Tourniquets are commonly used in orthopedic limb surgery to reduce intraoperative bleeding and improve surgical visibility, allowing for more precise procedures (1). However, tourniquets can also cause complications such as thromboembolism and ischemia–reperfusion injury (IRI) in muscles (2). Ischemia–reperfusion of large muscle tissues can induce rhabdomyolysis by generating inflammatory cells (3). The systemic inflammatory response generated by this process increases vascular permeability, decreases intravascular volume, and activates the renin-angiotensin-aldosterone system. This leads to renal vasoconstriction, reduced renal blood flow and oxygenation, and the generation of free radicals, primarily reactive oxygen species (ROS), which in severe cases can result in acute kidney injury (AKI) (4). The incidence of tourniquet-related AKI ranges from 0.8 to 17.2%, depending on whether patients have risk factors such as diabetes and hypertension (5). Currently, there are no specific measures to prevent or treat tourniquet-induced kidney or other organ damage (6).

Ischemic preconditioning (IPC) involves performing multiple transient cycles of ischemia and reperfusion to the target organ, which makes the organism resistant to further severe injury. This protective phenomenon was first identified in cardiac experiments by Murry et al. (7). A recent systematic review showed that IPC can be applied to the kidney, providing a protective effect (8). Remote ischemic preconditioning (RIPC) involves exposing tissues or organs distant from the target organ (usually upper or lower limbs) to brief cycles of ischemia and reperfusion to minimize subsequent damage to the target organ. Przyklenk et al. conducted the first study on RIPC in 1993 and concluded that RIPC could mitigate myocardial IRI (9). RIPC has the potential to protect a wide range of organs and tissues, including the heart, brain, kidneys, lungs, liver, and skin (10, 11). Multiple meta-analyses have indicated that the incidence of AKI after cardiac surgery is much higher than after myocardial infarction. RIPC before the induction of general anesthesia reduces peak serum creatinine (SCr) and neutrophil gelatinase-associated lipocalin (NGAL) levels and decreases the incidence of AKI by approximately 15% in patients with high-risk factors (12, 13). Studies on the short-term and long-term outcomes of kidney transplantation revealed that preoperative RIPC not only reduced the incidence of acute rejection in recipients but also led to sustained improvements in glomerular filtration rate over a period of five years, prolonging the use of the transplanted kidney (14, 15).

Many studies on RIPC have focused on renal function after cardiac surgery, with few exploring the role of RIPC in renal injury caused by tourniquets, particularly with repeated use (16, 17). Therefore, this study aims to investigate the renoprotective effect of RIPC on repeated tourniquet application in extremity surgery by detecting renal injury biomarkers, oxidative stress markers, and inflammatory factors in the blood. These results may help identify a safe, convenient, and effective perioperative renoprotective measure.

Tourniquets are commonly used in extremity surgeries for orthopedic patients to assist surgeons in better completing the surgery. However, muscle IRI from prolonged tourniquet use can lead to tissue and organ damage and, in severe cases, multiple systemic organ failure (18). RIPC has emerged in recent years as a convenient, non-invasive, and inexpensive protective method applicable to a wide range of organs. Current studies have found that preoperative use of RIPC reduces the incidence of postoperative AKI without adverse events (19). Our study found that RIPC could attenuate AKI induced by repeated tourniquet application in extremities, which may through inhibiting ROS production, oxidative stress and inflammatory responses, and inducing transient cell cycle arrest. This might provide a new strategy for mitigating perioperative AKI in the clinic.

The exact mechanism by which repeated tourniquet application and limb IRI lead to AKI is not fully elucidated. Most studies suggest that this phenomenon may be caused by chronic hypoperfusion and hypoxia of the kidney, related to the production of ROS such as superoxide, hydrogen peroxide, and hydroxyl radicals, as well as the involvement of inflammatory cells (20). During the ischemic phase, decreased aerobic metabolism and increased anaerobic metabolism led to the depletion of adenosine triphosphate (ATP), disruption of intracellular redox homeostasis, accumulation of acidic metabolites, and the failure of ATP-dependent sodium-potassium pumps and sodium-calcium exchangers. These disruptions result in increased intracellular Na⁺ and Ca²⁺, ultimately causing apoptosis (21). Superoxide is a key mediator of cell necrosis and tissue injury during reperfusion (22). It is produced when xanthine is oxidized to uric acid. During ischemia, decreased ATP leads to increased hypoxanthine, which is then oxidized during reperfusion to produce large amounts of superoxide (23, 24). SOD is an antioxidant enzyme that converts superoxide into oxygen and hydrogen peroxide, reducing ROS levels and protecting organs from oxidative damage. It is one of the most crucial antioxidant enzymes in cells (25). Researches have shown that SOD can protect the kidneys from chronic ischemic injury and prevent renal insufficiency through its antioxidant, vasodilatory, and antihypertensive effects (26). ROS-induced cell damage initiates lipid peroxidation, with MDA being a harmful end product that can result in cell damage and apoptosis, often used to indicate ROS levels (27, 28). Large amounts of ROS released into the blood during reperfusion elicit an inflammatory response, with the activation of inflammatory pathways and recruitment of inflammatory cells being early responses to renal injury.

To date, no single definitive measure has been found to prevent or mitigate AKI. Therefore, it is significant to explore simple methods to attenuate AKI caused by repeated tourniquet application and limb IRI by suppressing ROS levels and inflammatory responses. RIPC involves exposing the body to brief cycles of ischemia and reperfusion by pretreating organs and tissues far from the target organ, which has been shown to attenuate tissue and organ damage (29). The exact mechanism by which RIPC protects the kidney is not fully understood. However, most studies suggest it may involve the production of nitric oxide or nitrite, the release of damage-associated molecular patterns, activation of transient cell cycle arrest in renal tubular epithelial cells, and the clearance of damaged mitochondria via mitochondrial autophagy (30). RIPC may protect the kidney by activating natural defenses that cause renal tubular epithelial cells to undergo transient cell cycle arrest, enabling them to withstand subsequent oxidative stress or IRI. Many studies have identified TIMP-2 and IGFBP-7 as markers of cell cycle arrest (31). Alexander et al. revealed that RIPC could reduce [TIMP-2] × [IGFBP-7] levels in patients at high risk for AKI during cardiac surgery, contributing to a significant reduction in the incidence of AKI (12). Our study also confirmed that [TIMP-2] × [IGFBP-7] and KIM-1 levels were significantly reduced at T1, T2, and T3 in the RIPC group, suggesting that RIPC may attenuate tubular injury caused by repeated tourniquet application by inducing transient tubular epithelial cell cycle arrest.

Cys-C is a biomarker that is freely filtered through the glomeruli and completely absorbed by renal tubular epithelial cells, with minimal influence from external disturbances. Therefore, it has higher sensitivity and specificity in predicting AKI compared to SCr and BUN (32). Kasepalu et al. demonstrated that SCr, urea, and Cys-C were significantly reduced in the RIPC group of patients undergoing lower extremity revascularization (33). Similarly, our study found that the RIPC group exhibited lower levels of SCr and BUN at T4, and decreased level of Cys-C at T1, T2, and T3, indicating that RIPC could mitigate the effects of limb IRI induced by repeated tourniquet application on postoperative glomerular filtration function.

Oxidative stress markers and inflammation levels were also examined in this study. NGAL, primarily secreted by immune cells such as neutrophils, macrophages, and dendritic cells, is produced in response to inflammation and released after tubular injury during renal regeneration. It can be used as a predictor of renal function progression concerning renal failure (34). IL-18, an inflammatory factor released after acute ischemic injury, is considered an early diagnostic marker for AKI. Guo et al. demonstrated that RIPC attenuated postoperative NGAL levels compared to the control group (35). Luan et al. showed that IL-18 knockout ameliorated tubular injury and limited the progression of AKI to chronic kidney disease (36). Similarly, our study found that IL-18 and NGAL levels at T1, T2, and T3 were significantly lower in the RIPC group. Furthermore, RIPC also demonstrated the ability to mitigate oxidative stress in AKI induced by limb IRI by reducing MDA levels and increasing SOD levels in our study. These findings suggest that the protective effect of RIPC may be mediated through the inhibition of oxidative stress and inflammatory responses in AKI induced by limb IRI.

This study has several limitations. Firstly, it was conducted as a single-center study with a limited sample size, which may affect the generalizability of the results. Future studies with larger, multicenter cohorts are necessary to validate these findings. Secondly, the antioxidant and renoprotective properties of anesthetic drugs were not considered in this study. Although we designed the study to minimize the potential impact of anesthetics on the outcomes, future research should explore the synergistic effects of anesthetic agents and RIPC on renal protection. Despite these limitations, our study demonstrates that RIPC is a promising and non-invasive method to attenuate AKI induced by repeated tourniquet use in extremity surgeries. The findings suggest potential clinical applications for improving perioperative renal outcomes, highlighting the need for further investigation into RIPC’s protective mechanisms and broader applicability. Future research should focus on elucidating the precise mechanisms underlying RIPC’s renoprotective effects and evaluating its long-term clinical benefits across diverse surgical populations.