The University of Southampton
University of Southampton Institutional Repository
Warning ePrints Soton is experiencing an issue with some file downloads not being available. We are working hard to fix this. Please bear with us.

Examining lysyl oxidase-like modulation of collagen architecture in 3D spheroid models of idiopathic pulmonary fibrosis via second-harmonic generation microscopy

Examining lysyl oxidase-like modulation of collagen architecture in 3D spheroid models of idiopathic pulmonary fibrosis via second-harmonic generation microscopy
Examining lysyl oxidase-like modulation of collagen architecture in 3D spheroid models of idiopathic pulmonary fibrosis via second-harmonic generation microscopy

Significance: Idiopathic pulmonary fibrosis (IPF) patients have a poor prognosis with short lifespan following diagnosis as there are limited effective treatment options. Despite matrix stiffening being the hallmark of the disease there remains a lack of knowledge surrounding the underlying collagen alterations in the disease. Specifically, while increased collagen crosslinking has been implicated, the resulting effects on collagen macro/supramolecular changes have not been explored. Aim: We sought to determine if second-harmonic generation (SHG) microscopy could characterize differences in the collagen architecture in 3D spheroid models of IPF grown under different crosslinking modulation conditions (promotion and inhibition). Approach: We used SHG metrics based on the fiber morphology, relative SHG brightness, and macro/supramolecular structure by SHG polarization analyses to compare the structure of the IPF spheroids. Results: Comparison of the fiber morphology of the spheroids showed that the control group had the longest, straightest, and thickest fibers. The spheroids with crosslink enhancement and inhibition had the highest and lowest SHG conversion efficiencies, respectively, consistent with the resulting harmonophore density. SHG polarization analyses showed that the peptide pitch angle, alignment of collagen molecules, and overall chirality were altered upon crosslink modulation and were also consistent with reduced organization relative to the control group. Conclusions: While no single SHG signature is associated with crosslinking, we show that the suite of metrics used here is effective in delineating alterations across the collagen architecture sizescales. The results largely mirror those of human tissues and demonstrate that the combination of 3D spheroid models and SHG analysis is a powerful approach for hypothesis testing the roles of operative cellular and molecular factors in IPF.

collagen, crosslinking, polarization, second-harmonic generation, stiffness
1083-3668
James, Darian S.
7af7b639-e65a-4afb-abcc-7d529fbb444d
Brereton, Christopher J.
948ca4ea-b04c-4b7a-bfe4-f79f184d7e43
Davies, Donna E.
7de8fdc7-3640-4e3a-aa91-d0e03f990c38
Jones, Mark G.
a6fd492e-058e-4e84-a486-34c6035429c1
Campagnola, Paul J.
8c7a886e-b4b7-4857-bd46-4e4889d7b8f9
James, Darian S.
7af7b639-e65a-4afb-abcc-7d529fbb444d
Brereton, Christopher J.
948ca4ea-b04c-4b7a-bfe4-f79f184d7e43
Davies, Donna E.
7de8fdc7-3640-4e3a-aa91-d0e03f990c38
Jones, Mark G.
a6fd492e-058e-4e84-a486-34c6035429c1
Campagnola, Paul J.
8c7a886e-b4b7-4857-bd46-4e4889d7b8f9

James, Darian S., Brereton, Christopher J., Davies, Donna E., Jones, Mark G. and Campagnola, Paul J. (2021) Examining lysyl oxidase-like modulation of collagen architecture in 3D spheroid models of idiopathic pulmonary fibrosis via second-harmonic generation microscopy. Journal of Biomedical Optics, 26 (6), [066501]. (doi:10.1117/1.JBO.26.6.066501).

Record type: Article

Abstract

Significance: Idiopathic pulmonary fibrosis (IPF) patients have a poor prognosis with short lifespan following diagnosis as there are limited effective treatment options. Despite matrix stiffening being the hallmark of the disease there remains a lack of knowledge surrounding the underlying collagen alterations in the disease. Specifically, while increased collagen crosslinking has been implicated, the resulting effects on collagen macro/supramolecular changes have not been explored. Aim: We sought to determine if second-harmonic generation (SHG) microscopy could characterize differences in the collagen architecture in 3D spheroid models of IPF grown under different crosslinking modulation conditions (promotion and inhibition). Approach: We used SHG metrics based on the fiber morphology, relative SHG brightness, and macro/supramolecular structure by SHG polarization analyses to compare the structure of the IPF spheroids. Results: Comparison of the fiber morphology of the spheroids showed that the control group had the longest, straightest, and thickest fibers. The spheroids with crosslink enhancement and inhibition had the highest and lowest SHG conversion efficiencies, respectively, consistent with the resulting harmonophore density. SHG polarization analyses showed that the peptide pitch angle, alignment of collagen molecules, and overall chirality were altered upon crosslink modulation and were also consistent with reduced organization relative to the control group. Conclusions: While no single SHG signature is associated with crosslinking, we show that the suite of metrics used here is effective in delineating alterations across the collagen architecture sizescales. The results largely mirror those of human tissues and demonstrate that the combination of 3D spheroid models and SHG analysis is a powerful approach for hypothesis testing the roles of operative cellular and molecular factors in IPF.

This record has no associated files available for download.

More information

Published date: 1 June 2021
Additional Information: Funding Information: PJC gratefully acknowledges support under 1R21HL126190-01A1 and NSF CBET – 1402757; and DSJ gratefully acknowledges support under NSF DGE-1747503. CJB acknowledges the support of the NIHR Southampton Biomedical Research Centre. MGJ acknowledges support from the Wellcome Trust (Grant No. 100638/Z/12/Z). Dr. Wolfgang Jarolimek from Pharmaxis kindly provided PXS-S2A. Publisher Copyright: © The Authors. Published by SPIE under a Creative Commons Attribution 4.0 Unported License. Distribution or reproduction of this work in whole or in part requires full attribution of the original publication, including its DOI. Copyright: Copyright 2021 Elsevier B.V., All rights reserved.
Keywords: collagen, crosslinking, polarization, second-harmonic generation, stiffness

Identifiers

Local EPrints ID: 451023
URI: http://eprints.soton.ac.uk/id/eprint/451023
ISSN: 1083-3668
PURE UUID: dda99757-5129-49ad-b959-4e9836bf74ba
ORCID for Donna E. Davies: ORCID iD orcid.org/0000-0002-5117-2991
ORCID for Mark G. Jones: ORCID iD orcid.org/0000-0001-6308-6014

Catalogue record

Date deposited: 03 Sep 2021 16:31
Last modified: 26 Nov 2021 02:52

Export record

Altmetrics

Contributors

Author: Darian S. James
Author: Christopher J. Brereton
Author: Donna E. Davies ORCID iD
Author: Mark G. Jones ORCID iD
Author: Paul J. Campagnola

Download statistics

Downloads from ePrints over the past year. Other digital versions may also be available to download e.g. from the publisher's website.

View more statistics

Atom RSS 1.0 RSS 2.0

Contact ePrints Soton: eprints@soton.ac.uk

ePrints Soton supports OAI 2.0 with a base URL of http://eprints.soton.ac.uk/cgi/oai2

This repository has been built using EPrints software, developed at the University of Southampton, but available to everyone to use.

We use cookies to ensure that we give you the best experience on our website. If you continue without changing your settings, we will assume that you are happy to receive cookies on the University of Southampton website.

×