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J Appl Physiol (1985). 2017 Jan 1;122(1):161-169. doi: 10.1152/japplphysiol.00838.2016. Epub 2016 Nov 17.

Nondestructive cryomicro-CT imaging enables structural and molecular analysis of human lung tissue.

Author information

1
University of British Columbia, Centre for Heart Lung Innovation, Vancouver, British Columbia, Canada; dragos.vasilescu@hli.ubc.ca.
2
Materials Science and Engineering, McMaster University, Hamilton, Ontario, Canada.
3
University of British Columbia, Centre for Heart Lung Innovation, Vancouver, British Columbia, Canada.
4
Paige Instruments, Woodland, California.
5
Section of Computational Biomedicine, Department of Medicine, Boston University Medical Center, Boston, Massachusetts.
6
Lung Transplant Unit, Department of Clinical and Experimental Medicine, KU Leuven, Leuven, Belgium.
7
Janssen Disease Interception Accelerator, Janssen Pharmaceutical Companies of Johnson and Johnson, Raritan, New Jersey; and.
8
Division of Thoracic Surgery, University of Pennsylvania, Philadelphia, Pennsylvania.

Abstract

Micro-computed tomography (CT) enables three-dimensional (3D) imaging of complex soft tissue structures, but current protocols used to achieve this goal preclude cellular and molecular phenotyping of the tissue. Here we describe a radiolucent cryostage that permits micro-CT imaging of unfixed frozen human lung samples at an isotropic voxel size of (11 µm)3 under conditions where the sample is maintained frozen at -30°C during imaging. The cryostage was tested for thermal stability to maintain samples frozen up to 8 h. This report describes the methods used to choose the materials required for cryostage construction and demonstrates that whole genome mRNA integrity and expression are not compromised by exposure to micro-CT radiation and that the tissue can be used for immunohistochemistry. The new cryostage provides a novel method enabling integration of 3D tissue structure with cellular and molecular analysis to facilitate the identification of molecular determinants of disease.

NEW & NOTEWORTHY:

The described micro-CT cryostage provides a novel way to study the three-dimensional lung structure preserved without the effects of fixatives while enabling subsequent studies of the cellular matrix composition and gene expression. This approach will, for the first time, enable researchers to study structural changes of lung tissues that occur with disease and correlate them with changes in gene or protein signatures.

KEYWORDS:

cryostage; gene expression; lung; micro-computed tomography; soft tissue imaging

PMID:
27856720
PMCID:
PMC6442661
DOI:
10.1152/japplphysiol.00838.2016
[Indexed for MEDLINE]
Free PMC Article

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