Non-invasive Skeletal Muscle Quantification in Small Animals Using Micro-computed Tomography

Summary

This protocol uses micro-computed tomography to enable the cost-effective quantification of lean mass, including skeletal muscle and visceral tissue, adipose tissue, and skeletal tissue in small animals. It distinguishes between lean and adipose tissue, which offers significant advantages for biomedical research, particularly in translational research in small animals.

Abstract

Skeletal muscle size, mass, and composition are critical properties for studying metabolic and muscle-related diseases, as they directly impact the understanding of disease progression and treatment outcomes. Quantifying a live animal's lean, adipose, and skeletal mass is important in metabolic, physiology, pharmacologic, and geroscience studies. However, obtaining accurate body composition measurements, especially of lean mass, remains challenging due to the inherent limitations of conventional assessment techniques. Micro-computed tomography (micro-CT) is a non-invasive radiological technique that enables high-resolution visualization of internal structures in small animal models. A standardized micro-CT method can significantly enhance translational research with more reliable and impactful results, particularly during aging studies or at different time points within the same animal. Despite its potential, the lack of standardization in image acquisition and analysis methods significantly hinders the comparability of results across different studies. Herein, we present a comprehensive and detailed low-cost protocol for lean mass analysis using micro-CT to address these challenges and promote consistency in research involving small animal models.

Introduction

Size, mass, and composition are crucial skeletal muscle properties for studying muscle-related and metabolic disease mechanisms¹. Sarcopenia, cachexia, atrophy, and myopathies share common phenotypes: reduction in mass, alteration in composition, and impaired muscle function^2,3,4,5. However, quantifying the body composition in a living animal remains highly complex and technically challenging⁶.

The primary methodologies for in vivo imaging and body compositi....

Protocol

All methods were approved by the Institutional Animal Care and Use Committee of the Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro (IACUC - UFRJ; A16/23-025-20). Scans were performed on male C57BL/6 mice aged 6 and 22 months.

1. Animal preparation

1. Anesthetize mice using 2% isoflurane in a mixture of oxygen delivered via vaporization. Use a precision vaporizer for both induction and maintenance with a mixture of 100% oxygen using a precision vaporizer.......

Discussion

Evaluation through tomography is an effective and non-invasive method for obtaining detailed body composition information. Micro-CT, in particular, offers valuable outcome measures for preclinical studies. In the bone field, micro-CT has different uses, as analyzing the micro-architecture²³ and bone remodeling²⁴ are particularly interesting. Assessing the morphology of internal biological structure is also relevant in biomedical research, as analyzing the density of vascula.......

Materials

Name	Company	Catalog Number	Comments
3DSlicer, version 5.6.2	3D Slicer platform		A free and open-source software for image analysis and scientific visualization.
Amide, version 1.0.1	Amide platform		A free and open-source software used for correcting the Hounsfield Unit values in the processed DICOM images.
Amira, version 4.1	Thermo Fisher Scientific		Used to extract air and water Hounsfield Unit values from the phantom's raw data and to convert images into DICOM files.
Isoforine	Cristália		Isoflurane is a non-flammable liquid anesthetic agent for use in general inhalation anesthesia by vaporization.
Triumph XO subsystem	Gamma Medica-Ideas Flex		Advanced imaging subsystem designed for preclinical small animal imaging, offering high-resolution CT and PET capabilities for quantitative and qualitative analysis.