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Thalidomide induces apoptosis during early mesodermal differentiation of human induced pluripotent stem cells

  • Saoko Tachikawa
  • Maho Shimizu
  • Kenshiro Maruyama
  • Kiyoshi Ohnuma
Article
  • 188 Downloads

Abstract

Thalidomide was once administered to pregnant women as a mild sedative; however, it was subsequently shown to be strongly teratogenic. Recently, there has been renewed interest in thalidomide because of its curative effects against intractable diseases. However, the teratogenicity of thalidomide is manifested in various ways and is still not fully understood. In the present study, we evaluated the effects of thalidomide on early mesodermal differentiation by examining the differentiation of human induced pluripotent stem cells (hiPSCs). The most common symptom of thalidomide teratogenicity is limb abnormality, which led us to hypothesize that thalidomide prevents early mesodermal differentiation. Therefore, mesodermal differentiation of hiPSCs was induced over a 6-d period. To induce early mesoderm differentiation, 1 d after seeding, the cells were incubated with the small molecule compound CHIR99021 for 3 d. Thalidomide exposure was initiated at the same time as CHIR99021 treatment. After 5 d of thalidomide exposure, the hiPSCs began expressing a mesodermal marker; however, the number of viable cells decreased significantly as compared to that of control cells. We observed that the proportion of apoptotic and dead cells increased on day 2; however, the proportion of dead cells on day 5 had decreased, suggesting that the cells were damaged by thalidomide during early mesodermal differentiation (days 0–2). Our findings may help elucidate the mechanism underlying thalidomide teratogenicity and bring us closer to the safe use of this drug.

Keywords

Thalidomide Human induced pluripotent cells Early mesodermal differentiation Teratogenicity 

Notes

Author contributions

S.T. and K.O. designed the project. S.T. performed all experiments. M.S. and K.M. assisted with S.T.’ s experiments. S.T. and K.O. wrote the manuscript, and all authors reviewed the manuscript.

Funding information

This work was supported in part by grants from the Ministry of Health, Labor, and Welfare of Japan, the Japan Agency for Medical Research and Development (AMED) (to K.O.), and the Foundation for Applied Research and Technological Uniqueness at Nagaoka University of Technology (to S.T.). The funding bodies had no role in the study design, data collection and analysis, the decision to publish, or manuscript preparation.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Supplementary material

11626_2018_234_MOESM1_ESM.docx (106 kb)
ESM 1 (DOCX 105 kb)

References

  1. Aikawa N, Kunisato A, Nagao K, Kusaka H, Takaba K, Ohgami K (2014) Detection of thalidomide embryotoxicity by in vitro embryotoxicity testing based on human iPS cells. J Pharm Sci 124(2):201–207.  https://doi.org/10.1254/jphs.13162FP CrossRefGoogle Scholar
  2. D'Amato RJ, Loughnan MS, Flynn E, Folkman J (1994) Thalidomide is an inhibitor of angiogenesis. Proc Natl Acad Sci U S A 91(9):4082–4085.  https://doi.org/10.1073/pnas.91.9.4082 CrossRefPubMedPubMedCentralGoogle Scholar
  3. Franks ME, Macpherson GR, Figg WD (2004) Thalidomide. Lancet 363(9423):1802–1811.  https://doi.org/10.1016/S0140-6736(04)16308-3 CrossRefPubMedGoogle Scholar
  4. Fratta ID, Sigg EB, Maiorana K (1965) Teratogenic effects of thalidomide in rabbits, rats, hamsters and mice. Toxicol Appl Pharmacol 7(2):268–286.  https://doi.org/10.1016/0041-008X(65)90095-5 CrossRefPubMedGoogle Scholar
  5. Furue MK, Na J, Jackson JP, Okamoto T, Jones M, Baker D, Hata R, Moore HD, Sato JD, Andrews PW (2008) Heparin promotes the growth of human embryonic stem cells in a defined serum-free medium. Proc Natl Acad Sci U S A 105(36):13409–13414.  https://doi.org/10.1073/pnas.0806136105 CrossRefPubMedPubMedCentralGoogle Scholar
  6. Guo CW, Kawakatsu M, Idemitsu M, Urata Y, Goto S, Ono Y, Hamano K, Li TS (2013) Culture under low physiological oxygen conditions improves the stemness and quality of induced pluripotent stem cells. J Cell Physiol 228(11):2159–2166.  https://doi.org/10.1002/jcp.24389 CrossRefPubMedGoogle Scholar
  7. Hayashi Y, Chan T, Warashina M, Fukuda M, Ariizumi T, Okabayashi K, Takayama N, Otsu M, Eto K, Furue MK, Michiue T, Ohnuma K, Nakauchi H, Asashima M (2010) Reduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions. PLoS One 5(11):e14099.  https://doi.org/10.1371/journal.pone.0014099 CrossRefPubMedPubMedCentralGoogle Scholar
  8. Ito T, Ando H, Suzuki T, Ogura T, Hotta K, Imamura Y, Yamaguchi Y, Handa H (2010) Identification of a primary target of thalidomide teratogenicity. Science 327(5971):1345–1350.  https://doi.org/10.1126/science.1177319 CrossRefPubMedGoogle Scholar
  9. Kameoka S, Babiarz J, Kolaja K, Chiao E (2014) A high-throughput screen for teratogens using human pluripotent stem cells. Toxicol Sci 137(1):76–90.  https://doi.org/10.1093/toxsci/kft239 CrossRefPubMedGoogle Scholar
  10. Moore KL, Torchia MG (2015) The developing human, 10th edition Clinically Oriented Embryology. ElsevierGoogle Scholar
  11. Knobloch J, Jungck D, Koch A (2011) Apoptosis induction by thalidomide: critical for limb teratogenicity but therapeutic potential in idiopathic pulmonary fibrosis? Curr Mol Pharmacol 4:26–61CrossRefPubMedGoogle Scholar
  12. Knobloch J, Ruther U (2008) Shedding light on an old mystery: thalidomide suppresses survival pathways to induce limb defects. Cell Cycle 7(9):1121–1127.  https://doi.org/10.4161/cc.7.9.5793 CrossRefPubMedGoogle Scholar
  13. Knobloch J, Shaughnessy JD Jr, Ruther U (2007) Thalidomide induces limb deformities by perturbing the Bmp/Dkk1/Wnt signaling pathway. FASEB J 21(7):1410–1421.  https://doi.org/10.1096/fj.06-7603com CrossRefPubMedGoogle Scholar
  14. Kumagai A, Suga M, Yanagihara K, Itoh Y, Takemori H, Furue MK (2016) A simple method for labeling human embryonic stem cells destined to lose undifferentiated potency. Stem Cells Transl Med 5(3):275–281.  https://doi.org/10.5966/sctm.2015-0145 CrossRefPubMedPubMedCentralGoogle Scholar
  15. Kunisada Y, Tsubooka-Yamazoe N, Shoji M, Hosoya M (2012) Small molecules induce efficient differentiation into insulin-producing cells from human induced pluripotent stem cells. Stem Cell Res 8(2):274–284.  https://doi.org/10.1016/j.scr.2011.10.002 CrossRefPubMedGoogle Scholar
  16. Lenz W (1962) Thalidomide and congenital abnormalities. Lancet 1:45CrossRefGoogle Scholar
  17. McBride WG (1961) Thalidomide and congenital abnormalities. Lancet 2:1358CrossRefGoogle Scholar
  18. Melchert M, List A (2007) The thalidomide saga. Int J Biochem Cell Biol 39(7-8):1489–1499.  https://doi.org/10.1016/j.biocel.2007.01.022 CrossRefPubMedGoogle Scholar
  19. Mellin GW, Katzenstein M (1962a) The saga of thalidomide. Neuropathy to embryopathy, with case reports of congenital anomalies. N Engl J Med 267(23):1184–1192.  https://doi.org/10.1056/NEJM196212062672305 CrossRefPubMedGoogle Scholar
  20. Mellin GW, Katzenstein M (1962b) The saga of thalidomide: neuropathy to embryopathy, with case reports of congenital anomalies. N Engl J Med 267:1238–1244CrossRefPubMedGoogle Scholar
  21. Miller MT, Stromland K (1999) Teratogen update: thalidomide: a review, with a focus on ocular findings and new potential uses. Teratology 60(5):306–321.  https://doi.org/10.1002/(SICI)1096-9926(199911)60:5<306::AID-TERA11>3.0.CO;2-Y CrossRefPubMedGoogle Scholar
  22. Nichols J, Smith A (2009) Naive and primed pluripotent states. Cell Stem Cell 4(6):487–492.  https://doi.org/10.1016/j.stem.2009.05.015 CrossRefPubMedGoogle Scholar
  23. Ninomiya H, Mizuno K, Terada R, Miura T, Ohnuma K, Takahashi S, Asashima M, Michiue T (2015) Improved efficiency of definitive endoderm induction from human induced pluripotent stem cells in feeder and serum-free culture system. In Vitro Cell Dev Biol Anim 51(1):1–8.  https://doi.org/10.1007/s11626-014-9801-y CrossRefPubMedGoogle Scholar
  24. Nowack E (1965) The sensitive phase in thalidomide embryopathy. Humangenetik 1(6):516–536.  https://doi.org/10.1007/BF00338341 CrossRefPubMedGoogle Scholar
  25. Ohgushi M, Matsumura M, Eiraku M, Murakami K, Aramaki T, Nishiyama A, Muguruma K, Nakano T, Suga H, Ueno M, Ishizaki T, Suemori H, Narumiya S, Niwa H, Sasai Y (2010) Molecular pathway and cell state responsible for dissociation-induced apoptosis in human pluripotent stem cells. Cell Stem Cell 7(2):225–239CrossRefPubMedGoogle Scholar
  26. Parman T, Wiley MJ, Wells PG (1999) Free radical-mediated oxidative DNA damage in the mechanism of thalidomide teratogenicity. Nat Med 5(5):582–585.  https://doi.org/10.1038/8466 CrossRefPubMedGoogle Scholar
  27. Ring DB, Johnson KW, Henriksen EJ, Nuss JM, Goff D, Kinnick TR, Ma ST, Reeder JW, Samuels I, Slabiak T, Wagman AS, Hammond ME, Harrison SD (2003) Selective glycogen synthase kinase 3 inhibitors potentiate insulin activation of glucose transport and utilization in vitro and in vivo. Diabetes 52(3):588–595.  https://doi.org/10.2337/diabetes.52.3.588 CrossRefPubMedGoogle Scholar
  28. Schardein JL (2000). Chem induced birth defects, Third Edition 41Google Scholar
  29. Seiler AE, Spielmann H (2011) The validated embryonic stem cell test to predict embryotoxicity in vitro. Nat Protoc 6(7):961–978.  https://doi.org/10.1038/nprot.2011.348 CrossRefPubMedGoogle Scholar
  30. Sheskin J (1965) Thalidomide in the treatment of lepra reactions. Clin Pharmacol Ther 6(3):303–306.  https://doi.org/10.1002/cpt196563303 CrossRefPubMedGoogle Scholar
  31. Singhal S, Mehta J, Desikan R, Ayers D, Roberson P, Eddlemon P, Munshi N, Anaissie E, Wilson C, Dhodapkar M, Zeddis J, Barlogie B (1999) Antitumor activity of thalidomide in refractory multiple myeloma. N Engl J Med 341(21):1565–1571.  https://doi.org/10.1056/NEJM199911183412102 CrossRefPubMedGoogle Scholar
  32. Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K, Yamanaka S (2007) Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 131:861–872CrossRefPubMedGoogle Scholar
  33. Tachikawa S, Nishimura T, Nakauchi H, Ohnuma K (2017) Thalidomide induces apoptosis in undifferentiated human induced pluripotent stem cells. In Vitro Cell Dev Biol Anim 53(9):841–851.  https://doi.org/10.1007/s11626-017-0192-8 CrossRefPubMedGoogle Scholar
  34. Takasato M, Er PX, Chiu HS, Maier B, Baillie GJ, Ferguson C, Parton RG, Wolvetang EJ, Roost MS, Lopes SM, Little MH (2016) Kidney organoids from human iPS cells contain multiple lineages and model human nephrogenesis. Nature 536(7615):238.  https://doi.org/10.1038/nature17982 CrossRefPubMedGoogle Scholar
  35. Thomson JA, Itskovitz-Eldor J, Shapiro SS, Waknitz MA, Swiergiel JJ, Marshall VS, Jones JM (1998) Embryonic stem cell lines derived from human blastocysts. Science 282(5391):1145–1147.  https://doi.org/10.1126/science.282.5391.1145 CrossRefPubMedGoogle Scholar
  36. Watanabe K, Ueno M, Kamiya D, Nishiyama A, Matsumura M, Wataya T, Takahashi J B, Nishikawa S, Nishikawa S, Muguruma K, Sasai Y (2007) A ROCK inhibitor permits survival of dissociated human embryonic stem cells. Nat Biotechnol 25(6):681–686.CrossRefPubMedGoogle Scholar
  37. Yoshida Y, Takahashi K, Okita K, Ichisaka T, Yamanaka S (2009) Hypoxia enhances the generation of induced pluripotent stem cells. Cell Stem Cell 5:237–241CrossRefPubMedGoogle Scholar
  38. Yu J, Vodyanik MA, Smuga-Otto K, Antosiewicz-Bourget J, Frane JL, Tian S, Nie J, Jonsdottir GA, Ruotti V, Stewart R, Slukvin II, Thomson JA (2007) Induced pluripotent stem cell lines derived from human somatic cells. Science 318(5858):1917–1920.  https://doi.org/10.1126/science.1151526 CrossRefPubMedGoogle Scholar

Copyright information

© The Society for In Vitro Biology 2018

Authors and Affiliations

  1. 1.Department of BioengineeringNagaoka University of TechnologyNagaokaJapan
  2. 2.Department of Science of Technology InnovationNagaoka University of TechnologyNagaokaJapan

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