Isabelle Garcia – Gilbert Vassart

Our research

RESEARCH
Fundamental research in the stem cell field is instrumental to ultimately develop therapies with improved efficacy in regenerative medicine and oncology. As one of the tissues with highest self-renewing rates under steady-state conditions, the gastrointestinal epithelium constitutes an excellent model to better understand how resident adult stem cells contribute to tissue homeostasis, damage-induced regeneration processes or cancer.

LGR receptors: function in developmental progenitors, intestinal stem cells under homeostasis, regeneration and tumorigenic processes)
In the past decade, significant advances in this field have allowed identifying intestinal stem cells and culturing them as “mini-guts” in a tridimensional ex vivo system (Figure 1). As an essential player in stem cell proliferation and cell-fate decision, the Wnt/β-catenin signaling cascade is submitted to an extremely fine-tune control to maintain both stemness and differentiation in the epithelium. Importantly, mutations in components of this pathway, such as those leading to loss-of-function of the APC tumor suppressor, are considered initiating and rate-limiting mutations for most colorectal cancers (CRC).
The Leucine-rich repeat-containing G protein-coupled receptors (Lgr4/Lgr5/Lgr6), belonging to the large family of drug-targetable GPCR receptors are key modulators of the Wnt pathway in intestinal stem cells. Nevertheless, the molecular mechanisms involving Lgr receptors activity and Wnt signaling are not yet fully dissected. Moreover, in mouse oncological models, there are evidences that cells expressing Lgr4 and Lgr5 are tumor-initiating cells but the role itself of each of these two receptors in the tumor cells has not yet been addressed. Our goal is to get further understanding about the biological function of Lgr receptors on Wnt signaling modulation in intestinal stem cells during development, under adult homeostasis, regeneration, and tumorigenic processes. By combining in vivo and ex vivo studies using mouse models, we aim to determine whether Lgrs could represent valuable drug targets to better control the Wnt pathway in regenerative medicine or oncology.

Dissection of Trop2-associated processes regeneration in adult gut
Whereas coexistence of several pools of stem cells has been reported in the adult intestinal and gastric epithelia that ensure tissue homeostasis under steady-state conditions, much less was known till recently about the identity of fetal progenitors involved in tissue morphogenesis. Our group has isolated and characterized these cells from the mouse small intestine and pre-glandular stomach at embryonic stage E14.5 These progenitors grow as undifferentiated spheroids and express a transcriptome very different from the adult stem cells. Remarkedly, these fetal cells are only transiently detected during the initial phases of gastric gland formation and intestinal villogenesis in the mouse, being then replaced by the precursors of adult Lgr5-positive stem cells (Figure 1). One of the markers highly expressed in these progenitors is the cell surface molecule Trop2/Tacstd2 (Tumor-associated calcium signal transducer 2), which is normally undetectable in adult homeostatic stomach and intestine tissues but overexpressed in gastric and colorectal tumors.


Schematic representation of the intestinal epithelium in the late embryonic development and in adult stages under homeostasis or after injury. When cultured ex vivo, fetal and adult-derived epithelial cells generate different kinds of structures (spheroids or organoids).

 

 

Recently, our laboratory has provided evidences that damage of the mouse adult stomach and intestine epithelia activates partial re-expression of a “fetal-like” genetic program in the epithelium, leading to the appearance of regenerating cells characterized by the representative cell surface molecule Trop2 (Figure 1). Fetal progenitors and adult regenerating Trop2 cells, involved in building or re-building brand-new epithelia respectively, share common signaling pathways involved in tissue morphogenesis, cell proliferation and migration. These findings may be particularly relevant for gastrointestinal disorders like the chronic inflammatory bowel diseases (IBDs) that are characterized by sub-mucosal accumulation of inflammatory cells and severe damage of the epithelial layer, resulting in development of refractory ulcers and in some cases, evolving towards cancer. Recent studies point to the importance of epithelial cells for efficient “mucosal healing” and maintenance of long-term remission in these relapsing-remitting diseases. Thus, it is essential to get better knowledge of the molecular and cellular mechanisms associated with epithelial repair in experimental models of IBD, in order to improve mucosal healing in patients in the future and prevent cancer. Our goal is to better characterize the mechanisms triggering the “de-differention-to-re-differentiation” conversion state that is necessary to fully restore the functionality of the epithelium. Since Trop2 is a relevant prognosis marker in gastric and colorectal cancer cells, one question to address is whether non-adequate persistence of a regenerative process can ultimately lead to malignant transformation. Moreover, Trop2 is a cell surface molecule reported to transduce various signaling pathways in cancer cells. We are currently investigating the potential signaling role of this marker in fetal progenitors and adult regenerating cells.

Group members


Marie-Isabelle Garcia : Principal Investigator (Marie.Garcia@ulb.ac.be)

phone # +32 (0)2 555 4190

She performed her PhD in Biochemistry at the University Paris VII (France) in 1996 and pursued with a postodoctoral position at the Institut Pasteur (France) where she studied the role of cell adhesion as a virulence factor in the infectious process of pathogenic E. coli strains. She performed a postdoctoral training at the University La Sapienza of Rome (Italy) to study the role of the major capsid protein VP1 from Polyomavirus in the early stages of infection and host specificity (1998-2004). She joined the IRIBHM institute in 2006 and developed research on intestinal stem cells and Lgr receptors.

Gilbert Vassart (gvassart@ulb.ac.be)

phone # +32 (0)2 555 4185

Gilbert Vassart pioneered the use of molecular genetics approaches to solve endocrinology issues. Over the past three decades his group identified gain- and loss-of-function mutations in several endocrine disease and cloned a host of G protein-coupled receptors (GPCRs), opening the way to what has been coined “reverse pharmacology”. In collaboration with Marc Parmentier, this led to the deorphanization of several GPCRs, including CCR5, the main co-receptor for infection by HIV-1.
See: Pubmed

Morgane Leprovots: technician
She obtained a bachelor’s degree in Cellular biology, genetics, microbiology and physiology in 2011 at the University of Rennes (France) and a vocational bachelor’s in biology in 2012 at the IUT Tours (France). She joined our lab in September 2013.

Romain Gerbier: postdoctoral position
He obtained his PhD in Cellular Pharmacology in 2011 working on the role of central neuropeptides in regulation of body fluid homeostasis and cardiovascular functions (Collège de France, France). He performed postdoctoral research experiences on development of metabolically stable apelin analogs (Collège de France, 2011-2013) and on the in vivo characterization of mouse MTR/5-HT2c heteromerization (Institut Cochin, France, 2013-2016). He has joined the group in October 2016 to study Lgr receptors biology.

Maryam Marefati: PhD student
She obtained a bachelor/master’s degree in biology-Zoology in 2011 from Ferdowsi University of Mashhad (Iran) and a Master of Molecular Biology in 2016 from the Vrije Universiteit Brussels where she has experimentally characterized hepatic stellate cells in a 3D culture. She has joined our laboratory in October 2017 to study regeneration processes in adult gastrointestinal tissues.

Alumni
Valeria Fernandez-Vallone: Post-doctoral position from 2012 to 2017. She has characterized intestinal and gastric fetal progenitors and regenerating cells in the mouse adult gastric epithelium.
Gabriela Vasile: she participated to the characterization of murine fetal progenitors (2011-2015).
Sofia Cheyroux: she studied the ex vivo function of particular GPCRs in intestinal progenitors and adult stem cells as a trainee (Feb-Aug. 2015).
Roxana Mustata: postdoctoral position from 2009 to 2012. She studied the role of the Lgr4 receptor in murine intestinal stem cells and characterized murine fetal intestinal progenitors.
Tom Van Loy: postdoctoral position from 2009 to 2011. He worked on Lgr de-orphanization and participated to the study of the role of the Lgr4 receptor in murine intestinal stem cells. He also contributed to de-orphanization of the Drosophila ortholog Lgr2 ligand (Bursicon).
Maria-Angela Ghiani: medical doctor trainee in molecular biology from 2006 to 2007. She studied the role of Lgr5 receptor during murine intestinal development.
Fernando Mendive: postdoctoral position from 2002 to 2006. He worked on Lgr structure-function and on the role of Lgr4 in male fertility. He also contributed to de-orphanization of the Drosophila ortholog Lgr2 ligand (Bursicon).
Gregory Van Schoore: Obtained his PhD in 2008 where he defined the distribution of the Lgr4 receptor in the mouse tissues and he contributed to the study of the role of Lgr4 in male fertility.
Sylvie Claeysen: postdoctoral position from 2000 to 2002. She started the study of Lgrs in our lab.

Publications

LGRs receptors as peculiar GPCRs involved in cancer (Review). Garcia MI. J Stem Cell Res Med 2017, doi: 10.15761/JSCRM.1000116.

Ex vivo culture of gastric spheroids. Fernandez Vallone V., Leprovots M., Vassart G and Garcia MI. Bio-protocols . 2017. DOI 10.21769/2089.

Ex vivo culture of adult antral glands. Fernandez Vallone V., Leprovots M., Vassart G and Garcia MI. Bio-protocols. 2017. DOI 10.21769/2088.

LGR4 and LGR5 Regulate Hair Cell Differentiation in the Sensory Epithelium of the Developing Mouse Cochlea. Zak, Magdalena; Van Oort, Thys; Hendriksen, Ferry G; Garcia, Marie-Isabelle; Vassart, Gilbert; Grolman, Wilko. Front Cell Neurosci. 2016; 10: 186. doi: 10.3389/fncel.2016.00186 PMCID: PMC4988241

Stem Cells in Adult Homeostasis, Regeneration and Tissue Development of the Digestive Tract Epithelium. Valeria Fernandez Vallone and Marie-Isabelle Garcia. Int J Stem Cell Res Ther 2016, 3:038 Volume 3 | Issue 2 ISSN: 2469-570X

Building and rebuilding gastric epithelium: Trop2 as a common denominator. Valeria Fernandez Vallone and Marie-Isabelle Garcia. Atlas of Science, another view on science. Published online August 15th, 2016.

Trop2 marks transient gastric fetal epithelium and adult regenerating cells after epithelial damage: Fernandez Vallone V, Leprovots M, Strollo S, Vasile G, Lefort A, Libert F, Vassart G, Garcia MI.
Development. 2016 May 1;143(9):1452-63. doi: 10.1242/dev.131490. Epub 2016 Mar 17.

Identification of Lgr5-independent spheroid-generating progenitors of the mouse fetal intestinal epithelium: Mustata RC, Vasile G, Fernandez-Vallone V, Strollo S, Lefort A, Libert F, Monteyne D, Pérez-Morga D, Vassart G, Garcia MI. Cell Rep. 2013 Oct 31;5(2):421-32.

Identification des progéniteurs de l’épithélium intestinal foetal: M/S médecine/sciences. 2014 4:353-355. Garcia MI, Vassart G.

Chapter entitled :”Discovery and Characterization of Leucine-Rich Repeat-Containing G Protein-Coupled Receptors from the book “The Genetics of G Protein-Coupled Receptors: Methods for the Post-Genomic Era” edited by Craig W. Stevens, PhD; Springer Science+Business Media, LLC; Humana Press. MI Garcia, V. Fernandez-Vallone and G. Vassart.

Lgr4 is required for Paneth cell differentiation and maintenance of intestinal stem cells ex vivo. Mustata RC, Van Loy T, Lefort A, Libert F, Strollo S, Vassart G, Garcia MI. EMBO Rep. 2011 Jun;12(6):558-64.

LGR5 deficiency deregulates Wnt signaling and leads to precocious Paneth cell differentiation in the fetal intestine. Garcia MI, Ghiani M, Lefort A, Libert F, Strollo S, Vassart G. Dev Biol. 2009 Jul 1;331(1):58-67.

Three-dimensional reconstruction of efferent ducts in wild-type and Lgr4 knock-out mice. Lambot MA, Mendive F, Laurent P, Van Schoore G, Noël JC, Vanderhaeghen P, Vassart G. Anat Rec (Hoboken). 2009 Apr;292(4):595-603.

Defective postnatal development of the male reproductive tract in LGR4 knockout mice. Mendive F, Laurent P, Van Schoore G, Skarnes W, Pochet R, Vassart G. Dev Biol. 2006 Feb 15;290(2):421-34.

Expression pattern of the orphan receptor LGR4/GPR48 gene in the mouse. Van Schoore G, Mendive F, Pochet R, Vassart G. Histochem Cell Biol. 2005 Jul;124(1):35-50.

Drosophila molting neurohormone bursicon is a heterodimer and the natural agonist of the orphan receptor DLGR2. Mendive FM, Van Loy T, Claeysen S, Poels J, Williamson M, Hauser F, Grimmelikhuijzen CJ, Vassart G, Vanden Broeck J. FEBS Lett.2005 Apr 11;579(10):2171-6.