Effect of simple preservation treatments on porcine chondrocyte isolation

Upon an animal’s loss of life, decomposition commences within minutes of death. Autolytic enzymes are released into adjacent tissues when cell membranes deteriorate owing to insufficient blood circulation and other homeostatic mechanisms. These processes can severely compromise the viability of any cells that might otherwise be used for research, reproduction, or other biomedical applications. Researchers and individuals experience cases of unexpected animal death, including livestock, companion animals, or exotic species. In such cases, there is a desire to clone the animal. This study evaluated whether simple, low-cost preservation treatments could maintain porcine auricular chondrocyte viability after a 24-hour storage period at levels comparable to conventional insulated cooling on ice. The goal was to improve practical preservation methods with potential downstream applications in somatic cell nuclear transfer (SCNT), creating an improved recommendation for clients sending tissue samples to our lab. This experiment is important to gain insight into scenarios where a high value animal abruptly passes and the desire to replicate its chromosomal DNA is high. Four replicates of porcine auricular cartilage (n = 4) were divided in half and stored in insulated cooling over four treatments: (1) saline with a commercial vitamin C supplement, (2) saline with pure L-ascorbic acid, (3) saline control, and (4) an ice-only control. After 24 hours, chondrocytes were isolated by enzymatic digestion and evaluated for total cell yield and viability using trypan blue exclusion. Statistical analysis revealed no relevant differences in cell yield (P = 0.70) or viability (P = 0.68) between the treatment groups. Mean viability was numerically highest in treatment 2 (68.35 ± 11.02% viable cells) and lowest in treatment 1 (48.08 ± 21.22%), compared to controls (55.99 ± 14.18%). Total cell yield ranged from 0.66 ± 0.21 × 10⁶ cells/g (Treatment 1) to 1.03 ± 0.83 × 10⁶ cells/g (Treatment 2). The doubling times of cultured cells from all groups averaged 4.32 days, suggesting consistent growth rates regardless of preservation method. These results highlight the inherent resilience of cartilage; its dense extracellular matrix and avascular nature help delay postmortem degradation. Additionally, even basic preservation approaches, including simple solutions made from over-the-counter ingredients, effectively sustained chondrocyte viability for 24 hours. This finding is particularly relevant with its implications for livestock breeding, companion animal cloning, and conservation efforts, where simple tissue preservation can expand the feasibility of SCNT. Future research should investigate longer storage durations and interspecies comparisons to determine whether these low-cost treatments can reliably maintain cell integrity beyond 24 hours.

Name thesis:degree_name

M.S.

Level thesis:degree_level

Thesis

Discipline thesis:degree_discipline

Animal Sciences

Grantor

University of Illinois Urbana-Champaign

Year dc:date

2025

Author dc:creator

• Wilson, Kenneth C.

Contributors dc:contributor

• Wheeler, Matthew B.
• Knox, Robert V
• Emmert, Jason L.

• Somatic Cell Nuclear Transfer
• Post-mortem Tissue Preservation

Statement dc:rights

• Copyright 2025 Kenneth Wilson

Language dc:language

en, eng

Handle dc:identifier

https://hdl.handle.net/2142/129902

OAI record oai:identifier

https://www.ideals.illinois.edu/oai-pmh?verb=GetRecord&metadataPrefix=etdms&identifier=oai%3Awww.ideals.illinois.edu%3A2142%2F129902