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

International Journal of Molecular Sciences

Carsten Rendenbach¹  Daniel Kendoff²  Ufuk Sentürk³  Jacob Spinnen⁴  Tilo Dehne⁴  Shabnam Hemmati-Sadeghi⁴  Lennard K. Shopperly⁴  Michael Sittinger⁴  Anja A. Kühl⁵ 

[1] Department of Oral and Maxillofacial Surgery, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt Universität zu Berlin, and Berlin Institute of Health, 13353 Berlin, Germany;Department of Orthopaedic Surgery, Helios Klinikum Berlin-Buch, 13125 Berlin, Germany;Department of Orthopedics, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt Universität zu Berlin, and Berlin Institute of Health, 13353 Berlin, Germany;Department of Rheumatology, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt Universität zu Berlin, and Berlin Institute of Health, 10117 Berlin, Germany;iPATH Histopathology Core Unit, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt Universität zu Berlin, and Berlin Institute of Health, 13353 Berlin, Germany;

关键词: osteoarthritis;    osteochondral explant culture;    joint modelling;    pharmacological assay;    native tissue analysis;   

DOI  :  10.3390/ijms22126394

来源: DOAJ

【 摘 要 】

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.

【 授权许可】

Unknown