Thalidomide induces apoptosis in undifferentiated human induced pluripotent stem cells
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Thalidomide, which was formerly available commercially to control the symptoms of morning sickness, is a strong teratogen that causes fetal abnormalities. However, the mechanism of thalidomide teratogenicity is not fully understood; thalidomide toxicity is not apparent in rodents, and the use of human embryos is ethically and technically untenable. In this study, we designed an experimental system featuring human-induced pluripotent stem cells (hiPSCs) to investigate the effects of thalidomide. These cells exhibit the same characteristics as those of epiblasts originating from implanted fertilized ova, which give rise to the fetus. Therefore, theoretically, thalidomide exposure during hiPSC differentiation is equivalent to that in the human fetus. We examined the effects of thalidomide on undifferentiated hiPSCs and early-differentiated hiPSCs cultured in media containing bone morphogenetic protein-4, which correspond, respectively, to epiblast (future fetus) and trophoblast (future extra-embryonic tissue). We found that only the number of undifferentiated cells was reduced. In undifferentiated cells, application of thalidomide increased the number of apoptotic and dead cells at day 2 but not day 4. Application of thalidomide did not affect the cell cycle. Furthermore, immunostaining and flow cytometric analysis revealed that thalidomide exposure had no effect on the expression of specific markers of undifferentiated and early trophectodermal differentiated cells. These results suggest that the effect of thalidomide was successfully detected in our experimental system and that thalidomide eliminated a subpopulation of undifferentiated hiPSCs. This study may help to elucidate the mechanisms underlying thalidomide teratogenicity and reveal potential strategies for safely prescribing this drug to pregnant women.
KeywordsThalidomide Human induced pluripotent cells Teratogenicity
This work was supported in part by grants from the Ministry of Health, Labour, 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, decision to publish, or preparation of the manuscript.
S.T. and K.O. designed the project. S.T. performed all experiments. T.N. and H.N. provided a hiPS clone and helpful feedback on the manuscript. S.T. and K.O. wrote the manuscript and all authors reviewed the manuscript.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no competing financial interests.
- Ando M, Nishimura T, Yamazaki S, Yamaguchi T, Kawana-Tachikawa A, Hayama T, Nakauchi Y, Ando J, Ota Y, Takahashi S, Nishimura K, Ohtaka M, Nakanishi M, Miles JJ, Burrows SR, Brenner MK, Nakauchi H (2015) A safeguard system for induced pluripotent stem cell-derived rejuvenated T cell therapy. Stem Cell Rep 5:597–608CrossRefGoogle Scholar
- Chen G, Gulbranson DR, Hou Z, Bolin JM, Ruotti V, Probasco MD, Smuga-Otto K, Howden SE, Diol NR, Propson NE, Wagner R, Lee GO, Antosiewicz-Bourget J, Teng JM, Thomson JA (2011) Chemically defined conditions for human iPSC derivation and culture. Nat Methods 8:424–429CrossRefPubMedPubMedCentralGoogle Scholar
- 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:e14099CrossRefPubMedPubMedCentralGoogle Scholar
- Knobloch J, Shaughnessy JD, Ruther U (2007) Thalidomide induces limb deformities by perturbing the Bmp/Dkk1/Wnt signaling pathway. FASEB J 21(7):1410–1421Google Scholar
- Meganathan K, Jagtap S, Wagh V, Winkler J, Gaspar JA, Hildebrand D, Trusch M, Lehmann K, Hescheler J, Schluter H, Sachinidis A (2012) Identification of thalidomide-specific transcriptomics and proteomics signatures during differentiation of human embryonic stem cells. PLoS One 7:e44228CrossRefPubMedPubMedCentralGoogle Scholar
- Moore KL, Persaud TVN, Torchia MG (2015) The developing human Clinically oriented embryology, 10th edition. Elsevier Health Sciences, Netherlands AmsterdamGoogle Scholar
- Nishimura T, Kaneko S, Kawana-Tachikawa A, Tajima Y, Goto H, Zhu D, Nakayama-Hosoya K, Iriguchi S, Uemura Y, Shimizu T, Takayama N, Yamada D, Nishimura K, Ohtaka M, Watanabe N, Takahashi S, Iwamoto A, Koseki H, Nakanishi M, Eto K, Nakauchi H (2013) Generation of rejuvenated antigen-specific T cells by reprogramming to pluripotency and redifferentiation. Cell Stem Cell 12:114–126CrossRefPubMedGoogle Scholar
- 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 7:225–239CrossRefPubMedGoogle Scholar
- Parman T, Wiley MJ, Wells PG (1999) Free radical-mediated oxidative DNA damage in the mechanism of thalidomide teratogenicity. Nat Med 5:582–585Google Scholar
- Qin XY, Akanuma H, Wei F, Nagano R, Zeng Q, Imanishi S, Ohsako S, Yoshinaga J, Yonemoto J, Tanokura M, Sone H (2012) Effect of low-dose thalidomide on dopaminergic neuronal differentiation of human neural progenitor cells: a combined study of metabolomics and morphological analysis. Neurotoxicology 33:1375–1380CrossRefPubMedGoogle Scholar
- Schardein JL (2000) Chemically induced birth defects, third edition. Informa Healthcare, New York LondonGoogle Scholar
- Vukicevic S, Kleinman HK, Luyten FP, Roberts AB, Roche NS, Reddi AH (1992) Identification of multiple active growth factors in basement membrane Matrigel suggests caution in interpretation of cellular activity related to extracellular matrix components. Exp Cell Res 202:1–8CrossRefPubMedGoogle Scholar