Exploratory metabolomic analysis based on UHPLC-Q-TOF-MS/MS to study hypoxia-reoxygenation energy metabolic alterations in HK-2 cells

Abstract

Purpose

Renal ischemia-reperfusion injury(IRI)is a major cause of acute kidney injury(AKI), the injury and repair of renal tubular epithelial cells play an important role in the pathological process of IR-AKI. Metabolomics was used to detect cell metabolism alterations and metabolic reprogramming in the initial injury, peak injury, and recovery stage of human renal proximal tubular cells (HK-2 cells) to provide insights into clinical prevention and treatment of IRI-induced AKI.

Methods

An in vitro ischemia-reperfusion (H/R) injury and the recovery model of HK-2 cells were established at different times of hypoxia/reoxygenation. Comprehensive detection of metabolic alterations in HK-2 cells after H/R induction by nontarget metabolomics. Interconversion of glycolysis and fatty acid oxidation (FAO) in HK-2 cells after H/R induction was examined by western blotting and qRT-PCR.

Results

Multivariate data analysis found significant differences among the groups, with significant changes in metabolites such as glutamate, malate, aspartate, and L-palmitoylcarnitine. Hypoxia-reoxygenated HK-2 cells are accompanied by altered metabolisms such as disturbance of amino acid and nucleotide metabolism, dysregulation of lipid metabolism, increased glycolysis, and metabolic reprogramming, which manifests as a shift in energy metabolism from FAO to glycolysis.

Conclusion

The development of IRI-induced AKI in HK-2 cells is accompanied by the disturbance of amino acid, nucleotide, and tricarboxylic acid cycle metabolism and specifically metabolic reprogramming of FAO to glycolytic conversion. The timely recovery of energy metabolism in HK-2 cells is of great significance for treating and prognosis IRI-induced AKI.

Keywords:

• Metabolomics
• acute kidney injury
• metabolic reprogramming
• glycolysis
• metabolic pathways

Author contributions

Xiaoshuang Zhou and Rongshan Li designed the study. Xiaoyu Yang, Ailing Kang and Yuanyue Lu performed the experiments and confirm the authenticity of all the raw data. Xiaoyu Yang, Xiaoshuang Zhou and Rongshan Li analyzed the data and wrote the manuscript. Yafeng Li and Lili Guo provided laboratory support. All authors read and approved the final manuscript.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

All data are included in the manuscript.

Additional information

Funding

This work was supported by grants from the National Natural Science Foundation of China (NSFC) [Grant Numbers 82170708] and Key Research and Development (R&D) Projects of Shanxi Province [201803D31151].