Cytotechnology

, Volume 70, Issue 1, pp 129–140 | Cite as

Liver epithelioid progenitor cells derived from fetal Luxi bovine alleviate liver fibrosis

Article

Abstract

Liver epithelioid progenitor cells (LEPCs) have important roles in liver therapy because of their hepatic differentiation potency in vitro and in vivo. Despite many researches on humans, mice, and rats, equivalent progenitor cells derived from bovine are relatively rare. The purpose of our current study is to characterize bovine LEPCs, and research on the cure potency of this heteroplastic progenitor cells on mice liver fibrosis. We have used collagenase IV digesting and differential adhesion method to isolate slabstone shape, EpCAM, LGR5, NCAM1 and SOX9 positive progenitor cells from fetal Luxi bovine liver. When cultured in hepatic differentiation media containing 20 ng/ml Oncostatin M, LEPCs can differentiate into hepatocytes in vitro. After 4 weeks of intravenous tail vein injection into CCl4-injured mouse liver, LEPCs engrafted into liver parenchyma, differentiated into ALB positive hepatocytes, and could alleviate liver fibrosis through down regulating fibrosis genes-Tgfb1 and α-SMA as well as decreasing expression of collagen gene Col1a1, Col3a1, and Col4a1, and regain liver function by recovering ALT and AST. Our findings provided a useful tool for studying liver development in vitro, new cell resource for heterograft on mouse liver diseases, and a new platform for researches on immune rejection of heterogeneous cell transplantation.

Keywords

Hepatocyte differentiation Liver epithelioid progenitor cells Stem cells therapy Liver fibrosis Heterogeneous cell transplantation 

Abbreviations

AFP

Alpha fetoprotein

CDM

Cholangiocytic differentiation medium

EPCAM

Epithelial cell adhesion molecule

G6PC

Glucose-6-phosphatase catalytic subunit

HDM

Hepatic differentiation medium

HepSCs

Hepatic stem cells

iHepSCs

Induced hepatic stem cells

Itgb4

Integrin subunit beta 4

LGR5

Leucine rich repeat containing G protein coupled receptor 5

LPCs

Liver progenitor cells

LEPCs

Liver epithelioid progenitor cells

NCAM1

Neural cell adhesion molecule 1

OSM

Oncostatin M

PAS

Periodic acid-Schiff

PFA

Paraformaldehyde

SOX9

SRY (sex determining region Y)-box 9

Notes

Acknowledgements

This research was supported by the National Natural Science Foundation of China (Grant Nos: 31472099; 31672404), the Agricultural Science and Technology Innovation Program (ASTIP) (cxgc-ias-01) and the project National Infrastructure of Animal Germplasm Resources (2016).

Author’s contribution

KW and HL participated in experimental designs, data acquisition and analysis, and data interpretation, as well as drafting of the manuscript. JY, CM and ZZ were involved in data acquisition and data interpretation, as well as drafting the manuscript. DZ and WG were also involved in data acquisition and data interpretation, as well as drafting the manuscript. All authors read and approved the final manuscript.

Compliance with ethical standards

Conflict of interest

The authors declare that there is no conflict of interests.

Supplementary material

10616_2017_113_MOESM1_ESM.docx (15 kb)
Supplementary material 1 (DOCX 14 kb)
10616_2017_113_MOESM2_ESM.docx (14 kb)
Supplementary material 2 (DOCX 13 kb)
10616_2017_113_MOESM3_ESM.docx (13 kb)
Supplementary material 3 (DOCX 13 kb)

References

  1. Arends B, Spee B, Schotanus BA, Roskams T, van den Ingh TS, Penning LC, Rothuizen J (2009) In vitro differentiation of liver progenitor cells derived from healthy dog livers. Stem Cells and Dev 18:351–358. doi: 10.1089/scd.2008.0043 CrossRefGoogle Scholar
  2. Barker N et al (2007) Identification of stem cells in small intestine and colon by marker gene Lgr5. Nature 449:1003–1007. doi: 10.1038/nature06196 CrossRefGoogle Scholar
  3. Barker N, van Es JH, Jaks V, Kasper M, Snippert H, Toftgard R, Clevers H (2008) Very long-term self-renewal of small intestine, colon, and hair follicles from cycling Lgr5+ve stem cells. Cold Spring Harb Symp Quant Biol 73:351–356. doi: 10.1101/sqb.2008.72.003 CrossRefGoogle Scholar
  4. Barker N et al (2010) Lgr5(+ve) stem cells drive self-renewal in the stomach and build long-lived gastric units in vitro. Cell Stem Cell 6:25–36. doi: 10.1016/j.stem.2009.11.013 CrossRefGoogle Scholar
  5. Buzhor E et al (2013) Reactivation of NCAM1 defines a subpopulation of human adult kidney epithelial cells with clonogenic and stem/progenitor properties. Am J Pathol 183:1621–1633. doi: 10.1016/j.ajpath.2013.07.034 CrossRefGoogle Scholar
  6. Gao YH, Guan WJ, Ma YH (2015) A short review: research progress of bovine stem cells. Cell Mol Biol (Noisy-le-grand) 61:74–78Google Scholar
  7. Huch M, Boj SF, Clevers H (2013a) Lgr5(+) liver stem cells, hepatic organoids and regenerative medicine. Regen Med 8:385–387. doi: 10.2217/rme.13.39 CrossRefGoogle Scholar
  8. Huch M et al (2013b) In vitro expansion of single Lgr5+ liver stem cells induced by Wnt-driven regeneration. Nature 494:247–250. doi: 10.1038/nature11826 CrossRefGoogle Scholar
  9. Huch M et al (2015) Long-term culture of genome-stable bipotent stem cells from adult human liver. Cell 160:299–312. doi: 10.1016/j.cell.2014.11.050 CrossRefGoogle Scholar
  10. Kido T et al (2015) CPM is a useful cell surface marker to isolate expandable bi-potential liver progenitor cells derived from human iPS cells. Stem Cell Rep 5:508–515. doi: 10.1016/j.stemcr.2015.08.008 CrossRefGoogle Scholar
  11. Lee PH et al (2016) Antifibrotic activity of human placental amnion membrane-derived CD34+ mesenchymal stem/progenitor cell transplantation in mice with thioacetamide-induced liver injury. Stem Cells Transl Med 5:1473–1484. doi: 10.5966/sctm.2015-0343 CrossRefGoogle Scholar
  12. Li WL et al (2006) Isolation and characterization of bipotent liver progenitor cells from adult mouse. Stem Cells 24:322–332. doi: 10.1634/stemcells.2005-0108 CrossRefGoogle Scholar
  13. Lin JS et al (2015) Hepatic differentiation of human amniotic epithelial cells and in vivo therapeutic effect on animal model of cirrhosis. J Gastroenterol Hepatol 30:1673–1682. doi: 10.1111/jgh.12991 CrossRefGoogle Scholar
  14. Liu C et al (2016) Sox9 regulates self-renewal and tumorigenicity by promoting symmetrical cell division of cancer stem cells in hepatocellular carcinoma. Hepatology 64:117–129. doi: 10.1002/hep.28509 CrossRefGoogle Scholar
  15. Lu T, Hu P, Su X, Li C, Ma Y, Guan W (2014) Isolation and characterization of mesenchymal stem cells derived from fetal bovine liver. Cell Tissue Bank 15:439–450. doi: 10.1007/s10561-013-9410-0 CrossRefGoogle Scholar
  16. Okabe M et al (2009) Potential hepatic stem cells reside in EpCAM+ cells of normal and injured mouse liver. Development 136:1951–1960. doi: 10.1242/dev.031369 CrossRefGoogle Scholar
  17. Okaya A et al (2005) Oncostatin M inhibits proliferation of rat oval cells, OC15-5, inducing differentiation into hepatocytes. Am J Pathol 166:709–719. doi: 10.1016/S0002-9440(10)62292-4 CrossRefGoogle Scholar
  18. Paganelli M et al (2014) Downregulation of Sox9 expression associates with hepatogenic differentiation of human liver mesenchymal stem/progenitor cells. Stem cells Dev 23:1377–1391. doi: 10.1089/scd.2013.0169 CrossRefGoogle Scholar
  19. Park M et al (2015) Tonsil-derived mesenchymal stem cells ameliorate CCl4-induced liver fibrosis in mice via autophagy activation. Sci Rep 5:8616. doi: 10.1038/srep08616 CrossRefGoogle Scholar
  20. Schmelzer E et al (2007) Human hepatic stem cells from fetal and postnatal donors. J Exp Med 204:1973–1987. doi: 10.1084/jem.20061603 CrossRefGoogle Scholar
  21. Terris B, Cavard C, Perret C (2010) EpCAM, a new marker for cancer stem cells in hepatocellular carcinoma. J Hepatol 52:280–281. doi: 10.1016/j.jhep.2009.10.026 CrossRefGoogle Scholar
  22. Wang Y, Yu X, Chen E, Li L (2016) Liver-derived human mesenchymal stem cells: a novel therapeutic source for liver diseases. Stem Cell Research Ther 7:71. doi: 10.1186/s13287-016-0330-3 CrossRefGoogle Scholar
  23. Wauthier E et al (2008) Hepatic stem cells and hepatoblasts: identification, isolation, and ex vivo maintenance. Methods Cell Biol 86:137–225. doi: 10.1016/S0091-679X(08)00008-3 CrossRefGoogle Scholar
  24. Yamaza T et al (2015) In vivo hepatogenic capacity and therapeutic potential of stem cells from human exfoliated deciduous teeth in liver fibrosis in mice. Stem Cell Res Ther 6:171. doi: 10.1186/s13287-015-0154-6 CrossRefGoogle Scholar
  25. Yoon SM, Gerasimidou D, Kuwahara R, Hytiroglou P, Yoo JE, Park YN, Theise ND (2011) Epithelial cell adhesion molecule (EpCAM) marks hepatocytes newly derived from stem/progenitor cells in humans. Hepatology 53:964–973. doi: 10.1002/hep.24122 CrossRefGoogle Scholar
  26. Yovchev MI, Xue Y, Shafritz DA, Locker J, Oertel M (2014) Repopulation of the fibrotic/cirrhotic rat liver by transplanted hepatic stem/progenitor cells and mature hepatocytes. Hepatology 59:284–295. doi: 10.1002/hep.26615 CrossRefGoogle Scholar
  27. Yu B et al (2013) Reprogramming fibroblasts into bipotential hepatic stem cells by defined factors. Cell Stem Cell 13:328–340. doi: 10.1016/j.stem.2013.06.017 CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2017

Authors and Affiliations

  1. 1.College of Wildlife ResourcesNortheast Forestry UniversityHarbinChina
  2. 2.Institute of Animal ScienceChinese Academy of Agricultural SciencesBeijingChina
  3. 3.College of Life SciencesQufu Normal UniversityQufuChina

Personalised recommendations