Extended Abstract
Background and Aim: Cadmium (Cd)-induced kidney injury is a major public health concern due to its widespread environmental exposure and nephrotoxic effects. Cadmium accumulation in renal tissue can impair kidney function by inducing oxidative stress, inflammation, and cellular damage. In contrast, growing evidence suggests that regular physical exercise exerts protective effects on kidney health by improving protein metabolism, enhancing tubular reabsorption, and maintaining iron and urea homeostasis. In addition to exercise, naturally occurring dietary antioxidants have been shown to exert beneficial effects on the function of various organs. Among these antioxidants, gallic acid (GA), also known as 3,4,5-trihydroxybenzoic acid, is a naturally occurring polyphenolic compound found in a variety of fruits and plant-derived foods, including walnuts, sumac, hazelnuts, tea leaves, and oak. Owing to its potent antioxidant and anti-inflammatory properties, GA has attracted considerable attention as a potential therapeutic agent for reducing cadmium-induced tissue damage.
Although the independent effects of exercise and antioxidant supplementation on kidney tissue have been investigated, limited information is available regarding their combined effects on molecular pathways associated with cadmium-induced nephrotoxicity. Therefore, the present study aimed to investigate the combined effects of eight weeks of high-intensity interval training (HIIT) and gallic acid supplementation on the expression of SIRT1 and miR-9-5p in the kidney tissue of cadmium-exposed rats.
Materials and Methods: This experimental study employed a posttest-only design with a control group. A total of 42 male Sprague–Dawley rats, aged 8–9 months and weighing 250–270 g, were obtained from the Pasteur Institute Laboratory Animal Breeding and Reproduction Center. The sample size was determined based on previous studies using a similar experimental design, in which seven animals were allocated to each of six experimental groups, resulting in a total sample of 42 rats. Following purchase, the animals were transferred to the Exercise Physiology Laboratory of the Bitaran Research Center and allowed to acclimatize to the laboratory environment for one week under standard housing conditions. All experimental procedures were conducted in accordance with the ethical guidelines for the care and use of laboratory animals and were approved by the Biomedical Research Ethics Committee of the Islamic Azad University, Borujerd Branch (Approval No. IR.IAU.B.REC.1403.094). After the acclimatization period, the rats were randomly assigned to six groups (n=7 per group): healthy control (HC), cadmium (Cd), sham, Cd + gallic acid (GA), Cd + high-intensity interval training (HIIT), and Cd + HIIT + GA. Cadmium chloride was purchased from Sigma-Aldrich (USA). A dose equivalent to 5 mg/kg body weight was prepared daily by dissolving the appropriate amount of cadmium in the drinking water of the designated groups, allowing the animals to receive the required dose through voluntary water consumption. Gallic acid (GA; Sigma-Aldrich, USA; Catalog No. 27645) was administered at a dose of 20 mg/kg body weight per day. The required amount of GA was dissolved in a food and water flavoring solution to facilitate oral administration. Before the intervention, rats assigned to the exercise groups underwent a one-week familiarization period consisting of five treadmill-running sessions at a speed of 8 m/min for 5 min per session. Subsequently, the HIIT protocol was performed for eight weeks, with three sessions per week. Before initiating the training program, each rat’s maximum running speed was determined using the incremental treadmill test developed by Bedford et al. (1979) and subsequently standardized by Lindberg and colleagues. The HIIT program consisted of treadmill running at an intensity corresponding to 80–110% of maximal oxygen uptake (VO₂max), with exercise intensity and duration adjusted according to the experimental protocol. Data were analyzed using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test for multiple comparisons. Statistical significance was established at p<0.05.
Results: The Shapiro–Wilk test confirmed that all study variables were normally distributed. One-way analysis of variance (ANOVA) revealed significant differences among the experimental groups in the expression of miR-9-5p (F=261.31, p<0.001) and SIRT1 (F=272.97, p<0.001). Tukey’s post hoc analysis showed no significant difference in miR-9-5p expression between the healthy control (HC) and sham groups (p=0.47, MD=−0.05). Compared with the HC group, miR-9-5p expression was significantly reduced in the cadmium (Cd) group (p<0.001, MD=0.61). In contrast, miR-9-5p expression was significantly increased in the Cd + GA (p<0.001, MD=−0.54), Cd + HIIT (p<0.001, MD=−0.48), and Cd + HIIT + GA (p<0.001, MD=−1.15) groups compared with the Cd group. No significant difference was observed between the Cd + GA and Cd + HIIT groups (p=0.51, MD=−0.05). However, the Cd + HIIT + GA group exhibited significantly higher miR-9-5p expression than both the Cd + GA (p<0.001, MD=−0.60) and Cd + HIIT (p<0.001, MD=−0.66) groups (Figure 1). Similarly, no significant difference in SIRT1 expression was observed between the HC and sham groups (p=0.42, MD=−0.05). Cadmium exposure significantly reduced SIRT1 expression compared with the HC group (p<0.001, MD=−0.91). Treatment with GA (p<0.001, MD=−0.69), HIIT (p<0.001, MD=−0.63), and their combination (p<0.001, MD=−0.72) significantly increased SIRT1 expression compared with the Cd group. No significant differences were found between the Cd + GA and Cd + HIIT groups (p=0.31) or between the Cd + GA and Cd + HIIT + GA groups (p=0.83). However, SIRT1 expression was significantly higher in the Cd + HIIT + GA group than in the Cd + HIIT group (p=0.02, MD=0.09) (Figure 1).
Conclusion: The findings of the present study suggest that both high-intensity interval training (HIIT) and gallic acid (GA) attenuate cadmium-induced renal injury by modulating molecular pathways associated with oxidative stress and cellular protection. HIIT may enhance the endogenous antioxidant defense system through activation of the NRF2 signaling pathway, thereby increasing the expression and activity of antioxidant enzymes such as superoxide dismutase (SOD), glutathione peroxidase (GPx), and reduced glutathione (GSH). In addition, HIIT may improve mitochondrial function, promote mitochondrial biogenesis, enhance cellular energy metabolism, and inhibit apoptosis through SIRT1- and SIRT3-dependent signaling pathways. Exercise may also suppress inflammatory responses by modulating macrophage polarization and inhibiting TLR2 and NF-κB signaling.
Gallic acid appears to exert nephroprotective effects through complementary antioxidant and anti-inflammatory mechanisms. In addition to directly scavenging reactive oxygen species, GA may inhibit NF-κB, NADPH oxidase, and myeloperoxidase activity, reduce lipid peroxidation and nitric oxide production, suppress pro-inflammatory cytokines such as interleukin-6 (IL-6), and promote SIRT1-mediated mitochondrial biogenesis. Collectively, these molecular adaptations may contribute to the preservation of renal cellular integrity during cadmium exposure. The present findings further indicate that the combined administration of HIIT and GA produced greater beneficial effects on SIRT1 and miR-9-5p expression than either intervention alone. This enhanced response may be attributed to the simultaneous activation of endogenous antioxidant defenses, attenuation of oxidative stress and inflammatory signaling, improvement of mitochondrial homeostasis, and coordinated regulation of SIRT1 and miR-9-5p. Therefore, the combination of HIIT and GA may represent a promising strategy for mitigating cadmium-induced renal injury, although further mechanistic and clinical studies are required to confirm these findings.
Ethical Considerations: It should be noted that all ethical principles regarding the use of laboratory animals in this study were observed under the supervision of the Ethics Committee for Biomedical Research at the Islamic Azad University, Borujerd Branch (approval code: IR.IAU.B.REC.1403.094).
Compliance with Ethical Guidelines: The research followed the ethical standards of the Declaration of Helsinki and institutional guidelines.
Funding: No funding.
Conflicts of Interest: The authors declare no conflicts of interest regarding the publication of this study.