Cellular Plasticity of Defa4Cre-Expressing Paneth Cells in Response to Notch Activation and Intestinal Injury

Jennifer C Jones; Constance D Brindley; Nicholas H Elder; Martin G Myers Jr; Michael W Rajala; Christopher M Dekaney; Eoin N McNamee; Mark R Frey; Noah F Shroyer; Peter J Dempsey

doi:10.1016/j.jcmgh.2018.11.004

Cellular Plasticity of Defa4^Cre-Expressing Paneth Cells in Response to Notch Activation and Intestinal Injury

Cell Mol Gastroenterol Hepatol. 2019;7(3):533-554. doi: 10.1016/j.jcmgh.2018.11.004. Epub 2018 Nov 27.

Authors

Affiliations

¹ Cell Biology, Stem Cells and Development Graduate Program, University of Colorado Medical School, Aurora, Colorado.
² Division of Gastroenterology, Hepatology and Nutrition, Department of Pediatrics, University of Colorado Medical School, Aurora, Colorado.
³ Division of Metabolism, Endocrinology and Diabetes, Department of Internal Medicine, Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan.
⁴ Division of Gastroenterology, Department of Digestive Disease and Transplantation, Einstein Health Network, Philadelphia, Pennsylvania.
⁵ Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, North Carolina.
⁶ Mucosal Immunology Program, University of Colorado Medical School, Aurora, Colorado.
⁷ Saban Research Institute, Children's Hospital Los Angeles, Department of Pediatrics, Department of Biochemistry and Molecular Biology, Keck School of Medicine, University of Southern California, Los Angeles, California.
⁸ Section of Gastroenterology and Hepatology, Department of Medicine, Baylor College of Medicine, Houston, Texas.
⁹ Cell Biology, Stem Cells and Development Graduate Program, University of Colorado Medical School, Aurora, Colorado; Division of Gastroenterology, Hepatology and Nutrition, Department of Pediatrics, University of Colorado Medical School, Aurora, Colorado. Electronic address: [email protected].

Abstract

Background & aims: Loss of leucine-rich repeat-containing G-protein-coupled receptor 5-positive crypt base columnar cells provides permissive conditions for different facultative stem cell populations to dedifferentiate and repopulate the stem cell compartment. In this study, we used a defensin α4-Cre recombinase (Defa4Cre) line to define the potential of Paneth cells to dedifferentiate and contribute to intestinal stem cell (ISC) maintenance during normal homeostasis and after intestinal injury.

Methods: Small intestine and enteroids from Defa4^Cre;Rosa26 tandem dimer Tomato (tdTomato), a red fluoresent protein, (or Rosa26 Enhanced Yellow Fluorescent Protein (EYFP)) reporter, Notch gain-of-function (Defa4^Cre;Rosa26 Notch Intracellular Domain (NICD)-ires-nuclear Green Fluorescent Protein (nGFP) and Defa4^Cre;Rosa26^{reverse tetracycline transactivator-ires} Enhanced Green Fluorescent Protein (EGFP);TetO^NICD), A Disintegrin and Metalloproteinase domain-containing protein 10 (ADAM10) loss-of-function (Defa4^Cre;ADAM10^flox/flox), and Adenomatous polyposis coli (APC) inactivation (Defa4^Cre;APC^flox/flox) mice were analyzed. Doxorubicin treatment was used as an acute intestinal injury model. Lineage tracing, proliferation, and differentiation were assessed in vitro and in vivo.

Results: Defa4^Cre-expressing cells are fated to become mature Paneth cells and do not contribute to ISC maintenance during normal homeostasis in vivo. However, spontaneous lineage tracing was observed in enteroids, and fluorescent-activated cell sorter-sorted Defa4^Cre-marked cells showed clonogenic enteroid growth. Notch activation in Defa4^Cre-expressing cells caused dedifferentiation to multipotent ISCs in vivo and was required for adenoma formation. ADAM10 deletion had no significant effect on crypt homeostasis. However, after acute doxorubicin-induced injury, Defa4^Cre-expressing cells contributed to regeneration in an ADAM10-Notch-dependent manner.

Conclusions: Our studies have shown that Defa4^Cre-expressing Paneth cells possess cellular plasticity, can dedifferentiate into multipotent stem cells upon Notch activation, and can contribute to intestinal regeneration in an acute injury model.

Keywords: Chemotherapy; Defensin; Enteroid; Intestinal Stem Cells; Notch; Paneth Cell; Regeneration.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

ADAM10 Protein / metabolism
Adenoma / pathology
Adenomatous Polyposis Coli Protein / metabolism
Alleles
Animals
Cell Dedifferentiation
Cell Lineage
Cell Plasticity*
Clone Cells
Doxorubicin
Gene Deletion
Homeostasis
Hyperplasia
Integrases / metabolism*
Intestines / injuries*
Intestines / pathology*
Mice
Mitosis
Multipotent Stem Cells / metabolism
Organoids / growth & development
Organoids / pathology
Paneth Cells / metabolism*
Receptors, Notch / metabolism*
Regeneration
alpha-Defensins / metabolism*

Substances

Adenomatous Polyposis Coli Protein
Receptors, Notch
alpha-Defensins
Doxorubicin
Cre recombinase
Integrases
ADAM10 Protein

Abstract

Publication types

MeSH terms

Substances

Grants and funding