://doi.org/10.1158/1078-0432.CCR-14-0854.CrossRefPubMedGoogle Scholar
  40. 40.
    Kobayashi Y, Nikaido T, Zhai YL, Iinuma M, Shiozawa T, Shirota M, et al. In-vitro model of uterine leiomyomas: formation of ball-like aggregates. Hum Reprod. 1996;11:1724–30.CrossRefGoogle Scholar
  41. 41.
    Bertsch E, Qiang W, Zhang Q, Espona-Fiedler M, Druschitz S, Liu Y, et al. MED12 and HMGA2 mutations: two independent genetic events in uterine leiomyoma and leiomyosarcoma. Mod Pathol. 2014;27:1144–53.  https://doi.org/10.1038/modpathol.2013.243.CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    Mehine M, Kaasinen E, Heinonen HR, Mäkinen N, Kämpjärvi K, Sarvilinna N, et al. Integrated data analysis reveals uterine leiomyoma subtypes with distinct driver pathways and biomarkers. Proc Natl Acad Sci U S A. 2016;113:1315–20.  https://doi.org/10.1073/pnas.1518752113.CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    Ono M, Qiang W, Serna VA, Yin P, Coon JS 5th, Navarro A, et al. Role of stem cells in human uterine leiomyoma growth. PLoS One. 2012;7:e36935.  https://doi.org/10.1371/journal.pone.0036935.CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Obstetrics and GynecologyYamaguchi University Graduate School of MedicineUbeJapan

Personalised recommendations