Acidic preconditioning induced intracellular acid adaptation to protect renal injury via dynamic phosphorylation of focal adhesion kinase-dependent activation of sodium hydrogen exchanger 1

• Published in: Cell communication and signaling Vol. 22; no. 1; pp. 393 - 21
• Main Authors: Chen, Annan, Zhang, Jian, Yan, Zhixin, Lu, Yufei, Chen, Weize, Sun, Yingxue, Gu, Qiuyu, Li, Fang, Yang, Yan, Qiu, Shanfang, Lin, Xueping, Zhang, Dong, Teng, Jie, Fang, Yi, Shen, Bo, Song, Nana, Ding, Xiaoqiang
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 08.08.2024; BioMed Central; BMC
• Subjects: Acid-base equilibrium; Acidic preconditioning; Acidosis; Acids - metabolism; Actin; Adaptation; Analysis; Animal models; Animals; Apoptosis; Care and treatment; Carrier proteins; Caspase-3; Caspase-9; Cell adhesion; Cell adhesion & migration; Cell Line; Cell migration; Cytoskeleton; Development and progression; Epithelial cells; Fluorescent indicators; Focal adhesion kinase; Focal Adhesion Protein-Tyrosine Kinases - metabolism; Health aspects; Hemin; Homeostasis; Humans; Hydrogen; Hydrogen-Ion Concentration; Hypoxia; Immunofluorescence; Intracellular; Ischemia; Ischemic Preconditioning; Kidney - metabolism; Kidney - pathology; Kidney failure; Kidneys; Kinases; Localization; Male; Mice; Mice, Inbred C57BL; Muscle proteins; Na+/H+ transporter 1; NADPH Oxidase 4 - genetics; NADPH Oxidase 4 - metabolism; NOX4; NOX4 protein; pH effects; pHi adaptation; Phosphorylation; Physiological aspects; Prevention; Protein kinases; Proteins; Reactive oxygen species; Reactive Oxygen Species - metabolism; Renal epithelia cells; Reperfusion; Reperfusion Injury - metabolism; Reperfusion Injury - pathology; Signal transduction; Sodium-Hydrogen Exchanger 1 - genetics; Sodium-Hydrogen Exchanger 1 - metabolism; Western blotting; China; Acidic preconditioning; NOX4; Cytoskeletal remodeling; Focal adhesion kinase; Na+/H+ transporter 1; pHi adaptation; Renal epithelia cells
• ISSN: 1478-811X; 1478-811X
• DOI: 10.1186/s12964-024-01773-w

Disruptions in intracellular pH (pH ) homeostasis, causing deviations from the physiological range, can damage renal epithelial cells. However, the existence of an adaptive mechanism to restore pH to normalcy remains unclear. Early research identified H as a critical mediator of ischemic preconditioning (IPC), leading to the concept of acidic preconditioning (AP). This concept proposes that short-term, repetitive acidic stimulation can enhance a cell's capacity to withstand subsequent adverse stress. While AP has demonstrated protective effects in various ischemia-reperfusion (I/R) injury models, its application in kidney injury remains largely unexplored. An AP model was established in human kidney (HK2) cells by treating them with an acidic medium for 12 h, followed by a recovery period with a normal medium for 6 h. To induce hypoxia/reoxygenation (H/R) injury, HK2 cells were subjected to hypoxia for 24 h and reoxygenation for 1 h. In vivo, a mouse model of IPC was established by clamping the bilateral renal pedicles for 15 min, followed by reperfusion for 4 days. Conversely, the I/R model involved clamping the bilateral renal pedicles for 35 min and reperfusion for 24 h. Western blotting was employed to evaluate the expression levels of cleaved caspase 3, cleaved caspase 9, NHE1, KIM1, FAK, and NOX4. A pH-sensitive fluorescent probe was used to measure pH , while a Hemin/CNF microelectrode monitored kidney tissue pH. Immunofluorescence staining was performed to visualize the localization of NHE1, NOX4, and FAK, along with the actin cytoskeleton structure in HK2 cells. Cell adhesion and scratch assays were conducted to assess cell motility. Our findings demonstrated that AP could effectively mitigate H/R injury in HK2 cells. This protective effect and the maintenance of pH homeostasis by AP involved the upregulation of Na /H exchanger 1 (NHE1) expression and activity. The activity of NHE1 was regulated by dynamic changes in pH -dependent phosphorylation of Focal Adhesion Kinase (FAK) at Y397. This process was associated with NOX4-mediated reactive oxygen species (ROS) production. Furthermore, AP induced the co-localization of FAK, NOX4, and NHE1 in focal adhesions, promoting cytoskeletal remodeling and enhancing cell adhesion and migration capabilities. This study provides compelling evidence that AP maintains pH homeostasis and promotes cytoskeletal remodeling through FAK/NOX4/NHE1 signaling. This signaling pathway ultimately contributes to alleviated H/R injury in HK2 cells.