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Comparison of the effects of two methods of euthanasia on post mortem changes in rats: histopathological and molecular findings

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Abstract

Postmortem changes (PMCs) not only affect the results of experimental studies, but also determine postmortem intervals in forensic sciences. The present study aimed to assess the effects of two methods of laboratory animals’ euthanasia on PMCs in rats. In this experimental study, 10 female rats were randomly assigned to two equal groups and were euthanized using the inhalation of CO2 (gas-treated group) or over-dose intramuscular injection of ketamine/xylazine (drug-treated group). Kidney and liver tissue samples were collected at baseline and 2, 4, 6, 8, 10, 12, 24, and 48 h after euthanasia and were subjected to histopathological examinations. The expression of liver-specific microRNA-122 (miR-122) was also assessed in each time point via a SYBR green real-time polymerase chain reaction (PCR) assay. Finally, miR-122 target genes and related functional pathways were identified through bioinformatics analysis. The progression of PMCs in the drug-treated group was faster than the gas-treated group. The expression of miR-122 was significantly (P < 0.0001) upregulated in the drug-treated group at 4, 10, and 24 h in comparison to the gas-treated group; however, it was downregulated at 6, 8, and 48 h after euthanasia. The biosynthesis of amino acids, glycolysis/gluconeogenesis, and carbon metabolism as well as the glucagon, Hedgehog, cGMP-PKG, and neurotrophin signaling pathways were identified as the significant pathways related to miR-122 target genes. The findings indicated that methods of euthanasia for laboratory animals could cause changes at microscopic and molecular levels. Therefore, researchers should consider this issue in the design phase of their studies.

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Acknowledgements

Hereby, the authors would like to thank Ms. A. Keivanshekouh at the Research Consultation Center (RCC) of Shiraz University of Medical Sciences for improving the use of English in the manuscript.

Funding

This study was financially supported by grant no. 95–01-45–13692 from the Vice-chancellor for Research Affairs of Shiraz University of Medical Sciences, Shiraz, Iran.

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Authors and Affiliations

  1. Department of Comparative Biomedical Sciences, School of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences, Shiraz, Iran

    Amin Derakhshanfar, Mehdi Kian & Zeinab Dehghan

  2. Diagnostic Laboratory Sciences and Technology Research Center, School of Paramedical Sciences, Shiraz University of Medical Sciences, Shiraz, Iran

    Amin Derakhshanfar & Javad Moayedi

  3. Center of Comparative and Experimental Medicine, Shiraz University of Medical Sciences, Shiraz, Iran

    Amin Derakhshanfar, Abouzar Valizadeh & Javad Moayedi

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  1. Amin Derakhshanfar
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  2. Mehdi Kian
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Correspondence to Javad Moayedi.

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All animal experiments were approved by the local Ethics Committee of Shiraz University of Medical Sciences, Shiraz, Iran (Approval ID: IR.SUMS.REC.1395.S1241). All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.

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Derakhshanfar, A., Kian, M., Dehghan, Z. et al. Comparison of the effects of two methods of euthanasia on post mortem changes in rats: histopathological and molecular findings. Comp Clin Pathol 31, 815–826 (2022). https://doi.org/10.1007/s00580-022-03385-7

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