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A Novel Method Facilitating the Simple and Low-Cost Preparation of Human Osteochondral Slice Explants for Large-Scale Native Tissue Analysis

For in vitro modeling of human joints, osteochondral explants represent an acceptable compromise between conventional cell culture and animal models. However, the scarcity of native human joint tissue poses a challenge for experiments requiring high numbers of samples and makes the method rather uns...

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Autores principales: Spinnen, Jacob, Shopperly, Lennard K., Rendenbach, Carsten, Kühl, Anja A., Sentürk, Ufuk, Kendoff, Daniel, Hemmati-Sadeghi, Shabnam, Sittinger, Michael, Dehne, Tilo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8232634/
https://www.ncbi.nlm.nih.gov/pubmed/34203791
http://dx.doi.org/10.3390/ijms22126394
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author Spinnen, Jacob
Shopperly, Lennard K.
Rendenbach, Carsten
Kühl, Anja A.
Sentürk, Ufuk
Kendoff, Daniel
Hemmati-Sadeghi, Shabnam
Sittinger, Michael
Dehne, Tilo
author_facet Spinnen, Jacob
Shopperly, Lennard K.
Rendenbach, Carsten
Kühl, Anja A.
Sentürk, Ufuk
Kendoff, Daniel
Hemmati-Sadeghi, Shabnam
Sittinger, Michael
Dehne, Tilo
author_sort Spinnen, Jacob
collection PubMed
description For in vitro modeling of human joints, osteochondral explants represent an acceptable compromise between conventional cell culture and animal models. However, the scarcity of native human joint tissue poses a challenge for experiments requiring high numbers of samples and makes the method rather unsuitable for toxicity analyses and dosing studies. To scale their application, we developed a novel method that allows the preparation of up to 100 explant cultures from a single human sample with a simple setup. Explants were cultured for 21 days, stimulated with TNF-α or TGF-β3, and analyzed for cell viability, gene expression and histological changes. Tissue cell viability remained stable at >90% for three weeks. Proteoglycan levels and gene expression of COL2A1, ACAN and COMP were maintained for 14 days before decreasing. TNF-α and TGF-β3 caused dose-dependent changes in cartilage marker gene expression as early as 7 days. Histologically, cultures under TNF-α stimulation showed a 32% reduction in proteoglycans, detachment of collagen fibers and cell swelling after 7 days. In conclusion, thin osteochondral slice cultures behaved analogously to conventional punch explants despite cell stress exerted during fabrication. In pharmacological testing, both the shorter diffusion distance and the lack of need for serum in the culture suggest a positive effect on sensitivity. The ease of fabrication and the scalability of the sample number make this manufacturing method a promising platform for large-scale preclinical testing in joint research.
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spelling pubmed-82326342021-06-26 A Novel Method Facilitating the Simple and Low-Cost Preparation of Human Osteochondral Slice Explants for Large-Scale Native Tissue Analysis Spinnen, Jacob Shopperly, Lennard K. Rendenbach, Carsten Kühl, Anja A. Sentürk, Ufuk Kendoff, Daniel Hemmati-Sadeghi, Shabnam Sittinger, Michael Dehne, Tilo Int J Mol Sci Article For in vitro modeling of human joints, osteochondral explants represent an acceptable compromise between conventional cell culture and animal models. However, the scarcity of native human joint tissue poses a challenge for experiments requiring high numbers of samples and makes the method rather unsuitable for toxicity analyses and dosing studies. To scale their application, we developed a novel method that allows the preparation of up to 100 explant cultures from a single human sample with a simple setup. Explants were cultured for 21 days, stimulated with TNF-α or TGF-β3, and analyzed for cell viability, gene expression and histological changes. Tissue cell viability remained stable at >90% for three weeks. Proteoglycan levels and gene expression of COL2A1, ACAN and COMP were maintained for 14 days before decreasing. TNF-α and TGF-β3 caused dose-dependent changes in cartilage marker gene expression as early as 7 days. Histologically, cultures under TNF-α stimulation showed a 32% reduction in proteoglycans, detachment of collagen fibers and cell swelling after 7 days. In conclusion, thin osteochondral slice cultures behaved analogously to conventional punch explants despite cell stress exerted during fabrication. In pharmacological testing, both the shorter diffusion distance and the lack of need for serum in the culture suggest a positive effect on sensitivity. The ease of fabrication and the scalability of the sample number make this manufacturing method a promising platform for large-scale preclinical testing in joint research. MDPI 2021-06-15 /pmc/articles/PMC8232634/ /pubmed/34203791 http://dx.doi.org/10.3390/ijms22126394 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Spinnen, Jacob
Shopperly, Lennard K.
Rendenbach, Carsten
Kühl, Anja A.
Sentürk, Ufuk
Kendoff, Daniel
Hemmati-Sadeghi, Shabnam
Sittinger, Michael
Dehne, Tilo
A Novel Method Facilitating the Simple and Low-Cost Preparation of Human Osteochondral Slice Explants for Large-Scale Native Tissue Analysis
title A Novel Method Facilitating the Simple and Low-Cost Preparation of Human Osteochondral Slice Explants for Large-Scale Native Tissue Analysis
title_full A Novel Method Facilitating the Simple and Low-Cost Preparation of Human Osteochondral Slice Explants for Large-Scale Native Tissue Analysis
title_fullStr A Novel Method Facilitating the Simple and Low-Cost Preparation of Human Osteochondral Slice Explants for Large-Scale Native Tissue Analysis
title_full_unstemmed A Novel Method Facilitating the Simple and Low-Cost Preparation of Human Osteochondral Slice Explants for Large-Scale Native Tissue Analysis
title_short A Novel Method Facilitating the Simple and Low-Cost Preparation of Human Osteochondral Slice Explants for Large-Scale Native Tissue Analysis
title_sort novel method facilitating the simple and low-cost preparation of human osteochondral slice explants for large-scale native tissue analysis
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8232634/
https://www.ncbi.nlm.nih.gov/pubmed/34203791
http://dx.doi.org/10.3390/ijms22126394
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