目的:探究在低氧条件下乳酸对小鼠海马HT22细胞的神经保护作用。方法:将HT22细胞分成4组:对照组、乳酸组(LA组)、低氧组(Hypoxia组)和低氧加乳酸组(Hypoxia+LA组),通过CCK-8检测低氧条件下不同浓度乳酸对细胞活力的影响,观察细胞形态的变化;采用Western blot检测由PARK7(Parkinson's disease-associated protein 7, PARK7)基因编码的DJ-1的表达,并通过细胞免疫荧光法检测DJ-1分布和表达变化。结果:与低氧组相比,1∶104的乳酸的添加明显提高了细胞活力,具有统计学意义(P＜0.01);与对照组相比,低氧后DJ-1蛋白在细胞内分布与表达减少,具有统计学意义(P＜0.01);与低氧组相比,在低氧前加入乳酸增加了细胞中DJ-1的表达与分布,具有统计学意义(P＜0.05)。结论:低氧前乳酸加入可能通过激活DJ-1,维持细胞活性,缓解低氧损伤,从而发挥神经保护作用。
Objective: To investigate the neuroprotective effect of lactic acid on mouse hippocampal HT22 cells under hypoxic conditions. Methods: HT22 cells were divided into four groups: the control group, lactate group (LA group), hypoxia group and hypoxia + lactic acid group (Hypoxia + LA group), and CCK-8 was used to detect the effects of different concentrations of lactate on cell viability under hypoxic conditions, and the effects of morphological changes were observed. The expression of DJ-1 (Parkinson's disease-associated protein 7) encoded by PARK7 gene was detected by Western blot, and changes in DJ-1 distribution and expression were detected by cellular immunofluorescence. Results: Compared with the hypoxia group, the addition of 1∶104 lactic acid significantly increased cell viability, which was statistically significant (P＜0.01). Compared with the normal control group, the intracellular distribution and expression of DJ-1 protein decreased under hypoxia condition, which was statistically significant (P＜0.01). The addition of lactic acid before hypoxia increased the expression and distribution of DJ-1 in the cells compared with the hypoxia group, which was statistically significant (P＜0.05). Conclusion: Pre-hypoxic lactate incorporation may exert neuroprotective effects by activating DJ-1, maintaining cellular activity, and mitigating hypoxic injury.
[1] Thomas LW, Ashcroft M. Exploring the molecular interface between hypoxia-inducible factor signalling and mitochondria[J]. Cell Mol Life Sci, 2019, 76(9): 1759-1777.
[2] Fan YT, Li JS, Fang B. A tale of two:when neural stem cells encounter hypoxia[J]. Cell Mol Neurobiol, 2023, 43(5): 1799-1816.
[3] Liu J, Gu YK, Guo MY, et al. Neuroprotective effects and mechanisms of ischemic/hypoxic preconditioning on neurological diseases[J]. CNS Neurosci Ther, 2021, 27(8): 869-882.
[4] Magistretti PJ, Allaman I. A cellular perspective on brain energy metabolism and functional imaging[J]. Neuron, 2015, 86(4): 883-901.
[5] Brooks GA. Cell-cell and intracellular lactate shuttles[J]. J Physiol, 2009, 587(Pt 23): 5591-5600.
[6] Magistretti PJ, Allaman I. Lactate in the brain: from metabolic end-product to signalling molecule[J]. Nat Rev. Neurosci, 2018, 19(4): 235-249.
[7] van Hall G, Strømstad M, Rasmussen P, et al. Blood lactate is an important energy source for the human brain[J]. J Cereb Blood Flow Metab, 2009, 29(6): 1121-1129.
[8] 倪晓晨, 于世龙, 刘延庆, 等. DJ-1调控线粒体功能研究进展[J]. 中国药理学通报, 2023, 39(8): 1406-1411.
[9] Pellerin L, Magistretti PJ. Glutamate uptake into astrocytes stimulates aerobic glycolysis: a mechanism coupling neuronal activity to glucose utilization[J]. Proc Natl Acad Sci USA, 1994, 91(22): 10625-10629.
[10] Hyder F, Rothman DL, Bennett MR. Cortical energy demands of signaling and nonsignaling components in brain are conserved across mammalian species and activity levels[J]. Proc Natl Acad Sci USA, 2013, 110(9): 3549-3554.
[11] Wu A, Lee D, Xiong WC. Lactate metabolism, signaling, and function in brain development, synaptic plasticity, angiogenesis, and neurodegenerative diseases[J]. Int J Mol Sci, 2023, 24(17).
[12] Roumes H, Dumont U, Sanchez S, et al. Neuroprotective role of lactate in rat neonatal hypoxia-ischemia[J]. J Cereb Blood Flow Metab, 2021, 41(2): 342-358.
[13] Kennedy L, Glesaaen ER, Palibrk V, et al. Lactate receptor HCAR1 regulates neurogenesis and microglia activation after neonatal hypoxia-ischemia[J]. Elife, 2022, 11: e76451
[14] Nalbandian M, Takeda M. Lactate as a signaling molecule that regulates exercise-induced adaptations[J]. Biology, 2016, 5(4): 38.
[15] Margineanu MB, Mahmood H, Fiumelli H, et al. L-lactate regulates the expression of synaptic plasticity and neuroprotection genes in cortical neurons: a transcriptome analysis[J]. Front Mol Neurosci, 2018, 11:375.
[16] 颜杨林, 彭吾训, 张飞. DJ-1抗氧化应激作用及机制的研究进展[J]. 医学综述, 2021, 27(11): 2093-2098.
[17] Choong CJ, Mochizuki H. Involvement of mitochondria in Parkinson's disease[J]. Int J Mol Sci, 2023, 24(23): 17027.
[18] Shen ZY, Sun Q, Xia ZY, et al. Overexpression of DJ-1 reduces oxidative stress and attenuates hypoxia/reoxygenation injury in NRK-52E cells exposed to high glucose[J]. Int J Mol Med, 2016, 38(3): 729-736.
[19] Gao QS, Zhang YH, Xue H, et al. Brief inhalation of sevoflurane can reduce glial scar formation after hypoxic-ischemic brain injury in neonatal rats[J]. Neural Regen Res, 2021, 16(6): 1052-1061.
[20] Sarkar A, Singh MP. A complex interplay of DJ-1, LRRK2, and Nrf2 in the regulation of mitochondrial function in cypermethrin-induced Parkinsonism [J]. Mol Neurobiol, 2024, 61(2): 953-970.
[21] Lu Y, Zhang J, Han B, et al. Extracellular vesicles DJ-1 derived from hypoxia-conditioned hMSCs alleviate cardiac hypertrophy by suppressing mitochondria dysfunction and preventing ATRAP degradation[J]. Pharmacol Res, 2023, 187: 106607.
[22] Danileviciute E, Zeng N, Capelle CM, et al. PARK7/DJ-1 promotes pyruvate dehydrogenase activity and maintains T(reg) homeostasis during ageing[J]. Nat Metab, 2022, 4(5): 589-607.
