Visualizing Cathepsin K‐Cre Expression at the Single‐Cell Level with GFP Reporters. (21st December 2022)

• Record Type: Journal Article
• Authors: Chai, Wenhuan; Hao, Weiwei; Liu, Jintao; Han, Zhenglin; Chang, Shiyu; Cheng, Liben; Sun, Mingxin; Yan, Guofang; Liu, Zemin; Liu, Yin; Zhang, Guodong; Xing, Li; Chen, Hongqian; Liu, Peng
• Is Part Of: JBMR plus. Volume 7:Number 1(2023)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2022-12-21
• Subjects: AGING -- CATHEPSIN K -- CRE -- GFPS -- OSTEOBLASTS -- OSTEOCLASTS; Bones -- Diseases -- Periodicals; Bones -- Metabolism -- Periodicals; Orthopedics -- Periodicals; 612.75104
• DOI: 10.1002/jbm4.10706
• Languages: English
• ISSNs: 2473-4039

Abstract

ABSTRACT: The Cre/lox system is a fundamental tool for functional genomic studies, and a number of Cre lines have been generated to target genes of interest spatially and temporally in defined cells or tissues; this approach has greatly expanded our knowledge of gene functions. However, the limitations of this system have recently been recognized, and we must address the challenge of so‐called nonspecific/off‐target effects when a Cre line is utilized to investigate a gene of interest. For example, cathepsin K (Ctsk) has been used as a specific osteoclast marker, and Cre driven by its promoter is widely utilized for osteoclast investigations. However, Ctsk‐Cre expression has recently been identified in other cell types, such as osteocytes, periosteal stem cells, and tenocytes. To better understand Ctsk‐Cre expression and ensure appropriate use of this Cre line, we performed a comprehensive analysis of Ctsk‐Cre expression at the single‐cell level in major organs and tissues using two green fluorescent protein (GFP) reporters (ROSA nT‐nG and ROSA tdT) and a tissue clearing technique in young and aging mice. The expression profile was further verified by immunofluorescence staining and droplet digital RT‐PCR. The results demonstrate that Ctsk‐Cre is expressed not only in osteoclasts but also at various levels in osteoblast lineage cells and other major organs/tissues, particularly in the brain, kidney, pancreas, and blood vessels. Furthermore, Ctsk‐Cre expression increases markedly in the bone marrow, skeletal muscle, and intervertebral discs in aging mice. These data will be valuable for accurately interpreting data obtained from in vivo studies using Ctsk‐Cre mice to avoid potentially misleading conclusions. © 2022 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research. Abstract : We performed a comprehensive analysis of Ctsk‐Cre expression at the single‐cell level in major organs and tissues using two GFP reporters. These results demonstrate that Ctsk‐Cre is expressed not only in osteoclasts but also at various levels in osteoblast lineage cells and other major organs/tissues, particularly in the brain, kidney, pancreas, and blood vessels. These data will be valuable for accurately interpreting data to avoid potentially misleading conclusions.