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In This Article

  • Summary
  • Abstract
  • Introduction
  • Protocol
  • Results
  • Discussion
  • Disclosures
  • Acknowledgements
  • Materials
  • References
  • Reprints and Permissions

Summary

The present study identified a unique mechanism by which salidroside exerts mitochondrial protective effects on hypoxic HT22 cells, partly through the AMPK/Sirt1/HIF-1α pathway.

Abstract

Salidroside (Sal), an active ingredient of Rhodiola crenulata (Hook. f. et Thoms.) H. Ohba has been found to exert mitochondrial protective effects by improving metabolism and enhancing the energy supply of brain cells under hypoxic conditions. However, its mechanism of action has not been fully clarified. In the present study, high-performance liquid chromatography was first employed to analyze the effects of Sal on nucleotide (ATP, ADP, and AMP) levels. The cellular thermal shift assay (CETSA), a widely used molecular interaction method for validating and quantifying drug target engagement in cells and tissues across different species, was then chosen to confirm the affinity of Sal for AMPK/Sirt1/HIF-1α pathway-related proteins. The results revealed that Sal increased ATP and ADP levels in hypoxic HT22 cells while reducing AMP levels. Moreover, Sal exhibited stable binding to AMPKα, p-AMPKα, Sirt1, and HIF-1α proteins. In conclusion, Sal may exert mitochondrial protective effects by modulating the AMPK/Sirt1/HIF-1α pathway to regulate nucleotide content. This study provides a methodological reference for nucleotide content analysis in cell samples and contributes to the identification and discovery of targets for compounds derived from traditional Chinese medicine.

Introduction

The brain is highly sensitive to oxygen due to its high metabolic demands, limited glycolytic capacity, and dependence on oxidative phosphorylation. As a result, exposure to a low-oxygen environment at high altitudes can readily lead to hypobaric hypoxic brain injury (HHBI)1,2. Epidemiological studies indicate that when individuals unacclimated to high altitudes ascend rapidly to high-altitude regions, the incidence of acute mountain sickness can reach up to 75%, with a fatality rate of approximately 1% for severe cases. Furthermore, in the absence of medical care, mortality rates for high-altitude cerebral or pulmonary edema can be as high as 40%3,4.

HHBI presents with a broad spectrum of clinical symptoms. Mild to moderate cases may include headaches, dizziness, and memory loss5, while severe cases can result in cognitive impairment, altered consciousness, and potentially fatal outcomes5. The prevention and treatment of HHBI in high-altitude regions have become a key focus of medical research. Preventative strategies primarily involve adaptive training in high-altitude environments, including adequate rest, a well-balanced diet, proper nutrition, and appropriate physical exercise6,7. Additionally, pharmacological interventions aimed at protecting brain cells and alleviating cerebral hypoxia remain central to current HHBI research.

Mitochondria serve as the primary energy production centers within cells, synthesizing adenosine triphosphate (ATP) to meet cellular energy demands. Under hypoxic conditions, mitochondrial energy production declines, leading to reduced ATP levels and impaired cell function8. The hypoxic injury also disrupts mitochondrial regulation of Ca2+ and pH homeostasis, triggering apoptosis and necrosis9,10. There is a mutually reinforcing relationship between mitochondrial dysfunction and hypoxic brain injury. On the one hand, hypoxia-induced mitochondrial impairment exacerbates oxygen deficiency by further reducing cellular energy metabolism, creating a vicious cycle. On the other hand, mitochondrial dysfunction elevates intracellular Ca2+ levels, activating apoptotic cascades and leading to cell death11. Although the mechanisms underlying hypoxic brain injury remain complex and not fully understood, multiple studies have identified impaired neuronal mitochondrial energy metabolism as a critical factor in its pathogenesis12,13. Therefore, further exploration of mitochondrial function may provide valuable insights into potential therapeutic targets for hypoxic brain injury.

Salidroside (Sal) is an active ingredient extracted from the plateau plant Rhodiola crenulata (Hook. f. et Thoms.) H. Ohba and is widely used in food health products and pharmaceuticals14. The molecular formula of Sal is C14H20O7, and it is also known as 2-(4-hydroxyphenyl)-ethyl β-D-glucopyranoside. It possesses diverse pharmacological properties, including anti-hypoxia, antioxidant, anti-fatigue, anti-tumor, immunomodulatory, anti-inflammatory, and cardiovascular and cerebrovascular protective effects15,16,17. Among these, its anti-hypoxia effect is one of the most well-documented. Recent studies have increasingly highlighted the significant mitochondrial protective effects of Sal as a potential mechanism for its preventive and therapeutic effects on plateau-induced brain injury in mice14,18. However, the precise molecular mechanisms by which Sal influences ATP, ADP, and AMP levels remain poorly understood.

AMP-activated protein kinase (AMPK) acts as a key energy sensor that helps maintain cellular energy homeostasis. Activation of AMPK stimulates Sirtuin 1 (Sirt1), leading to an increase in intracellular NAD+ levels19. Studies have shown that Sirt1 can regulate hypoxia-inducible factor 1-alpha (HIF-1α) to coordinate the cellular response to hypoxia20. Previous research has demonstrated that Sal inhibits the opening of the neuronal mitochondrial permeability transition pore, regulates HIF-1α-mediated mitochondrial energy processes, attenuates neuronal apoptosis, and maintains blood-brain barrier integrity, thereby protecting rats from plateau-induced brain injury14,21. However, the effect of Sal on ATP and its metabolites, ADP and AMP, remains uncertain.

To investigate this, high-performance liquid chromatography (HPLC) was first employed to quantify the levels of these three nucleotides. Additionally, the cellular thermal shift assay (CETSA), a widely used biophysical technique introduced in 2013 to study ligand-protein interactions in intact cells22, was utilized. This method is commonly applied to validate and quantify drug-target engagement in cells and tissues across different species. Specifically, after co-incubating target cell lysates with the drug at varying temperatures for a set duration, the drug-bound protein exhibits increased thermal stability, making it less prone to denaturation and precipitation. The precipitated unbound proteins are then removed via centrifugation, and drug-target interactions are subsequently identified through western blot analysis of the supernatant22. To identify potential molecular targets of Sal, CETSA was selected to assess its binding affinity with AMPK/Sirt1/HIF-1α pathway-related proteins.

Protocol

The commercial details of the reagents and the equipment used in this study are provided in the Table of Materials.

1. Solution preparation

  1. Prepare complete Dulbecco's modified Eagle medium (DMEM) by adding 10% fetal bovine serum and 1% penicillin-streptomycin.
  2. Prepare a 20 mM Sal solution by dissolving 3 mg of Sal in 500 µL of phosphate-buffered saline (PBS)21.
  3. Filter 500 mL of chromatographically pure acetonitrile (mobile phase A) using a 0.45 µm organic membrane.
  4. Prepare 1000 mL of mobile phase B by adding 3.5 mL of reagent I to 1 L of deionized water. Adjust the pH to 6.15 with reagent II, following the instructions of the nucleotide (ATP, ADP, and AMP) content assay kit (see Table of Materials).
    NOTE: Reagent I and reagent II are included in the nucleotide (ATP, ADP, and AMP) content assay kit.
  5. Filter mobile phase B using a 0.22 µm aqueous membrane.
    NOTE: Mobile phase B should be used immediately after preparation. Mobile phases A and B should be sonicated for 30 min before use to remove air bubbles.
  6. Dilute a 1 µM ATP, ADP, and AMP standard stock solution with deionized water to obtain 0.5 µM, 0.1 µM, 0.05 µM, 0.01 µM, and 0.005 µM ATP, ADP, and AMP series standard solutions.
  7. Filter the series of standard solutions using a 0.22 µm needle-type microporous filter membrane.
    NOTE: Brown injection bottles should be used to store the serial standard solutions. The prepared series of standard solutions should be used as soon as possible.

2. Cell culture

NOTE: HT22 cell culture and the CoCl2-stimulated hypoxia model were established according to a previous report21.

  1. Culture HT22 cells at 37 °C with 5% CO2 until 80% confluence is reached in the Petri dish. Digest the cells with 0.25% trypsin for 1 min.
  2. Seed 2 × 105 cells from step 2.1 into 6-well plates with complete DMEM (control group with no drug treatment) or complete DMEM containing the drug (250 µM CoCl2, 250 µM CoCl2 + 10 µM Dor, 250 µM CoCl2 + 20 µM Sal, and 250 µM CoCl2 + 10 µM Dor + 20 µM Sal). Incubate for 24 h at 37 °C with 5% CO2.
    NOTE: Three parallel samples were set up in each group.

3. Nucleotide (ATP, ADP, and AMP) content assay

  1. Add 1 mL of extraction reagent I to each well in the 6-well plates from step 2.2.
    NOTE: Extraction reagent I is included in the nucleotide (ATP, ADP, and AMP) content assay kit (see Table of Materials).
  2. Lyse the cells by sonicating on an ice bath using an ultrasonic cell disruption apparatus (power: 300 W, ultrasonic waves for 3 s, 7 s interval, total time: 3 min).
  3. Centrifuge the lysed cells at 10,304 × g for 10 min at 4 °C using a high-speed refrigerated centrifuge.
  4. Transfer 0.75 mL of supernatant from step 3.3 into a 2.0 mL tube. Add 0.75 mL of extraction reagent II, shake and mix using a vortex mixer, and centrifuge again at 10,304 × g for 10 min at 4 °C.
    NOTE: Extraction reagent II is included in the nucleotide (ATP, ADP, and AMP) content assay kit.
  5. Pipette the supernatant from step 3.4 and filter it using a 0.22 µm needle-type microporous filter membrane into a brown vial.
    NOTE: ATP, ADP, and AMP in the extracted sample are not stable. All experimental procedures should be performed at low temperatures or on ice. Samples should be processed and tested as soon as possible.
  6. Open the chromatography software and set the following parameters: injection volume: 10 µL; column temperature: 27 °C; flow rate: 0.8 mL/min; detection wavelength: 254 nm; detection duration: 70 min. Set the gradient elution procedure according to Table 1.
  7. Click the execute button to automatically inject the sample and detect the content of ATP, ADP, and AMP in serial standard solutions (step 1.7) and sample solutions (step 3.5) according to the program set in step 3.6.
    NOTE: At the end of the experiment, wash the chromatographic column with 98% mobile phase B to prevent clogging.
  8. Plot the standard curves of ATP, ADP, and AMP using graphing and statistical analysis software, with serial standard concentrations on the x-axis and peak areas on the y-axis.
  9. Calculate the ATP, ADP, and AMP content in the samples using the standard curve equation from step 3.8.

4. Cellular thermal shift assay (CETSA)

  1. Lyse normal HT22 cells for 20 min in a Petri dish using protease inhibitor-containing lysate (lysate: protease inhibitor = 100:1) by repeatedly pipetting the cells.
    NOTE: Perform this step on an ice bath to prevent protein degradation.
  2. Collect the cell lysate from step 4.1 and sonicate on an ice bath using an ultrasonic cell disruption apparatus (power: 300 W, ultrasonic waves for 3 s, 7 s interval, total time: 3 min). Centrifuge at 10,304 × g, 4 °C for 20 min and collect the supernatant.
    NOTE: Divide the supernatant into 14 portions: 7 groups for coincubation with Sal and 7 groups for coincubation with PBS as the control. For the Sal-treated group, add 100 µL of supernatant and 0.1 µL of 20 µM Sal per sample. For the control group, add 100 µL of supernatant and 0.1 µL of PBS per sample.
  3. Incubate each sample at room temperature for 30 min, then further incubate at 37 °C, 42 °C, 47 °C, 52 °C, 57 °C, 62 °C, and 67 °C for 3 min using a metal heating temperature control instrument.
  4. Centrifuge the samples from step 4.3 at 10,304 × g for 20 min at 4 °C and collect the supernatant using a pipette.
  5. Measure the total protein concentration of the supernatant from step 4.4 using a BCA protein concentration assay kit, following the manufacturer's instructions (see Table of Materials).
  6. Prepare 10% separation gel using a PAGE gel rapid preparation kit, following the manufacturer's instructions (see Table of Materials).
  7. Load 3 µL of pre-stained color protein marker and 14 µL of samples into the wells of the separation gel (final protein amount per well: 20 µg).
  8. Run the gel using electrophoresis buffer for 1 h at 80-100 mV to obtain separated proteins.
  9. Transfer the proteins from step 4.8 onto a PVDF membrane using rapid membrane transfer solution and a sandwich structure (cotton-filter paper-PVDF membrane-protein gel-filter paper-cotton) at 400 mA for 30 min .
  10. Block the PVDF membranes from step 4.9 with 5% bovine serum albumin (BSA) solution for 1-2 h at room temperature.
    NOTE: Prepare 5% BSA solution by dissolving 5 mg of BSA powder in 100 mL of TBST solution containing 100 mL of TBS buffer and 0.05% Tween 20 .
  11. Incubate the PVDF membranes from step 4.10 with 5 mL diluted primary antibody at 4 °C overnight.
    NOTE: Dilute the primary antibody in BSA solution (1:1000 ratio).
  12. Wash the PVDF membranes three times with TBST for 5 min each to remove unbound primary antibodies.
  13. Incubate the membranes from step 4.12 with diluted horseradish peroxidase (HRP)-coupled secondary antibody for 2 h at room temperature on a decolorization shaker.
    NOTE: Dilute the HRP-coupled secondary antibody in BSA solution (1:10,000 ratio) .
  14. Wash the membranes three times with TBST for 5 min each to remove residual secondary antibody.
  15. Cover the membranes from step 4.14 with ECL chemiluminescence developer, then image the target protein signal using an imaging system.
    NOTE: Perform this step in the dark to prevent fluorescence quenching.
  16. Analyze and quantify the gray value of target proteins using ImageJ software (see T able of Materials).

Results

The standard curves for ATP, ADP, and AMP detected by HPLC were Y = 7006.5X - 222.99, Y = 5217.3X - 17.796, and Y = 9280.1X + 22.749, respectively (Figure 1A-C). The nucleotide contents measured in each group by HPLC were calculated using the standard curves (Figure 1D-I). It was found that CoCl2 significantly reduced ATP and ADP levels in HT22 cells compared to the control group (

Discussion

Mitochondria are key organelles involved in the therapeutic prevention of HHBI23,24,25. Previous studies by the group have confirmed that Sal regulates AMPK, Sirt1, and HIF-1α protein expression, enhancing neuronal mitochondrial function and protecting against HHBI21,24. However, the direct effect of Sal on nucleotides in hypoxic cells requires further investigation...

Disclosures

The authors have nothing to disclose.

Acknowledgements

This work was supported by the National Natural Science Foundation of China (82274207 and 82474185), the Science & Technology Department of Sichuan Province (2024NSFSC1845), the Science Foundation for Youths of Science & Technology Department of Sichuan Province (2023NSFSC1776), the Key Research and Development Program of Ningxia (2023BEG02012), Youth Talent Support Project of the China Association of Chinese Medicine for 2024-2026 (2024-QNRC2-B07) and the Xinglin Scholar Research Promotion Project of Chengdu University of TCM (XKTD2022013 and QJJJ2024027).

AUTHOR CONTRIBUTION:
Xiaobo Wang, Yating Zhang, Ya Hou, Rui Li and Xianli Meng conceived this project. Yating Zhang, Ya Hou, and Tingting Kuang performed the experiments and analyzed the data. Yating Zhang and Hong Jiang wrote the manuscript. Xiaobo Wang and Xianli Meng revised the manuscript. All of the authors have read and approved the final manuscript.

Materials

NameCompanyCatalog NumberComments
AcetonitrileAladdinA104440
0.22 µm aqueous membraneJintengJTMF0445
0.22 µm needle type microporous filter membraneJintengJTSFM013001
0.45 µm organic membraneJintengJTMF0448
Agilent OpenLab software  AgilentVersion 2.X
Antibody-AMPKαCell Signaling Technology#2532
Antibody-HIF-1αCell Signaling Technology#41560
Antibody-p-AMPKαCell Signaling Technology#50081
Antibody-Sirt1Cell Signaling Technology#2028
Antibody-β-actinCell Signaling Technology#4970
BCA protein concentration assay kitBoster Biological Technology17E17B46
Bovine serum albuminCywin Innovation (Beijing) Biotechnology Co., Ltd.SW127-02
Broad-spectrum phosphatase inhibitor (100×)Boster Biological TechnologyAR1183
Chromatographic columnAgilentSB-C18
CoCl2Sigma15862
Decolorization shakerKylin-BellTS-2
DorsomorphinMedChemExpress (MCE)HY-13418A
Dulbecco's modified eagle mediumGibco8121587
Electrophoresis bufferNCM Biotech20230801
Fetal bovine serumGibco2166090RP
Goat Anti-rabbit IgG H&L (TRITC)ZenBioScience Co., Ltd.511202
GraphPad Prism softwareGraphPad software, LLCVersion 9.0.0
High performance liquid chromatographyAgilent1260 Infinity II Prime 
High speed refrigerated centrifugeThermo Fisher ScientificLegend Micro 17R
HRP conjugated affinipure goat anti-rabbit IgG(H+L)Boster Biological Technology Co., Ltd.BA1054
HT22 cellsGuangzhou Jennio Biotech Co., Ltd.JNO-02001
Hypersensitive ECL chemiluminescence kitNCM BiotechP10300
Image J softwareNational Institutes of Healthv1.8.0
Metal heating temperature control instrumentBaiwan Electronictechnology Co., Ltd.HG221-X3
MethanolAladdinM116118
Nucleotide (ATP, ADP, and AMP) content assay kitBeijing Solarbio Science & Technology Co., Ltd.BC5114
PAGE gel rapid preparation kitBiosharpPL566B-5
Penicillin-streptomycinNCM BiotechC125C5
Phosphate buffered saline (1×)Gibco8120485
Pre-stained color protein marker (10-180 kDa)Cywin Innovation (Beijing) Biotechnology Co., Ltd.SW176-02
Protein loading buffer (5x)Boster Biological TechnologyAR1112
PVDF (0.45 μm)Cywin Innovation (Beijing) Biotechnology Co., Ltd.SW120-01
Rapid membrane transfer solutionCywin Innovation (Beijing) Biotechnology Co., Ltd.SW171-02
RIPA lysateBoster Biological Technology Co., Ltd.AR0105
SalidrosideChengdu Herbpurify Co., Ltd.RFS-H0400191102
TBS bufferNCM Biotech23HA0102
Transmembrane bufferNCM Biotech23CA2000
Trypsin (0.25%, 1×)HyCloneJ210045
Tween 20Shanghai Canspec Scientific Instruments Co., Ltd.PM12012
Ultrasonic cell disruption apparatusNingbo Xinyi ultrasonic equipment Co., Ltd.JY92-IIDN
Visionworks imaging systemAnalytik JenaUVP ChemStudio
Vortex mixerKylin-BellXW-80A

References

  1. Marutani, E., et al. Ichinose, F. Sulfide catabolism ameliorates hypoxic brain injury. Nat Commun. 12 (1), 3108 (2021).
  2. Chen, X., Ma, W., Li, Y. Current situation of Chinese and Western medicine research on hypoxic brain injury at high altitude. Jilin Medical J. 43 (11), 3099-3101 (2022).
  3. Huo, Y., Zhao, A., Li, X., Li, J., Wang, R. Animal models of acute plateau disease. Chinese Pharmacol Bull. 37 (01), 26-30 (2021).
  4. Netzer, N., Strohl, K., Faulhaber, M., Gatterer, H., Burtscher, M. Hypoxia-related altitude illnesses. J Travel Med. 20 (4), 247-255 (2013).
  5. Lefferts, W. K. et al. Effect of hypoxia on cerebrovascular and cognitive function during moderate intensity exercise. Physiol Behav. 165, 108-118 (2016).
  6. Burtscher, J., Mallet, R. T., Burtscher, M., Millet, G. P. Conditioning the brain: From exercise to hypoxia. Exerc Sport Sci Rev. 49 (4), 291-292 (2021).
  7. Koester-Hegmann, C. et al. High-altitude cognitive impairment is prevented by enriched environment including exercise via VEGF signaling. Front Cell Neurosci. 12, 532 (2019).
  8. Coimbra-Costa, D., Alva, N., Duran, M., Carbonell, T., Rama, R. Oxidative stress and apoptosis after acute respiratory hypoxia and reoxygenation in rat brain. Redox Biol. 12, 216-225 (2017).
  9. Nair, S. et al. Neuroprotection offered by mesenchymal stem cells in perinatal brain injury: Role of mitochondria, inflammation, and reactive oxygen species. J Neurochem. 158 (1), 59-73 (2021).
  10. Rodríguez, M., Valez, V., Cimarra, C., Blasina, F., Radi, R. Hypoxic-Ischemic encephalopathy and mitochondrial dysfunction: facts, unknowns, and challenges. Antioxid Redox Sign. 33 (4), 247-262 (2020).
  11. Li, T. et al. Overexpression of apoptosis inducing factor aggravates hypoxic-ischemic brain injury in neonatal mice. Cell Death Dis. 11 (1), 77 (2020).
  12. March-Diaz, R. et al. Hypoxia compromises the mitochondrial metabolism of Alzheimer's disease microglia via HIF1. Nature Aging. 1 (4), 385-399 (2021).
  13. Aabdien, A., Mallard, C., Hagberg, H. Mitochondrial dynamics, mitophagy and biogenesis in neonatal hypoxic-ischaemic brain injury. FEBS Lett. 592 (5), 812-830 (2018).
  14. Bai, X. L., Deng, X. L., Wu, G. J., Li, W. J., Jin, S. Rhodiola and salidroside in the treatment of metabolic disorders. Mini-Rev Med Chem. 19 (19), 1611-1626 (2019).
  15. Ji, R. et al. Salidroside alleviates oxidative stress and apoptosis via AMPK/Nrf2 pathway in DHT-induced human granulosa cell line KGN. Arch Biochem Biophys. 715, 109094 (2022).
  16. You, L., Zhang, D., Geng, H., Sun, F., Lei, M. Salidroside protects endothelial cells against LPS-induced inflammatory injury by inhibiting NLRP3 and enhancing autophagy. BMC Complement Med. 21 (1), 146 (2021).
  17. Yang, S. X., et al. Salidroside alleviates cognitive impairment by inhibiting ferroptosis via activation of the Nrf2/GPX4 axis in SAMP8 mice. Phytomedicine. 114, 154762 (2023).
  18. Hou, Y. et al. Salidroside intensifies mitochondrial function of CoCl2-damaged HT22 cells by stimulating PI3K-AKT-MAPK signaling pathway. Phytomedicine. 109, 154568 (2023).
  19. Yang, H., et al. Black soybean peptide mediates the AMPK/SIRT1/NF-κB signaling pathway to alleviate Alzheimer's-related neuroinflammation in lead-exposed HT22 cells. Int J Biol Macromol. 286, 138404 (2025).
  20. Yoon, H., Shin, S. H., Shin, D. H., Chun, Y. S., Park, J. W. Differential roles of Sirt1 in HIF-1α and HIF-2α mediated hypoxic responses. Biochem Bioph Res Co. 444 (1), 36-43 (2014).
  21. Hou, Y. et al. Rhodiola crenulata alleviates hypobaric hypoxia-induced brain injury by maintaining BBB integrity and balancing energy metabolism dysfunction. Phytomedicine. 128, 155529 (2024).
  22. Jafari, R. et al. The cellular thermal shift assay for evaluating drug target interactions in cells. Nat Protoc. 9 (9), 2100-2122 (2014).
  23. Tang, Y. et al. Salidroside attenuates CoCl2-simulated hypoxia injury in PC12 cells partly by mitochondrial protection. Eur J Pharmacol. 912, 174617 (2021).
  24. Wang, X. et al. Salidroside, a phenyl ethanol glycoside from Rhodiola crenulata, orchestrates hypoxic mitochondrial dynamics homeostasis by stimulating Sirt1/p53/Drp1 signaling. J Ethnopharmacol. 293, 115278 (2022).
  25. Kristián, T. Metabolic stages, mitochondria and calcium in hypoxic/ischemic brain damage. Cell Calcium. 36 (3-4), 221-233 (2004).
  26. Carling, D. AMPK signalling in health and disease. Curr Opin Cell Biol. 45, 31-37 (2017).
  27. Steinberg, G. R., Hardie, D. G. New insights into activation and function of the AMPK. Nat Rev. Mol Cell Bio. 24 (4), 255-272 (2023).
  28. Zhang, N. et al. Restoration of energy homeostasis under oxidative stress: Duo synergistic AMPK pathways regulating arginine kinases. PLoS Genet. 19 (8), e1010843 (2023).
  29. Wang, Y. et al. Irisin ameliorates neuroinflammation and neuronal apoptosis through integrin αVβ5/AMPK signaling pathway after intracerebral hemorrhage in mice. J Neuroinflamm. 19 (1), 82 (2022).
  30. Liu, H., Li, Y., Xiong, J. The role of hypoxia-inducible factor-1 alpha in renal disease. Molecules. 27 (21), 7318 (2022).
  31. Wu, H. et al. Mitochondrial dysfunction promotes the transition of precursor to terminally exhausted T cells through HIF-1α-mediated glycolytic reprogramming. Nat Commun. 14 (1), 6858 (2023).
  32. Joo, H. Y. et al. NADH elevation during chronic hypoxia leads to VHL-mediated HIF-1α degradation via SIRT1 inhibition. Cell Biosci. 13 (1), 182 (2023).
  33. Ham, P. B., Raju, R. Mitochondrial function in hypoxic ischemic injury and influence of aging. Prog Neurobiol. 157, 92-116 (2017).
  34. Dal Cortivo, G., Barracchia, C. G., Marino, V., D'Onofrio, M., Dell'Orco, D. Alterations in calmodulin-cardiac ryanodine receptor molecular recognition in congenital arrhythmias. Cell Mol Life Sci. 79 (2), 127 (2022).
  35. Wang, Z. Q. et al. Cyclovirobuxine D inhibits triple-negative breast cancer via YAP/TAZ suppression and activation of the FOXO3a/PINK1-Parkin pathway-induced mitophagy. Phytomedicine. 136, 156287 (2024).

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