The cAMP-PKA signaling pathway induces apoptosis and promotes hypothermic liver injury

doi:10.5847/wjem.j.1920-8642.2025.114

Abstract

Abstract:

BACKGROUND: The mechanisms underlying hypothermic liver injury necessitate investigation for the development of effective diagnostic and therapeutic approaches. We aim to establish a model of hypothermic liver injury to explore the hepatic alterations, thereby facilitating the prevention and treatment of the liver injury associated with hypothermia.

METHODS: The mice were placed in a −20 °C environment, to establish a hypothermic injury model. The liver function, metabolites, and proteins expression were measured by thromboelastography, histopathology, metabolomics and western blotting, respectively. Furthermore, apoptosis and pathway changes in the liver cells conducted with target metabolites were examined and verified.

RESULTS: According to the prolonged righting reflex recovery time and death occurrence, the mice with the anal temperature (AT) dropping to 20 °C or 15 °C were used to establish a model of hypothermia. The model mice showed changes in alanine aminotransferase (ALT), aspartate transaminase (AST), and coagulation indicators. HE staining results indicated that liver tissue in the AT 20°C mice had large hemorrhagic patches, while the AT 15°C mice displayed significant congestion, along with extensive infiltration of inflammatory cells around the central vein. Metabolomic and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of target metabolites revealed a significant increase in 3-hydroxybutyric acid and changes in the cyclic adenosine monophosphate (cAMP) signaling pathway in the liver tissue of hypothermic mice. The hypothermic mice showed decreases in levels of cAMP, protein kinase A C-α (PKA C-α), and phosphorylated BCL-2/BCL-XL-associated death promoter (p-Bad) and an increase in BCL-2/BCL-XL-associated death promoter (Bad) level in the liver. These protein changes and apoptosis were intensified by 3-hydroxybutyric acid in liver cells.

CONCLUSION: Hypothermia may induce apoptosis in the liver cell which may be related to the changes of the cAMP-PKA pathway proteins expression. These findings provide a basis for the treatment of hypothermic injury.

Key words: Hypothermia, Liver, Apoptosis, Metabolomics, 3-hydroxybutyric acid

Ying Liu, Feng Cheng, Zewei Wang, Du Juan, Zhixin Yuan, Xiaoye Tian, Ruihang Ma, Hongxu Jin. The cAMP-PKA signaling pathway induces apoptosis and promotes hypothermic liver injury[J]. World Journal of Emergency Medicine, 2025, 16(6): 543-551.

Figures/Tables 6

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References 31

1	Eglin CM, Wright J, Shepherd AI, Massey H, Hollis S, Towse J, et al. Plasma biomarkers of endothelial function, inflammation and oxidative stress in individuals with non-freezing cold injury. Exp Physiol. 2023; 108(3): 448-64. doi: 10.1113/EP090722 pmid: 36808666
2	Spencer MR, Hedegaard HB. QuickStats: death rates attributed to excessive cold or hypothermia among persons aged ≥15 years, by urban-rural status and age group - national vital statistics system, United States, 2019. MMWR Morb Mortal Wkly Rep. 2021; 70(7): 258. doi: 10.15585/mmwr.mm7007a6
3	van Veelen MJ, Brodmann Maeder M. Hypothermia in trauma. Int J Environ Res Public Health. 2021; 18(16): 8719. doi: 10.3390/ijerph18168719
4	Endorf FW, Nygaard RM. Social determinants of poor outcomes following frostbite injury: a study of the national inpatient sample. J Burn Care Res. 2021; 42(6): 1261-5. doi: 10.1093/jbcr/irab115 pmid: 34139760
5	Nasu T, Kainuma R, Ota H, Mizumura K, Taguchi T. Increased nociceptive behaviors and spinal c-Fos expression in the formalin test in a rat repeated cold stress model. Neurosci Res. 2024;198: 30-8.
6	Dong XC. Sirtuin 6-a key regulator of hepatic lipid metabolism and liver health. Cells. 2023; 12(4): 663. doi: 10.3390/cells12040663
7	Alabdulaali B, Al-Rashed F, Al-Onaizi M, Kandari A, Razafiarison J, Tonui D, et al. Macrophages and the development and progression of non-alcoholic fatty liver disease. Front Immunol. 2023;14: 1195699.
8	Teng T, Sun GD, Ding HW, Song X, Bai GD, Shi BM, et al. Characteristics of glucose and lipid metabolism and the interaction between gut microbiota and colonic mucosal immunity in pigs during cold exposure. J Anim Sci Biotechnol. 2023; 14(1): 84. doi: 10.1186/s40104-023-00886-5
9	Liu Y, Xue N, Zhang B, Lv H, Li S. Cold stress induced liver injury of mice through activated NLRP3/caspase-1/GSDMD pyroptosis signaling pathway. Biomolecules. 2022; 12(7): 927. doi: 10.3390/biom12070927
10	Hu LL, Liu JJ, Zhang WH, Wang TY, Zhang N, Lee YH, et al. Functional metabolomics decipher biochemical functions and associated mechanisms underlie small-molecule metabolism. Mass Spectrom Rev. 2020; 39(5-6): 417-33. doi: 10.1002/mas.21611 pmid: 31682024
11	Zhang ZH, Wang XY, Wang JC, Jia ZY, Liu YM, Xie X, et al. Metabonomics approach to assessing the metabolism variation and endoexogenous metabolic interaction of ginsenosides in cold stress rats. J Proteome Res. 2016; 15(6): 1842-52. doi: 10.1021/acs.jproteome.6b00015 pmid: 27150950
12	Qiu S, Cai Y, Yao H, Lin CS, Xie YQ, Tang SQ, et al. Small molecule metabolites: discovery of biomarkers and therapeutic targets. Signal Transduct Target Ther. 2023; 8(1): 132.
13	Okamatsu-Ogura Y, Kuroda M, Tsutsumi R, Tsubota A, Saito M, Kimura K, et al. UCP1-dependent and UCP1-independent metabolic changes induced by acute cold exposure in brown adipose tissue of mice. Metabolism. 2020;113: 154396.
14	Shen Y, Wu SD, Chen Y, Li XY, Zhu Q, Nakayama K, et al. Alterations in gut microbiome and metabolomics in chronic hepatitis B infection-associated liver disease and their impact on peripheral immune response. Gut Microbes. 2023; 15(1): 2155018. doi: 10.1080/19490976.2022.2155018
15	Hu JJ, Shen YL, Chen YW. Metabonomics application on screening serum biomarkers of golden hamsters with nonalcoholic steatohepatitis induced by high-fat diet. Comb Chem High Throughput Screen. 2023; 26(13): 2280-92. doi: 10.2174/1386207326666230223095745 pmid: 36815651
16	Savioli G, Ceresa IF, Bavestrello Piccini G, Gri N, Nardone A, Russa RL, et al. Hypothermia: beyond the narrative review-the point of view of emergency physicians and medico-legal considerations. J Pers Med. 2023; 13(12): 1690. doi: 10.3390/jpm13121690
17	Maule AL, Kotas KS, Scatliffe-Carrion KD, Ambrose JF. Cold weather injuries among the active and reserve components of the U.S. Armed Forces, July 2019-June 2024. MSMR. 2024; 31(11): 2-13. pmid: 39621826
18	Song XY, Zha Y, Liu J, He P, He LJ. Associations between liver function parameters and poor clinical outcomes in peritoneal dialysis patients. Ther Apher Dial. 2023; 27(1): 12-8. doi: 10.1111/tap.v27.1
19	Yalçın R, Kart A, Özmen Ö, Zeybek E. Protective effects of resveratrol against fumonisin B1-induced liver toxicity in mice. Arh Hig Rada Toksikol. 2023; 74(2): 115-9. doi: 10.2478/aiht-2023-74-3648 pmid: 37357877
20	Zaki MSA, Abadi AM, El-Kott AF, Mohamed G, Alrashdi BM, Eid RA, et al. Protective efficacy of luteolin against aflatoxinB1-induced toxicity, oxidative damage, and apoptosis in the rat liver. Environ Sci Pollut Res Int. 2023; 30(18): 52358-68. doi: 10.1007/s11356-023-26085-3
21	Wang XL, He Y, Mackowiak B, Gao B. MicroRNAs as regulators, biomarkers and therapeutic targets in liver diseases. Gut. 2021; 70(4): 784-95. doi: 10.1136/gutjnl-2020-322526 pmid: 33127832
22	Liu Y, Xu B, Hu YJ, Liu P, Lian S, Lv HM, et al. O-GlcNAc/Akt pathway regulates glucose metabolism and reduces apoptosis in liver of piglets with acute cold stress. Cryobiology. 2021;100: 125-32.
23	Yao RZ, Yang YY, Lian S, Shi HZ, Liu P, Liu Y, et al. Effects of acute cold stress on liver O-GlcNAcylation and glycometabolism in mice. Int J Mol Sci. 2018; 19(9): 2815. doi: 10.3390/ijms19092815
24	Pernes G, Morgan PK, Huynh K, Mellett NA, Meikle PJ, Murphy AJ, et al. Characterization of the circulating and tissue-specific alterations to the lipidome in response to moderate and major cold stress in mice. Am J Physiol Regul Integr Comp Physiol. 2021; 320(2): R95-R104.
25	Chen Y, Ji H, Guo JR, Chen Y, Li WJ, Wang SP, et al. Non-targeted metabolomics analysis based on LC-MS to assess the effects of different cold exposure times on piglets. Front Physiol. 2022;13: 853995.
26	Sun SX, Cao XJ, Gao J. C24: 0 avoids cold exposure-induced oxidative stress and fatty acid β-oxidation damage. iScience. 2021; 24(12): 103409. doi: 10.1016/j.isci.2021.103409
27	Zhang YD, Li ZH, Liu XY, Chen XY, Zhang SJ, Chen YM, et al. 3-hydroxybutyrate ameliorates insulin resistance by inhibiting PPARγ Ser 273 phosphorylation in type 2 diabetic mice. Signal Transduct Target Ther. 2023;8: 190.
28	Aslam M, Ladilov Y. Emerging role of cAMP/AMPK signaling. Cells. 2022; 11(2): 308. doi: 10.3390/cells11020308
29	Ma CL, Li L, Yang GM, Zhang ZB, Zhao YN, Zeng XF, et al. Neuroprotective effect of gastrodin in methamphetamine-induced apoptosis through regulating cAMP/PKA/CREB pathway in cortical neuron. Hum Exp Toxicol. 2020; 39(8): 1118-29. doi: 10.1177/0960327120911438 pmid: 32162539
30	Yu LT, Liu SX, Zhou RX, Sun H, Su XJ, Liu Q, et al. Atorvastatin inhibits neuronal apoptosis via activating cAMP/PKA/p-CREB/BDNF pathway in hypoxic-ischemic neonatal rats. FASEB J. 2022; 36(4): e22263. doi: 10.1096/fsb2.v36.4
31	Xia Y, He F, Moukeila Yacouba MB, Zhou HM, Li JF, Xiong Y, et al. Adenosine A2a receptor regulates autophagy flux and apoptosis to alleviate ischemia-reperfusion injury via the cAMP/PKA signaling pathway. Front Cardiovasc Med. 2022;9: 755619.

Variables	Control (n=9)	AT 20 °C (n=45)	AT 15 °C (n=45)	AT 13 °C (n=9)
-20 °C exposure duration (min)	0	79.17±2.32	84.17±2.48	85.50±2.59
Righting reflex recovery time (s)	0.23±0.06	0.70±0.07*	disappear	disappear
Death rate	0	0	0	56% (5/9)

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[8]	Li-wen Du, Bao-qing Xu, Kai Xun, Fang-qi Zhang. Glutamine supplementation attenuates intestinal apoptosis by inducing heat shock protein 70 in heatstroke rats [J]. World Journal of Emergency Medicine, 2023, 14(1): 37-43.
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[10]	Cheng Hang, Li-jun Liu, Zhao-yun Huang, Jian-liang Zhu, Bao-chun Zhou, Xiao-zhen Li. Optimal indicator for changing the filter during the continuous renal replacement therapy in intensive care unit patients with acute kidney injury: A crossover randomized trial [J]. World Journal of Emergency Medicine, 2022, 13(3): 196-201.
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