In Vivo Clonal Analysis of Cardiomyocytes

Ngoc B Nguyen; G Esteban Fernandez; Yichen Ding; Tzung Hsiai; Reza Ardehali

doi:10.1007/978-1-0716-0668-1_18

In Vivo Clonal Analysis of Cardiomyocytes

Methods Mol Biol. 2021:2158:243-256. doi: 10.1007/978-1-0716-0668-1_18.

Authors

Ngoc B Nguyen^{1

2

3}, G Esteban Fernandez⁴, Yichen Ding¹, Tzung Hsiai¹, Reza Ardehali^{5

6

7

8}

Affiliations

¹ Division of Cardiology, Department of Internal Medicine, David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, CA, USA.
² Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research, University of California, Los Angeles, Los Angeles, CA, USA.
³ Molecular, Cellular and Integrative Physiology Graduate Program, University of California, Los Angeles, Los Angeles, CA, USA.
⁴ Children's Hospital Los Angeles and the Saban Research Institute, Los Angeles, CA, USA.
⁵ Division of Cardiology, Department of Internal Medicine, David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, CA, USA. RArdehali@mednet.ucla.edu.
⁶ Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research, University of California, Los Angeles, Los Angeles, CA, USA. RArdehali@mednet.ucla.edu.
⁷ Molecular, Cellular and Integrative Physiology Graduate Program, University of California, Los Angeles, Los Angeles, CA, USA. RArdehali@mednet.ucla.edu.
⁸ Molecular Biology Institute, University of California, Los Angeles, Los Angeles, CA, USA. RArdehali@mednet.ucla.edu.

PMID: 32857378
DOI: 10.1007/978-1-0716-0668-1_18

Abstract

The development of the CreER/LoxP system has enabled temporal control and cell type specificity of gene activation or repression. A common application of this system involves lineage tracing and examining the proliferative capacity of cells of interest through clonal analysis. Here, we describe a method of performing 2- and 3-dimensional clonal analysis of cardiomyocytes (CMs) using the Rainbow reporter mouse model. We outline the process of using the Cell Counter plug-in tool in ImageJ to quantify the number of clones as well as the number of cells within each clone. For 3-dimensional analysis, we describe the tissue clearing technique, CLARITY, in conjunction with light-sheet imaging to obtain digital slices of the whole heart that can be reconstructed and clone volumes quantified using the Imaris software.

Keywords: Cardiac regeneration; Cell proliferation; Clonal analysis; CreER/LoxP system; Fluorescent reporter; Imaris; Light-sheet imaging; Tissue clearing.

Publication types

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

MeSH terms

Animals
Cell Differentiation*
Cell Lineage*
Cell Separation / methods*
Clone Cells
Disease Models, Animal
Genes, Reporter*
Mice
Myocytes, Cardiac / cytology*
Pluripotent Stem Cells / cytology*
Regeneration*

Abstract

Publication types

MeSH terms

Grants and funding