The collection and preservation of postmortem genetic material from recently deceased animals of rare and endangered species represent a critical yet underexplored avenue in conservation biology. While extensive research has been conducted on the human postmortem interval (PMI), there is a notable gap in understanding the postmortem preservation of germplasm in endangered species.
Objectives:
This study aimed to investigate the dynamics of apoptosis in various tissues of the Yangtze sturgeon at different postmortem time points, and to provide a reference for identifying the optimal time window for germplasm preservation in rare and endangered fish in the wild.
Methods:
Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphated nick-end labeling assay and tissue morphology analyses were used to investigate apoptosis in the brain, heart, fin, liver, gonad, muscle, spleen, and skin of the Yangtze sturgeon at five different time points 0, 4, 8, 12, and 16 hours postmortem.
Results:
The results revealed a dynamic pattern of apoptosis. All tissues exhibited a time-dependent increase in apoptotic rate, indicating a clear correlation between PMI and apoptosis progression. This temporal pattern underscores the importance of timely genetic resource preservation, as the integrity of genetic material deteriorates progressively after death. Histomorphological analysis further demonstrated progressive degradation of tissue structure, especially in metabolically active tissues such as the gonad and fin.
Conclusion:
Based on the findings, we recommend that the genetic resources of the Yangtze sturgeon be preserved as soon as possible after death, particularly within the first 12 hours when tissue integrity remains sufficient for viable cell isolation or cryopreservation. This window is critical for metabolically active tissues, which show marked changes over time and may be important for postmortem identification. Further research should explore cryopreservation and antioxidant treatments to extend the preservation window for germplasm resources, ensuring the long-term viability of these valuable genetic materials.
LiJ, WangC, PanW, et al.Migration and distribution of adult hatchery reared Yangtze sturgeons (Acipenser dabryanus) after releasing in the upper Yangtze River and its implications for stock enhancement. J Appl Ichthyol, 2021; 37(1):3–11; doi: 10.1111/jai.14117
2.
BoscariE, WuJ, JiangT, et al.The last giants of the Yangtze River: A multidisciplinary picture of what remains of the endemic Chinese sturgeon. Sci Total Environ, 2022; 843:157011; doi: 10.1016/j.scitotenv.2022.157011
3.
ZhangH, KangM, ShenL, et al.Rapid change in Yangtze fisheries and its implications for global freshwater ecosystem management. Fish Fish, 2020; 21(3):601–620; doi: 10.1111/faf.12449
4.
ZhangL, WangH, GessnerJ, et al.To save sturgeons, we need river channels around hydropower dams. Proc Natl Acad Sci U S A, 2023; 120(13):e2217386120; doi: 10.1073/pnas.2217386120
5.
ChenM, XuX, WuX, et al.Centennial-scale study on the spatial-temporal evolution of riparian wetlands in the Yangtze River of China. Int J Appl Earth Obs, 2022; 113:102874; doi: 10.1016/j.jag.2022.102874
6.
FanX, WeiQ, ChangJ, et al.A review on conservation issues in the upper Yangtze River–a last chance for a big challenge: Can Chinese paddlefish (Psephurus gladius), Dabry′ s sturgeon, (Acipenser dabryanus) and other fish species still be saved? J Appl Ichthyol, 2006; 22(s1):32–39; doi: 10.1111/j.1439-0426.2007.00926.x
7.
LiuY, BlackburnH, TaylorSS, et al.Development of germplasm repositories to assist conservation of endangered fishes: Examples from small-bodied livebearing fishes. Theriogenology, 2019; 135(1):138–151; doi: 10.1016/j.theriogenology.2019.05.020
8.
ChenY, WuX, LiuX, et al.Biochemical, transcriptomic and metabolomic responses to total dissolved gas supersaturation and their underlying molecular mechanisms in Yangtze sturgeon (Acipenser dabryanus). Environ Res, 2023; 216(Pt 1):114457; doi: 10.1016/j.envres.2022.114457
9.
LiP, GuoW, QiaoXM, et al.Morphology and ultrastructure of Dabry’s sturgeon (Acipenser dabryanus) spermatozoa using scanning and transmission electron microscopy. J Appl Ichthyol, 2019; 35(1):192–201; doi: 10.1111/jai.13836
10.
XiaoK, HuangH, ShiX, et al.The effects of tachykinin1 gene products on prepubertal Dabry’s Sturgeon (Acipenser dabrynus) pituitary hormone secretion and gene expression. Animals (Basel), 2024; 14(2):227; doi: 10.3390/ani14020227
11.
YeH, TakeuchiY, DuH, et al.Spermatogonia from cryopreserved testes of critically endangered chinese sturgeon efficiently colonized and preferentially proliferated in the recipient gonads of Yangtze sturgeon. Mar Biotechnol (NY), 2022; 24(1):136–150; doi: 10.1007/s10126-022-10092-5
12.
LiuJ, LiuX, ZengQ, et al.Establishment and characterization of a cell line derived from fin of the endangered Yangtze sturgeon (Acipenser dabryanus). In Vitro Cell Dev Biol Anim, 2020; 56(8):650–658; doi: 10.1007/s11626-020-00488-9
13.
YeH, ZhouC, YueH, et al.Cryopreservation of germline stem cells in American paddlefish (Polyodon spathula). Anim Reprod Sci, 2021; 224:106667; doi: 10.1016/j.anireprosci.2020.106667
14.
JudyckaS, DietrichMA, NyncaJ, et al.Preservation of fish male germplasm in Poland. Front Mar Sci, 2024; 11:1407895; doi: 10.3389/fmars.2024.1407895
15.
Martínez-PáramoS, HorváthÁ, LabbéC, et al.Cryobanking of aquatic species. Aquaculture, 2017; 472(1):156–177; doi: 10.1016/j.aquaculture.2016.05.042
16.
Sampaio-SilvaF, MagalhaesT, CarvalhoF, et al.Profiling of RNA degradation for estimation of post morterm interval. PLoS One, 2013; 8(2):e56507; doi: 10.1371/journal.pone.0056507
17.
TozzoP, ScrivanoS, SanavioM, et al.The role of DNA degradation in the estimation of post-mortem interval: A systematic review of the current literature. Int J Mol Sci, 2020; 21(10):3540; doi: 10.3390/ijms21103540
18.
RichterF, PlehnJE, BesslerL, et al.RNA marker modifications reveal the necessity for rigorous preparation protocols to avoid artifacts in epitranscriptomic analysis. Nucleic Acids Res, 2022; 50(8):4201–4215; doi: 10.1093/nar/gkab1150
19.
ZhangW, DongK, SunJ, et al.The relationship between postmortem interval and protein changes in mice. J Forensic Leg Med, 2024; 101:102618; doi: 10.1016/j.jflm.2023.102618
20.
DeCosterMA, MaddiS, DuttaV, et al.Microscopy and image analysis of individual and group cell shape changes during apoptosis. In: Microscopy: Science, Technology, Applications and Education. (Méndez-VilasA, DíazJ. eds.) Formatex: Spain; 2010; pp. 836–843.
21.
WangCJ, ZhouZG, HolmqvistA, et al.Survivin expression quantified by Image Pro-Plus compared with visual assessment. Appl Immunohistochem Mol Morphol, 2009; 17(6):530–535; doi: 10.1097/PAI.0b013e3181a13bf2
22.
R Core Team. R: A language and environment for statistical computing. R Foundation for Statistical Computing: Vienna, Austria, 2018. http://www.R-project.org
23.
ComizzoliP, HoltWV. Breakthroughs and new horizons in reproductive biology of rare and endangered animal species. Biol Reprod, 2019; 101(3):514–525; doi: 10.1093/biolre/ioz031
24.
MarshSM, HoffmannM, BurgessND, et al.Prevalence of sustainable and unsustainable use of wild species inferred from the IUCN Red List of Threatened Species. Conserv Biol, 2022; 36(2):e13844; doi: 10.1111/cobi.13844
25.
ChapmanT, MilesS, TrivediC. Capturing, protecting and restoring plant diversity in the UK: RBG Kew and the Millennium Seed Bank. Plant Divers, 2019; 41(2):124–131; doi: 10.1016/j.pld.2018.06.001
26.
BrarDS, KhushGS. Wild relatives of rice: a valuable genetic resource for genomics and breeding research. The wild Oryza genomes. Springer: Cham, Switzerland; 2018; doi: 10.1007/978-3-319-71997-9_1(2018)
27.
LoiP, PtakG, BarboniB, et al.Genetic rescue of an endangered mammal by cross-species nuclear transfer using post-mortem somatic cells. Nat Biotechnol, 2001; 19(10):962–964; doi: 10.1038/nbt1001-962
28.
OhHJ, KimMK, JangG, et al.Cloning endangered gray wolves (Canis lupus) from somatic cells collected postmortem. Theriogenology, 2008; 70(4):638–647; doi: 10.1016/j.theriogenology.2008.04.032
29.
CortezJV, HardwickeK, GrupenCG, et al.Cloned foal born from postmortem-obtained ear sample refrigerated for 5 days before fibroblast isolation and decontamination of the infected monolayer culture. Cell Reprogram, 2024; 26(1):33–36; doi: 10.1089/cell.2023.0076
30.
MirzayansR, MurrayD. Do TUNEL and other apoptosis assays detect cell death in preclinical studies? Int J Mol Sci, 2020; 21(23):9090; doi: 10.3390/ijms21239090
31.
WynenH, TaylorE, HeylandA. Thyroid hormone-induced cell death in sea urchin metamorphic development. J Exp Biol, 2022; 225(23):jeb244560; doi: 10.1242/jeb.244560
32.
BehrmannMS, TrakselisMA. In vivo fluorescent TUNEL detection of single stranded DNA gaps and breaks induced by dnaB helicase mutants in Escherichia coli. Methods Enzymol, 2022; 672:125–142; doi: 10.1016/bs.mie.2022.02.021
33.
LooDT. In Situ Detection of Apoptosis by the TUNEL Assay: An Overview of Techniques. In: DNA Damage Detection In Situ, Ex Vivo, and In Vivo. Methods in Molecular Biology, vol 682. (DidenkoV. eds) Humana Press: Totowa, NJ; 2011.
34.
WijsmanJH, JonkerRR, KeijzerR, et al.A new method to detect apoptosis in paraffin sections: In situ end-labeling of fragmented DNA. J Histochem Cytochem, 1993; 41(1):7–12; doi: 10.1177/41.1.7678025
35.
PengX, CaiCY, YiSH, et al.Image analysis for degradation of DNA in retinal nuclei of rat after death. Chin J Forensic Med, 2009; 24(1):15–18; doi: 10.3969/j.issn.1001-5728.2009.01.005
36.
BryantBH, BoekelheideK. Time-dependent changes in post-mortem testis histopathology in the rat. Toxicol Pathol, 2007; 35(5):665–671; doi: 10.1080/01926230701459994
37.
MahmoudNA, HassanAA, Abdel RahimAH, et al.Molecular versus histopathological examination of the prostate gland in the estimation of post-mortem interval (an experimental study). Egypt J Forensic Sci Appli Toxicol, 2018; 18(1):35–54; doi: 10.1093/qjmed/hcy200.054
38.
LiWY, LiY, WengL, et al.Study on the properties of recombinant prolyl endopeptidase from abalone (Haliotis discus hannai) muscle. Sci Technol Food Industry, 2021; 42(7):129–135; doi: 10.13386/j.issn1002-0306.2020060196 (in Chinese with English abstract)
39.
GaoH, YangS, GaoJ, et al.An experimental study to estimate the early postmortem interval based on the degradation of lncRNAs in rat brain tissue. Sci Rep, 2024; 14(1):19586; doi: 10.1038/s41598-024-70678-y
40.
PittnerS, MonticelliFC, PfistererA, et al.Postmortem degradation of skeletal muscle proteins: A novel approach to determine the time since death. Int J Legal Med, 2016; 130(2):421–431; doi: 10.1007/s00414-015-1210-6
41.
FanW, DaiX, YeY, et al.Estimation of postmortem interval under different ambient temperatures based on multi-organ metabolomics and machine learning algorithm. J. Forensic. Leg. Med, 2025:1–15; doi: 10.1007/s00414-025-03523-0
42.
KrompecherT. Experimental evaluation of rigor mortis V. Effect of various temperatures on the evolution of rigor mortis. Forensic Sci Int, 1981; 17(1):19–26; doi: 10.1016/0379-0738(81)90184-5
43.
VarettoL, CurtoO. Long persistence of rigor mortis at constant low temperature. Forensic Sci Int, 2005; 147(1):31–34; doi: 10.1016/j.forsciint.2004.03.033
