Mitochondria structural reorganization during mouse embryonic stem cell derivation

Abstract

Mouse embryonic stem (ES) cells are widely used in developmental biology and transgenic research. Despite numerous studies, ultrastructural reorganization of inner cell mass (ICM) cells during in vitro culture has not yet been described in detail. Here, we for the first time performed comparative morphological and morphometric analyses of three ES cell lines during their derivation in vitro. We compared morphological characteristics of blastocyst ICM cells at 3.5 and 4.5 days post coitum on feeder cells (day 6, passage 0) with those of ES cells at different passages (day 19, passage 2; day 25, passage 4; and passage 15). At passage 0, there were 23–36% of ES-like cells with various values of the medium cross-sectional area and nucleocytoplasmic parameters, 55% of fibroblast-like (probably trophoblast derivatives), and ~ 19% of dying cells. ES-like cells at passage 0 contained autolysosomes and enlarged mitochondria with reduced numerical density per cell. There were three types of mitochondria that differed in matrix density and cristae width. For the first time, we revealed cells that had two and sometimes three morphologically distinct mitochondria types in the cytoplasm. At passage 2, there were mostly ES cells with a high nucleocytoplasmic ratio and a cytoplasm depleted of organelles. At passage 4, ES cell morphology and morphometric parameters were mostly stable with little heterogeneity. According to our data, cellular structures of ICM cells undergo destabilization during derivation of an ES cell line with subsequent reorganization into the structures typical for ES cells. On the basis of ultrastructural analysis of mitochondria, we believe that the functional activity of these organelles changes during early stages of ES cell formation from the ICM.

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Abbreviations

dpc:

days post coitum

EDTA:

ethylenediaminetetraacetic acid

ER:

endoplasmic reticulum

ESC:

embryonic stem cell

FBS:

fetal bovine serum

ICM:

inner cell mass

KSR:

knockout serum replacement

LIF ESGRO:

leukemia inhibitory factor

NEAA:

non-essential amino acid

TEM:

transmission electron microscopy

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Acknowledgments

We are thankful to Antonina I. Zhelezova and Aleftina N. Golubitsa for assistance with the experiments on mice. The study was supported by the Federal Agency for Scientific Organizations program for support of bioresource collections. We thank the Collective Center of ICG SB RAS “Collection of Pluripotent Human and Mammalian Cell Cultures for Biological and Biomedical Research” (http://ckp.icgen.ru/cells/) for providing the cell lines, and we are grateful to the Interinstitutional Shared Center for Microscopic Analysis of Biological Objects (ICG SB RAS, Novosibirsk) for providing the microscopy equipment for this study. The English language was corrected and certified by shevchuk-editing.com.

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Correspondence to Ksenia N. Morozova.

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All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.

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Suldina, L.A., Morozova, K.N., Menzorov, A.G. et al. Mitochondria structural reorganization during mouse embryonic stem cell derivation. Protoplasma 255, 1373–1386 (2018). https://doi.org/10.1007/s00709-018-1236-y

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Keywords

  • Embryonic stem cell
  • Mitochondria
  • Electron microscopy
  • Cell ultrastructure