Abstract
Although the feeder-free culture system has been established, the microenvironment provided by the feeder cells still possesses a unique advantage in maintaining the long-term stability and the rapid proliferation of pluripotent stem cells (PSCs). The aim of this study is to discover the adaptive ability of PSCs upon changes of feeder layers. In this study, the morphology, pluripotent marker expression, differentiation ability of bovine embryonic stem cells (bESCs) cultured on low-density, or methanol fixed mouse embryonic fibroblasts were examined by immunofluorescent staining, Western blotting, real-time reverse transcription polymerase chain reaction, and RNA-seq. The results showed that the changes of feeder layers did not induce the rapid differentiation of bESCs, while resulting in the differentiation initiation and alteration of pluripotent state of bESCs. More importantly, the expression of endogenous growth factors and extracellular matrix were increased, and the expression of cell adhesion molecules was altered, which indicated that bESCs may compensate some functions of the feeder layers upon its changes. This study shows the PSCs have the self-adaptive ability responded to the feeder layer alteration.
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Data availability
All data generated or analyzed during this study are included in this published article and its Supplementary Information files. Gene expression data of MC-F7 and 1/7MC-F7 or MC-F7 and MT-F7 are available in the GEO databases under the accession number GSE213631 or GSE199731.
References
Bendall SC, Stewart MH, Menendez P, George D, Vijayaragavan K, Werbowetski-Ogilvie T, Ramos-Mejia V, Rouleau A, Yang J, Bosse M, Lajoie G, Bhatia M (2007) IGF and FGF cooperatively establish the regulatory stem cell niche of pluripotent human cells in vitro. Nature 448:1015–1021. https://doi.org/10.1038/nature06027
Bogliotti YS, Wu J, Vilarino M, Okamura D, Soto DA, Zhong C, Sakurai M, Sampaio RV, Suzuki K, Izpisua Belmonte JC, Ross PJ (2018) Efficient derivation of stable primed pluripotent embryonic stem cells from bovine blastocysts. Proc Natl Acad Sci U S A 115:2090–2095. https://doi.org/10.1073/pnas.1716161115
Campos LS (2005) Beta1 integrins and neural stem cells: making sense of the extracellular environment. BioEssays 27:698–707. https://doi.org/10.1002/bies.20256
Chang BS, Choi YJ, Kim JH (2015) Collagen complexes increase the efficiency of iPS cells generated using fibroblasts from adult mice. J Reprod Dev 61:145–153. https://doi.org/10.1262/jrd.2014-081
Chen LH, Sung TC, Lee HH, Higuchi A, Su HC, Lin KJ, Huang YR, Ling QD, Kumar SS, Alarfaj AA, Munusamy MA, Nasu M, Chen DC, Hsu ST, Chang Y, Lee KF, Wang HC, Umezawa A (2019) Xeno-free and feeder-free culture and differentiation of human embryonic stem cells on recombinant vitronectin-grafted hydrogels. Biomater Sci 7:4345–4362. https://doi.org/10.1039/c9bm00418a
Chen SS, Fitzgerald W, Zimmerberg J, Kleinman HK, Margolis L (2007) Cell-cell and cell-extracellular matrix interactions regulate embryonic stem cell differentiation. Stem Cells 25:553–561. https://doi.org/10.1634/stemcells.2006-0419
Choi JS, Yang HJ, Kim BS, Kim JD, Kim JY, Yoo B, Park K, Lee HY, Cho YW (2009) Human extracellular matrix (ECM) powders for injectable cell delivery and adipose tissue engineering. J Control Release 139:2–7. https://doi.org/10.1016/j.jconrel.2009.05.034
D’Amour KA, Agulnick AD, Eliazer S, Kelly OG, Kroon E, Baetge EE (2005) Efficient differentiation of human embryonic stem cells to definitive endoderm. Nat Biotechnol 23:1534–1541. https://doi.org/10.1038/nbt1163
Desai N, Rambhia P, Gishto A (2015) Human embryonic stem cell cultivation: historical perspective and evolution of xeno-free culture systems. Reprod Biol Endocrinol 13:9. https://doi.org/10.1186/s12958-015-0005-4
Eiselleova L, Peterkova I, Neradil J, Slaninova I, Hampl A, Dvorak P (2008) Comparative study of mouse and human feeder cells for human embryonic stem cells. Int J Dev Biol 52:353–363. https://doi.org/10.1387/ijdb.082590le
Ellis SJ, Tanentzapf G (2010) Integrin-mediated adhesion and stem-cell-niche interactions. Cell Tissue Res 339:121–130. https://doi.org/10.1007/s00441-009-0828-4
Evans MJ, Kaufman MH (1981) Establishment in culture of pluripotential cells from mouse embryos. Nature 292:154–156. https://doi.org/10.1038/292154a0
Gerecht S, Bettinger CJ, Zhang Z, Borenstein JT, Vunjak-Novakovic G, Langer R (2007) The effect of actin disrupting agents on contact guidance of human embryonic stem cells. Biomaterials 28:4068–4077. https://doi.org/10.1016/j.biomaterials.2007.05.027
Giancotti FG, Ruoslahti E (1999) Integrin signaling. Science 285:1028–1032. https://doi.org/10.1126/science.285.5430.1028
Guo R, Ye X, Yang J, Zhou Z, Tian C, Wang H, Wang H, Fu H, Liu C, Zeng M, Yang J, Liu L (2018) Feeders facilitate telomere maintenance and chromosomal stability of embryonic stem cells. Nat Commun 9:2620. https://doi.org/10.1038/s41467-018-05038-2
Han X, Xiang J, Li C, Wang J, Wang C, Zhang Y, Li Z, Lu Z, Yue Y, Li X (2020) MLL1 combined with GSK3 and MAP2K inhibition improves the development of in vitro-fertilized embryos. Theriogenology 146:58–70. https://doi.org/10.1016/j.theriogenology.2020.01.051
Hayal TB, Doğan A (2021) Feeder-free human embryonic stem cell culture under defined culture conditions. Methods Mol Biol. https://doi.org/10.1007/7651_2021_404
Hayashi Y, Caboni L, Das D, Yumoto F, Clayton T, Deller MC, Nguyen P, Farr CL, Chiu HJ, Miller MD, Elsliger MA, Deacon AM, Godzik A, Lesley SA, Tomoda K, Conklin BR, Wilson IA, Yamanaka S, Fletterick RJ (2015) Structure-based discovery of NANOG variant with enhanced properties to promote self-renewal and reprogramming of pluripotent stem cells. Proc Natl Acad Sci U S A 112:4666–4671. https://doi.org/10.1073/pnas.1502855112
Hughes C, Radan L, Chang WY, Stanford WL, Betts DH, Postovit LM, Lajoie GA (2012) Mass spectrometry-based proteomic analysis of the matrix microenvironment in pluripotent stem cell culture. Mol Cell Proteomics 11:1924–1936. https://doi.org/10.1074/mcp.M112.020057
Hunt GC, Singh P, Schwarzbauer JE (2012) Endogenous production of fibronectin is required for self-renewal of cultured mouse embryonic stem cells. Exp Cell Res 318:1820–1831. https://doi.org/10.1016/j.yexcr.2012.06.009
Hynes RO (2009) The extracellular matrix: not just pretty fibrils. Science 326:1216–1219. https://doi.org/10.1126/science.1176009
Hyslop L, Stojkovic M, Armstrong L, Walter T, Stojkovic P, Przyborski S, Herbert M, Murdoch A, Strachan T, Lako M (2005) Downregulation of NANOG induces differentiation of human embryonic stem cells to extraembryonic lineages. Stem Cells 23:1035–1043. https://doi.org/10.1634/stemcells.2005-0080
Joddar B, Nishioka C, Takahashi E, Ito Y (2015) Chemically fixed autologous feeder cell-derived niche for human induced pluripotent stem cell culture. J Mater Chem B 3:2301–2307. https://doi.org/10.1039/c4tb01635a
Kimura Y, Kasai K, Miyata S (2018) Feeder-free culture for mouse induced pluripotent stem cells by using UV/ozone surface-modified substrates. Mater Sci Eng C Mater Biol Appl 92:280–286. https://doi.org/10.1016/j.msec.2018.06.053
Lee JH, Lee EJ, Lee CH, Park JH, Han JY, Lim JM (2009) Requirement of leukemia inhibitory factor for establishing and maintaining embryonic stem cells in mice. Fertil Steril 92:1133–1140. https://doi.org/10.1016/j.fertnstert.2008.07.1733
Lee ST, Yun JI, Jo YS, Mochizuki M, van der Vlies AJ, Kontos S, Ihm JE, Lim JM, Hubbell JA (2010) Engineering integrin signaling for promoting embryonic stem cell self-renewal in a precisely defined niche. Biomaterials 31:1219–1226. https://doi.org/10.1016/j.biomaterials.2009.10.054
Luchsinger C, Arias ME, Vargas T, Paredes M, Sanchez R, Felmer R (2014) Stability of reference genes for normalization of reverse transcription quantitative real-time PCR (RT-qPCR) data in bovine blastocysts produced by IVF, ICSI and SCNT. Zygote 22:505–512. https://doi.org/10.1017/S0967199413000099
Nagaoka M, Si-Tayeb K, Akaike T, Duncan SA (2010) Culture of human pluripotent stem cells using completely defined conditions on a recombinant E-cadherin substratum. BMC Dev Biol 10:60. https://doi.org/10.1186/1471-213X-10-60
Nakagawa M, Taniguchi Y, Senda S, Takizawa N, Ichisaka T, Asano K, Morizane A, Doi D, Takahashi J, Nishizawa M, Yoshida Y, Toyoda T, Osafune K, Sekiguchi K, Yamanaka S (2014) A novel efficient feeder-free culture system for the derivation of human induced pluripotent stem cells. Sci Rep 4:3594. https://doi.org/10.1038/srep03594
Nakashima Y, Omasa T (2016) What kind of signaling maintains pluripotency and viability in human-induced pluripotent stem cells cultured on Laminin-511 with serum-free medium? Biores Open Access 5:84–93. https://doi.org/10.1089/biores.2016.0001
Ngangan AV, McDevitt TC (2009) Acellularization of embryoid bodies via physical disruption methods. Biomaterials 30:1143–1149. https://doi.org/10.1016/j.biomaterials.2008.11.001
Niessen CM, Leckband D, Yap AS (2011) Tissue organization by cadherin adhesion molecules: dynamic molecular and cellular mechanisms of morphogenetic regulation. Physiol Rev 91:691–731. https://doi.org/10.1152/physrev.00004.2010
Nikkhah M, Edalat F, Manoucheri S, Khademhosseini A (2012) Engineering microscale topographies to control the cell-substrate interface. Biomaterials 33:5230–5246. https://doi.org/10.1016/j.biomaterials.2012.03.079
Oh JH, Do HJ, Yang HM, Moon SY, Cha KY, Chung HM, Kim JH (2005) Identification of a putative transactivation domain in human Nanog. Exp Mol Med 37:250–254. https://doi.org/10.1038/emm.2005.33
Przybyla L, Voldman J (2012) Probing embryonic stem cell autocrine and paracrine signaling using microfluidics. Annu Rev Anal Chem (Palo Alto Calif) 5:293–315. https://doi.org/10.1146/annurev-anchem-062011-143122
Rahman S, Patel Y, Murray J, Patel KV, Sumathipala R, Sobel M, Wijelath ES (2005) Novel hepatocyte growth factor (HGF) binding domains on fibronectin and vitronectin coordinate a distinct and amplified Met-integrin induced signalling pathway in endothelial cells. BMC Cell Biol 6:8. https://doi.org/10.1186/1471-2121-6-8
Raymond K, Deugnier MA, Faraldo MM, Glukhova MA (2009) Adhesion within the stem cell niches. Curr Opin Cell Biol 21:623–629. https://doi.org/10.1016/j.ceb.2009.05.004
Ren Y, Ma Z, Yu T, Ling M, Wang H (2018) Methanol fixed fibroblasts serve as feeder cells to maintain stem cells in the pluripotent state in vitro. Sci Rep 8:7780. https://doi.org/10.1038/s41598-018-26238-2
Saha S, Ji L, de Pablo JJ, Palecek SP (2008) TGFbeta/Activin/Nodal pathway in inhibition of human embryonic stem cell differentiation by mechanical strain. Biophys J 94:4123–4133. https://doi.org/10.1529/biophysj.107.119891
Soto DA, Navarro M, Zheng C, Halstead MM, Zhou C, Guiltinan C, Wu J, Ross PJ (2021) Simplification of culture conditions and feeder-free expansion of bovine embryonic stem cells. Sci Rep 11:11045. https://doi.org/10.1038/s41598-021-90422-0
Stingl J, Eirew P, Ricketson I, Shackleton M, Vaillant F, Choi D, Li HI, Eaves CJ (2006) Purification and unique properties of mammary epithelial stem cells. Nature 439:993–997. https://doi.org/10.1038/nature04496
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 131:861–872. https://doi.org/10.1016/j.cell.2007.11.019
Takahashi K, Yamanaka S (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126:663–676. https://doi.org/10.1016/j.cell.2006.07.024
Talbot NC, Sparks WO, Powell AM, Kahl S, Caperna TJ (2012) Quantitative and semiquantitative immunoassay of growth factors and cytokines in the conditioned medium of STO and CF-1 mouse feeder cells. In Vitro Cell Dev Biol Anim 48:1–11. https://doi.org/10.1007/s11626-011-9467-7
Turner AE, Flynn LE (2012) Design and characterization of tissue-specific extracellular matrix-derived microcarriers. Tissue Eng Part C Methods 18:186–197. https://doi.org/10.1089/ten.TEC.2011.0246
Villa-Diaz LG, Ross AM, Lahann J, Krebsbach PH (2013) Concise review: The evolution of human pluripotent stem cell culture: from feeder cells to synthetic coatings. Stem Cells 31:1–7. https://doi.org/10.1002/stem.1260
Wang L, Zhang R, Ma R, Jia G, Jian S, Zeng X, Xiong Z, Li B, Li C, Lv Z, Bai X (2020) Establishment of a feeder and serum-free culture system for human embryonic stem cells. Zygote 28:175–182. https://doi.org/10.1017/s0967199419000625
Wu J, Okamura D, Li M, Suzuki K, Luo C, Ma L, He Y, Li Z, Benner C, Tamura I, Krause MN, Nery JR, Du T, Zhang Z, Hishida T, Takahashi Y, Aizawa E, Kim NY, Lajara J, Guillen P, Campistol JM, Esteban CR, Ross PJ, Saghatelian A, Ren B, Ecker JR, Izpisua Belmonte JC (2015) An alternative pluripotent state confers interspecies chimaeric competency. Nature 521:316–321. https://doi.org/10.1038/nature14413
Xu C, Inokuma MS, Denham J, Golds K, Kundu P, Gold JD, Carpenter MK (2001) Feeder-free growth of undifferentiated human embryonic stem cells. Nat Biotechnol 19:971–974. https://doi.org/10.1038/nbt1001-971
Xu W, Hao R, Wang J, Gao L, Han X, Li C, Fang S, Zhang H, Li X (2022) Methanol fixed feeder layers altered the pluripotency and metabolism of bovine pluripotent stem cells. Sci Rep 12:9177. https://doi.org/10.1038/s41598-022-13249-3
Yang Y, Liu B, Xu J, Wang J, Wu J, Shi C, Xu Y, Dong J, Wang C, Lai W, Zhu J, Xiong L, Zhu D, Li X, Yang W, Yamauchi T, Sugawara A, Li Z, Sun F, Li X, Li C, He A, Du Y, Wang T, Zhao C, Li H, Chi X, Zhang H, Liu Y, Li C, Duo S, Yin M, Shen H, Belmonte JCI, Deng H (2017) Derivation of pluripotent stem cells with in vivo embryonic and extraembryonic potency. Cell 169:243-257 e25. https://doi.org/10.1016/j.cell.2017.02.005
Yoshida Y, Takahashi K, Okita K, Ichisaka T, Yamanaka S (2009) Hypoxia enhances the generation of induced pluripotent stem cells. Cell Stem Cell 5:237–241. https://doi.org/10.1016/j.stem.2009.08.001
Yue XS, Fujishiro M, Nishioka C, Arai T, Takahashi E, Gong JS, Akaike T, Ito Y (2012) Feeder cells support the culture of induced pluripotent stem cells even after chemical fixation. PLoS ONE 7:e32707. https://doi.org/10.1371/journal.pone.0032707
Zheng R, Geng T, Wu DY, Zhang T, He HN, Du HN, Zhang D, Miao YL, Jiang W (2021a) Derivation of feeder-free human extended pluripotent stem cells. Stem Cell Rep 16:1686–1696. https://doi.org/10.1016/j.stemcr.2021.06.001
Zheng R, Geng T, Wu DY, Zhang T, He HN, Du HN, Zhang D, Miao YL, Jiang W (2021b) Derivation of feeder-free human extended pluripotent stem cells. Stem Cell Rep 16:2410–2414. https://doi.org/10.1016/j.stemcr.2021.07.019
Funding
This work was financially supported by the Science and Technology Major Project of the Inner Mongolia Autonomous Region of China (2020ZD0007), the National Natural Science Foundation of China (32160172), the Science Research Foundation of Baotou Medical College (BYJJ-ZRQM202215), the Major Program of the Inner Mongolia Natural Science Foundation (2020ZD10), the National Transgenic Project (2016ZX0801000-002 and 2016ZX08010005-001), and the Science and Technology Major Project of the Inner Mongolia Autonomous Region of China to the State Key Laboratory of Reproductive Regulation and Breeding of Grassland Livestock (zdzx2018065 and 2021ZD0048).
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Xueling Li and Yongli Yue designed the project and reviewed this paper. Wenqiang Xu, Lingna Gao, and Wei Li conducted the experiments; Wenqiang Xu analyzed the results and wrote the manuscript. The bioinformatics analyses of RNA-seq were conducted by Jing Wang.
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Xu, W., Gao, L., Li, W. et al. The adaptation of bovine embryonic stem cells to the changes of feeder layers. In Vitro Cell.Dev.Biol.-Animal 59, 85–99 (2023). https://doi.org/10.1007/s11626-022-00731-5
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DOI: https://doi.org/10.1007/s11626-022-00731-5