Temperature-dependent alterations in skeletal muscle during the postmortem interval: Structural, metabolic, and lncRNA expression changes.

BACKGROUND

Estimation of the postmortem interval (PMI) remains a challenge in forensic practice. Although environmental temperature is known to influence postmortem biological processes, its effects on structural, metabolic, and transcriptional changes in skeletal muscle remain poorly characterized.

METHODS

Male C57 mice were euthanized and carcasses were maintained at 30 °C, 20 °C, or 4 °C. Skeletal muscle samples were collected at 0, 4, 8, 12, 24, and 36 h postmortem. Histological alterations were assessed by H&E staining. Adenylate energy status was evaluated by measuring ATP, ADP, and AMP using high-performance liquid chromatography, followed by calculation of energy charge and the ATP/ADP ratio. Autophagy-related markers were analyzed by Western blot, transmission electron microscopy, and immunofluorescence. Long non-coding RNA (lncRNA) expression profiles were examined by sequencing, and selected candidates were validated by quantitative PCR.

RESULTS

Skeletal muscle structure progressively deteriorated with increasing PMI, with changes occurring earlier at 30 °C and delayed at 4 °C. ATP levels declined postmortem in all groups, accompanied by increased AMP and reduced energy charge and ATP/ADP ratio. The decline in energy status was accelerated at higher temperature and was slowed at lower temperature. Autophagy-related markers showed early elevation followed by a later reduction, with peak changes occurring earlier at higher temperature and later at lower temperature. Transcriptomic analysis identified 1643 high-confidence lncRNAs. Differential expression patterns varied among temperature groups. Six candidate lncRNAs displayed distinct time-dependent expression trajectories under different temperature conditions, and their temporal trends were described using fourth-order polynomial regression (R2 = 0.87-0.92).

CONCLUSION

Postmortem skeletal muscle exhibits structural, metabolic, and transcriptional alterations that are influenced by environmental temperature. These temperature-dependent temporal patterns may provide molecular information relevant to PMI assessment.