Mariana D. Dabeva, M.D., Ph.D., D.Sc.
Research on hepatic stem cells has entered a new era of controversy, excitement, and great expectations. Work in our laboratory focuses on the tissue specific liver stem/progenitor cells.
Rat liver stem cells are specified early during embryogenesis (9-9.5 dpc) from the foregut endoderm. The genetic program for their differentiation is executed through several steps with the help of signaling factors, signal transducers, and transcription factors, some of which are critical in the pathway of liver progenitor cell formation and progression along hepatocytic and bile duct epithelial cell lineages. Our current hypothesis is that the fetal liver contains a subset of hepatic stem/progenitor cells that have not reached terminal differentiation and are arrested in the adult quiescent liver as dormant/slow cycling, unidentifiable stem cells or partially differentiated progenitor cells. Under certain conditions, when the liver parenchyma is extensively damaged, liver stem/progenitor cells can be activated by yet unknown mechanisms to proliferate and differentiate into mature hepatocytes and biliary epithelial cells and repopulate the liver.
Because of the tremendous biological and pathobiological significance and applicability of liver stem/progenitor cells, our research program aims at: (i) their identification and isolation from fetal and adult liver, (ii) their propagation and maintenance in culture, (iii) elucidation of molecular mechanisms leading to their activation, proliferation and bipotential differentiation into hepatocytes or cholangiocytes, and (iv) identification of the genes involved in these processes. To achieve these goals, we are applying modern methods of gene analysis and are studying novel gene expression patterns in rat fetal liver and in models of stem/progenitor cells activation: D-galactosamine injured liver, retrorsine treated liver in conjunction with partial hepatectomy and 2-acetyl-amino fluorine treated rats also subjected to partial hepatectomy.
Using the suppression-subtractive hybridization (SSH) approach, we have constructed a cDNA library from 13-day fetal liver, subtracted from normal adult liver andidentified fetal liver epithelial cell specific clones. These clones are currently being characterized and experiments are being conducted to determine the function of these genes in fetal liver. One of the clones is the proprotein convertase subtilisin/kexin 9 (Pcsk9). Recently, it was shown that expression of Pcsk-9 is down-regulated in the liver of animals fed a cholesterol rich diet. Mutations in this gene appear to be the cause of a novel form of familial hypercholesterolemia. How this convertase relates to cholesterol homeostasis is an enigma. We hypothesize that Pcsk-9 has important functions in fetal liver, some related but others not related to cholesterol homeostasis, and we are currently studying the function of this convertase in hepatic cells and liver.
Applying cDNA microarray technology, we have studied the gene expression profile of fetal liver epithelial cells at ED 13, 14, 15, 16, 18, with newborn liver as a reference, using 9,000 cDNA mouse chips produced in the Microarray Facility of AECOM. The clones were sorted into functional groups and clusters and those implicated in epithelial cell growth, proliferation and differentiation are being studied further. We have found markers for identification of liver stem/progenitor cells and are using antibodies to some of the identified cell surface proteins for isolation of fetal stem/progenitor cells.
Another aim of our research is to establish a permanent cell line of ED13 fetal liver stem/progenitor cells that can be used for further research of their biology and also for cell transplantation. Different extracellular matrix proteins and growth factors affecting the maintenance, proliferation and differentiation of liver stem/progenitor cells in culture are being studied.
Dabeva MD, Petkov PM, Sandhu J, Oren R, Laconi E, Hurston E, Shafritz DA. Proliferation and differentiation of fetal liver epithelial progenitor cells after transplantation into adult rat liver. Am J Pathol. 2000; 156:2017-2031.
Petkov PM, Kim K, Sandhu J, Shafritz DA, Dabeva MD. Identification of differentially expressed genes in epithelial stem/progenitor cells of fetal rat liver. Genomics. 2000; 68:197-209.
Sandhu JS, Petkov PM, Dabeva MD, Shafritz DA.Stem cell properties and repopulation of the rat liver by fetal liver epithelial progenitor cells. Am J Pathol. 2001; 159:1323-1334.
Shafritz DA, Dabeva MD, Grompe, M. Liver repopulation through cell transplantation. In: The Liver: Biology and Pathobiology, 4th Ed., eds., Arias IM, Boyer JL, Chisari FV, Fausto N, Schachter D, Shafritz DA, Lippincott Williams & Wilkins, Chapter 61, 2001, pp. 927-940.
Shafritz DA, Dabeva MD. Liver stem cells and model systems for liver repopulation. J Hepatol. 2002; 36:552-564. Review.
Oertel M, Rosencrantz R, Chen YQ, Thota PN, Sandhu JS, Dabeva MD, Pacchia AL, Adelson ME, Dougherty JP, Shafritz DA. Repopulation of rat liver by fetal hepatoblasts and adult hepatocytes transduced ex vivo with lentiviral vectors. Hepatology. 2003; 37:994-1005.
Dabeva MD, Shafritz DA.Hepatic stem cells and liver repopulation. Semin Liver Dis. 2003; 23:349-362. Review.
Petkov PM, Zavadil J, Goetz D, Chu T, Carver R, Rogler CE, Bottinger EP, Shafritz DA, Dabeva MD. Gene expression pattern in hepatic stem/progenitor cells during rat fetal development using complementary DNA microarrays. Hepatology. 2004; 39:617-627.
Menthena A, Deb N, Oertel M, Grozdanov PN, Sandhu J, Shah S, Guha C, Shafritz DA, Dabeva MD. Bone marrow progenitors are not the source of expanding oval cells in injured liver. Stem Cells. 2004; 22:1049-1061.
Grozdanov PN, Petkov PM, Karagyozov LK, Dabeva MD.Expression and localization of PCSK9 in rat hepatic cells. Biochem Cell Biol. 2006; 84:80-92.
Oertel M, Menthena A, Dabeva MD, Shafritz DA. Cell competition leads to a high level of normal liver reconstitution by transplanted fetal liver stem/progenitor cells. Gastroenterology 2006; 130:507-520.
Grozdanov PN, Yovchev MI, Dabeva MD. The oncofetal protein glypican-3 is a novel marker of hepatic progenitor/oval cells. Lab Invest. 2006; 86:1272-1284.
Yovchev MI, Grozdanov PN, Joseph B, Gupta S, Dabeva MD. Novel hepatic progenitor cell surface markers in the adult rat liver. Hepatology 2007; 45:139-149.
Yovchev MI, Grozdanov PN, Zhou H, Racherla H, Guha C, Dabeva MD. Identification of adult hepatic progenitor cells capable of repopulating injured rat liver. Hepatology. 2008; 47:636-647.
Yovchev MI, Zhang J, Neufeld DS, Grozdanov PN, Dabeva MD. Thymus cell antigen-1-expressing cells in the oval cell compartment. Hepatology. 2009; 50:601-611.
Yovchev MI, Dabeva MD, Oertel M. Isolation, characterization, and transplantation of adult liver progenitor cells. Methods Mol Biol. 2013; 976:37-51.
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