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. 2024 Oct 8;46(2):2406395. doi: 10.1080/0886022X.2024.2406395

Reserpine alleviates cisplatin-induced acute kidney injury via anti-ferroptosis and cGAS/STING pathway

Nahua Xu a,#, Rong Mu a,#, Siyuan Deng b, Ye Han c, Yanyun Shi d, Xuemei Fu c, Hui Li a,c,e,✉, Qi Yao e,✉
PMCID: PMC11463010  PMID: 39377110

Abstract

Cisplatin plays a pivotal role in the chemotherapy treatment of various cancers, but its use is often limited due to its nephrotoxic side effects. Identifying compounds that can mitigate cisplatin-induced nephrotoxicity is therefore of great importance. This study focused on evaluating the protective effects of reserpine against cisplatin-induced acute kidney injury. Reserpine was found to significantly safeguard against kidney damage caused by cisplatin, as indicated by the decreased levels of serum creatinine, blood urea nitrogen, and lactate dehydrogenase induced by cisplatin. Moreover, reserpine improved kidney histology damage caused by cisplatin treatment, with hematoxylin-eosin and periodic acid-Schiff staining revealing notable recovery from renal injury. Mechanistically, reserpine mitigated oxidative stress triggered by cisplatin and exhibits the ability to inhibit ferroptosis both in vivo and in vitro. Additionally, reserpine blocked the activation of the cGAS/STING signaling pathway and the subsequent expression of inflammatory genes, thus reducing inflammation-driven kidney damage. In summary, the findings suggest that reserpine offers a promising new strategy for preventing nephrotoxicity induced by cisplatin.

Keywords: Reserpine, acute kidney injury, cisplatin, ferroptosis

Graphical Abstract

graphic file with name IRNF_A_2406395_UF0001_C.jpg

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Schematic diagram showing proposed mechanism on the role of reserpine in protection of cisplatin-induced AKI (Created with bioicons.com). Cisplatin treatment leads to oxidative stress and mitochondrial dysfunction, which releases mitochondrial DNA, activating the cGAS/STING/TBK1/IRF3 signaling cascade and consequently inflammatory cytokine expression. Reserpine could ameliorate cisplatin induced AKI and inflammation by targeting the anti-oxidant pathway and by inhibiting the activation of cGAS/STING signaling.

1. Introduction

Cisplatin is a platinum-based inorganic compound, which is widely used as an effective anticancer agent against various tumors, including lung, stomach, and ovaries [1,2]. It works by forming DNA crosslinks and adducts in cancer cells, initiating a DNA damage response, leading to cell cycle arrest and cell death [3–5]. Despite its efficacy, clinical application of cisplatin is hampered by its toxicity, particularly its nephrotoxic effects, which can lead to acute kidney injury (AKI) and chronic kidney disease [6,7]. Cisplatin is predominantly eliminated by the kidneys via glomerular filtration and tubular secretion, causing high concentration of cisplatin in renal tissue and, as a result, significant nephrotoxicity [8–10]. Approximately 30% of patients undergoing cisplatin therapy experience AKI with decline in renal function [6,11]. The nephrotoxicity of cisplatin is both cumulative and dose-dependent, often necessitating a reduction in dosage or discontinuation of treatment [12]. Repeated AKI episodes can lead to chronic renal issues, including tubular dysfunction and renal failure [13]. The underlying mechanisms of cisplatin-induced AKI include damage to the proximal tubules, cell death, oxidative stress, inflammation, and vascular damage within the kidneys [14–16]. Currently, there are no effective treatments for managing cisplatin-induced renal damage, highlighting an urgent need for new therapeutic strategies.

Recent research underscores a critical role of oxidative stress in cisplatin-induced nephrotoxicity, marked by increased levels of malondialdehyde (MDA) and 4-Hydroxynonenal (4-HNE), alongside reduced activity of antioxidant enzymes like superoxide dismutase and catalase [17–19]. Cisplatin deactivates thiol-containing antioxidants such as glutathione and metallothionein within renal tubular cells [20,21]. It also inhibits antioxidant enzymes, leading to elevated reactive oxygen species (ROS) levels [22,23]. These ROS impair mitochondrial complex enzymes, disrupt the oxidative phosphorylation chain, and cause ATP depletion, lipid peroxidation, and ultimately, cell death [14,24].

Inflammation plays a significant role in cisplatin-induced nephrotoxicity, with increased renal TNF-α expression observed in cisplatin-treated mice [25]. Inhibition or genetic deletion of TNF-α significantly mitigates renal dysfunction and damage caused by cisplatin, indicating its pivotal role in the nephrotoxic process [26–28]. Additionally, AKI is characterized by mitochondrial dysfunction and activation of the innate immune system. The cGAS-STING pathway, which detects cytosolic DNA and activates innate immunity, is implicated in the inflammatory response to cisplatin-induced renal damage [29–31]. Leakage of mitochondrial DNA into the cytosol, possibly through BAX/BAK mitochondrial membrane pores activates cGAS-STING signaling, ultimately exacerbating inflammation and AKI [32,33]. This process is attenuated in STING-deficient mice [29].

Reserpine is a traditional antihypertensive medicine that inhibits alpha-adrenergic transmission [34–36]. Hypertension is one of the common causes of kidney injury. Whether reserpine, as an antihypertensive drug, can alleviate kidney injury is yet to be determined. In this study we found that reserpine protects cisplatin-induced AKI in mouse model. Our findings reveal that reserpine significantly reduces kidney damage caused by cisplatin, as evidenced by lower serum markers and improved kidney histology. Reserpine also diminishes cGAS/STING-mediated inflammation and immune cell infiltration. Mechanistically, it appears that reserpine may alleviate cisplatin-induced oxidative stress by upregulating the anti-ferroptosis pathway NRF2/SLC7A11/GPX4.

2. Materials and methods

2.1. Chemicals and reagents

Cisplatin (Targetmol, T1564), Reserpine (MCE, HY-N0480A), RSL3 (MCE, HY-100218A), Imidazole ketone erastin (IKE, Targetmol, T5523), Liproxstatin-1 (Lip1, MCE, HY-12726), dimethyl sulfoxide (DMSO, Solarbio, D8371), NGAL (ABclonal, A24538, 1:1000), GPX4 (Huabio, ET1706-45, 1:10000), NRF2(Proteintech, 16396-1-AP, 1:2000), 4-HNE (Invitrogen, MA5-27570, 1:1000), SLC7A11 (CST, 12691S, 1:1000), cGAS (CST, 15102S, 1:1000), STING (CST, 50494 T, 1:1000), TBK1 (CST, 3504 T, 1:1000), p-TBK1 (CST, 5483 T, 1:1000), p-IRF3 (CST, 29047 T, 1:1000), Tubulin (ABclonal, AC024, 1:500), GAPDH (ABclonal, AC002, 1:10000), Lamin B1 (ABclonal, A1910, 1:1000), Peroxidase AffiniPure™ Goat Anti-Mouse IgG (Jackson, 111-035-008), Peroxidase AffiniPure™ Goat Anti-Rabbit IgG (H + L) (Jackson, 111-035-003).

2.2. Animal

The animal study conducted in this research was thoroughly reviewed and granted approval by the Laboratory Animal Welfare and Ethics Committee of the Third Military Medical University, with the Animal Ethical Statement number AMUWEC20242007. Strict adherence to the approved guidelines was ensured throughout the entire process. Adult male mice, aged 6–8 weeks and weighing between 20–22 grams, were procured from Chongqing Lepitt Biotechnology Co., Ltd., with a C57BL/6J genetic background. The mice were housed in cages maintained at a constant temperature of 21.0 °C and humidity level of 55 ± 5%, ensuring their comfort and welfare. They were provided with unlimited access to food and water, crucial for their health and well-being. Prior to the commencement of the experiment, all mice were reared in an appropriate environment and allowed to acclimate for a duration of one week.

2.3. Cisplatin-induced acute renal injury model

To investigate the therapeutic potential of reserpine in cisplatin-induced acute renal injury, mice were allocated into three distinct groups: the vehicle control group, the cisplatin-treated group, and the combination group of cisplatin and reserpine. Acute renal injury was induced in the mice via intraperitoneal injection of cisplatin at a dosage of 20 mg/kg. Prior to cisplatin injection, the mice in the combination group received an injection of reserpine at a concentration of 5 mg/kg. The control group received an injection of the equivalent volume of normal saline. 72 h following the cisplatin injection, all mice were sacrificed, and their blood and kidneys were harvested for subsequent in-depth analysis.

2.4. Serum biochemical analysis

The contents of serum creatinine (SCr), blood urea nitrogen (BUN), and lactate dehydrogenase (LDH) in the serum of mice in each experimental group were measured by commercial kits. The commercial kits included BUN colorimetric assay kit (Elabscience, E-BC-K183-M), Cr colorimetric assay kit (Elabscience, E-BC-K188-M), LDH activity assay kit (Elabscience, E-BC-K046-M). Initially, mouse blood was collected in EP tubes and allowed to rest at room temperature for 20 min. Subsequently, the tubes were centrifuged at 4 °C and 3000 revolutions per minute (rpm) for 15 min. The supernatant was then extracted for measurement, with the measurement steps being carried out in accordance with the manufacturer’s guidelines.The absorbance at the optimal wavelength was detected using microplate reader (Thermo Fisher, Varioskan Flash), and the concentration of SCr, BUN and LDH in mice serum was calculated according to the standard curve.

2.5. Western blot (WB) analysis

1% Sodium dodecyl sulfate (Biofroxx, 3250GR500) which containing 1 mM PMSF (SolarBio, 329-98-6) was used to lyse the kidney tissue. Repeatedly grinding the sheared kidney tissue on ice, we centrifuged the sample to separate the supernatant. Subsequently, the supernatant was subjected to a metal bath maintained at 100 °C for 10 min to ensure complete denaturation of proteins. After adding LDS loading buffer (Invitrogen, NP 0008), protein was separated by electrophoresis. Then protein was transferred to PVDF membrane (Merck millipore, IPVH00010), blocked with 5% skim milk (Biofroxx, 1172GR500) in TBST (Beyotime, ST663) for 1 h. The membrane was incubated overnight at 4 °C with primary antibody, and the secondary antibody of goat anti-rabbit or anti-mouse IgG of horseradish peroxidase (HRP) was incubated with the membrane for 1 h at room temperature. The blots were imaged by imager (Bio-Rad, ChemiDoc MP Imaging System).

2.6. Renal pathology assessment

Renal tissue was fixed in 4% paraformaldehyde (Beyotime, P0099) overnight. The mouse kidney sections embedded in paraffin were dewaxed and hydrated. H&E dye solution (Servicebio, G1120) and PAS dye solution set (Servicebio, G1008) were used to stain the sections according to the manufacturer’s instructions, and the stained sections were scanned and photographed under an optical microscope (Nikon, E100). Random fields were captured from each H&E-stained section at 400X magnification, and kidney injury was examined in a blinded manner and scored based on the percentage of cortical tubule damage: 0 indicates no damage, 1 indicates 1–25%, 2 indicates 26–50%, 3 indicates 51–75%, and 4 indicates 76–100%.

2.7. Renal immunohistochemical staining

The mouse kidney sections embedded in paraffin were dewaxed and hydrated. These sections were then placed in a repair cassette filled with EDTA antigen retrieval buffer (pH 8.0) and subjected to antigen retrieval in a microwave oven (Galanz, P70D20TL-P4). Following antigen retrieval, endogenous peroxidase activity was blocked, and the slides were sealed with serum to prevent nonspecific binding of antibodies. Subsequently, the slides were incubated overnight at 4 °C with primary antibodies specific for CD68 (Aifang Biological, SAF006), Ly6G (Aifang Biological, SAF013), and TNF-α (Aifang Biological, AF06294). After incubating the secondary antibody (Aifang Biological, AFIHC003) for 2 h, the sections were stained with DAB chromogenic solution (Aifang Biological, AFIHC004), which produces a visible brown coloration at the sites of antibody binding. The nuclei were then counterstained with hematoxylin (Aifang Biological, AFIHC005). Finally, the stained sections were sealed, scanned using a microscope (Nikon, E100), and images were captured with an imaging system (Nikon, DS-U3).

2.8. RNA-sequencing (RNA-seq)

Fresh kidney tissues from the vehicle group, cisplatin group, and cisplatin + reserpine group were collected and stored in Animal tissue RNA stable preservation solution (Beyotime, R0118). The RNA extraction, database building, sequencing and analysis were all completed by Hangzhou Lianchuan Biotechnology Co., Ltd. Total RNA extraction was performed utilizing Trizol reagent (Thermo Fisher, 15596018), adhering to the prescribed protocol of the manufacturer. The quantity and purity of the extracted total RNA were assessed using the Bioanalyzer 2100 with the RNA 6000 Nano LabChip Kit (Agilent, CA, USA, 5067-1511). Only high-quality RNA samples, indicated by a RNA Integrity Number (RIN) greater than 7.0, were selected for subsequent sequencing library construction. Sequencing was carried out using 2 × 150bp paired-end (PE150) technology on an Illumina NovaseqTM6000 sequencer, following the recommended protocol provided by the vendor. Differential gene expression analysis was conducted using DESeq2 software to compare two distinct groups, and edgeR was employed for the comparison between individual samples. Genes were identified as differentially expressed based on a false discovery rate (FDR) threshold of less than 0.05 and an absolute fold-change value of greater than or equal to 2.

2.9. Cell culture and treatment

HK-2/Caov3/HT0180 cells were obtained from Wuhan Pricella Biotechnology Co., Ltd. All cells were cultured in DMEM medium (Gibco, C11995500BT) supplemented with 10% fetal bovine serum (FSP500, EXCELL) and 1% penicillin–streptomycin (Beyotime, C0222). Cells were cultured in an incubator with 5% CO2 at 37 °C. Cells were seeded at 4 × 103 cells/well in 96‐well plates before 24 h, and synchronously treated with the drugs at the indicated concentrations for an additional 24 h. cell viability was detected using the CCK‐8 assay (Meilunbio, MA0218-5).

2.10. Data and statistics analysis

GraphPad Prism software (version 9.5.0) was used for statistical analysis. Data are expressed as mean ± SD. Differences among three groups were calculated using ANOVA, while the Student’s t-test was employed to analyze the differences between two groups. p-values are denoted as follows: *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; ns, not significant. Advanced heatmap and Venn plots were performed using the OmicStudio tools at https://www.omicstudio.cn.

3. Results

3.1. Effect of reserpine on SCr, BUN and LDH in cisplatin induced AKI

To explore the impact of reserpine on cisplatin-induced AKI, mice were treated with reserpine 1 h before receiving cisplatin. After 72 h of cisplatin treatment, the mice were euthanized for kidney damage assessment (Figure 1). Serum creatinine (SCr) and blood urea nitrogen (BUN) levels are established indicators of kidney function. Cisplatin administration significantly elevated SCr and BUN levels, but pretreatment with reserpine markedly mitigated these increases (Figure 2A, B). Serum lactate dehydrogenase (LDH) levels, which signify cell death, were also elevated following cisplatin treatment but were reduced by reserpine pretreatment (Figure 2C). Additionally, N-acetyl-β-D-glucosaminidase (NGAL, gene name Lcn2), another critical marker of kidney injury, was upregulated by cisplatin at both the protein and mRNA levels but was suppressed by reserpine pretreatment (Figure 2D-F). These findings indicate that reserpine effectively diminishes cisplatin-induced acute kidney damage and cellular mortality.

Figure 1.

Figure 1.

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(A) Experimental protocol of cisplatin induced renal injury in mice. C57BL/6J mice were treated with intraperitoneal injection of reserpine 1 h before cisplatin (Created with scidraw.io). Blood and kidneys samples were collected after 72 h. (B) Chemical structures of reserpine and cisplatin.

Figure 2.

Figure 2.

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Reserpine alleviates cisplatin induced AKI. Serum levels of SCr (A), BUN (B), and LDH (C) were analyzed by the end of experiments in vehicle, cisplatin and cisplatin + reserpine groups (n = 6). (D) Western blot analysis of NGAL in vehicle, cisplatin and cisplatin + reserpine groups (n = 3). (E) Quantification of NGAL protein expression in vehicle, cisplatin and cisplatin + reserpine groups (n = 3). (F) The mRNA levels of Lcn2 (Lcn2 encodes NGAL) in vehicle, cisplatin and cisplatin + reserpine groups analyzed by RNA-seq (n = 3). Each bar represents the mean ± SD. **p < 0.01, ***p < 0.001, ****p < 0.0001.

3.2. The impact of reserpine on histopathological changes in cisplatin-induced AKI

Histological analysis using Hematoxylin and Eosin (H&E) and Periodic Acid-Schiff (PAS) staining revealed significant renal tubular epithelial cell damage caused by cisplatin, including cell swelling, desquamation, necrosis, tubular cast formation, and increased infiltration of inflammatory cells into the renal tubule interstitium. Reserpine pretreatment significantly ameliorated these histopathological changes, making the renal morphology more similar to the control group (Figure 3A, B). The renal tubular injury score, based on PAS staining, further confirmed the protective effect of reserpine against kidney damage (Figure 3C). These results underscore reserpine’s potential in alleviating renal damage induced by cisplatin.

Figure 3.

Figure 3.

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Reserpine relieved cisplatin induced histological injury. (A) Representative H&E staining images (100X and 400X) of kidney in vehicle, cisplatin and cisplatin + reserpine groups. In cisplatin group, H&E staining showed that some renal tubular epithelial cells were necrotic (yellow arrow), the nucleus was condensed and the structure was destroyed, showing eosinophilic homogeneity. Water-like degeneration of renal tubular epithelial cells (blue arrow), cell swelling, loose cytoplasm and light staining are common. Eosinophils can be seen in a small amount of renal tubules (green arrow). (B) PAS staining (400X) in vehicle, cisplatin and cisplatin + reserpine groups, black arrow indicates glycogen deposited in renal tubules. (C) The renal tubular injury score was calculated. Each bar represents the mean ± SD (n = 5). ***p < 0.001, ****p < 0.0001.

3.3. Reserpine reduces immune infiltration and TNF-α expression in cisplatin-induced AKI

Further analysis revealed that reserpine might also decrease immune cell infiltration following cisplatin treatment. Examination of neutrophil and macrophage infiltration, identified by surface markers Ly6G and CD68 respectively, showed significant reductions in immune cell infiltration with reserpine pretreatment. Concurrently, cisplatin-induced upregulation of TNF-α expression (Figure 4), which contributes to inflammatory kidney damage, was mitigated by reserpine.

Figure 4.

Figure 4.

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Reserpine reduces immune infiltration and TNF-α expression in cisplatin-induced AKI. (A) Immunohistochemical analysis of LY6G, CD68 and TNF-α in vehicle, cisplatin and cisplatin + reserpine groups (scale bar = 100 μm), Quantitative analysis is shown in (B). Results are expressed as mean ± SD, (n = 6). ***p < 0.001, ****p < 0.0001.

3.4. Reserpine mitigates oxidative stress and ferroptosis

Oxidative stress plays a crucial role in cisplatin-induced renal damage. Reserpine significantly lowered levels of 4-HNE (Figure 5A), a lipid peroxidation marker, following cisplatin treatment. Given the emerging role of ferroptosis, a form of programmed cell death characterized by lipid peroxidation, in cisplatin-induced renal injury, we conducted RNA-seq to examine changes in ferroptosis regulator gene expression (Figure 5B). Reserpine reversed the cisplatin-induced expression changes of key ferroptosis regulators, including downregulation of ferroptosis promoters like ACSL4, Alox5, and Alox5ap (Figure 5C). Analysis of ferroptosis inhibitors NRF2, GPX4, and SLC7A11 showed a significant decrease after cisplatin treatment, which was rescued by reserpine (Figure 6A, B). These findings suggest that reserpine may protect against cisplatin-induced renal injury by inhibiting ferroptosis.

Figure 5.

Figure 5.

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Reserpine alleviates oxidative stress induced by cisplatin. (A) Western blot analysis of 4-HNE in vehicle, cisplatin and cisplatin + reserpine groups. The quantified result is shown on the right (n = 3). (B) Venn diagram of all RNA_seq genes and ferroptosis genes set (obtained from the FerrDb V2), 466 genes in the intersection were analyzed. Volcano plot showing comparison of expression changes of those 466 ferroptosis-related genes in cisplatin vs vehicle, and cisplatin + reserpine vs cisplatin groups (q-value < 0.05). (C) Heatmap of key ferroptosis-related genes expression in vehicle, cisplatin and cisplatin + reserpine groups analyzed by RNA-seq (n = 3). *p < 0.05.

Figure 6.

Figure 6.

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Reserpine effectively inhibits ferroptosis. (A)Western blot analysis of NRF2, SLC7A11, GPX4 in vehicle, cisplatin and cisplatin + reserpine groups. Quantitative analysis of proteins is shown in (B) (n ≥ 3). HK-2 cells were treated with DMSO/RSL3 0.5 µM (C)/IKE 10 µM (D), combined with reserpine 10 µM or Lip1 0.2 µM for 24 h. Cell viability was determined by CCK-8 assay. Caov3 cells (E) and HT1080 cells (F) were treated with concentration gradients of cisplatin, combined with DMSO or reserpine (10 µM) for 24 h. Cell viability was determined by CCK‐8 assay. Results are expressed as mean ± SD, (n = 3). *p < 0.05, **p < 0.01, ****p < 0.0001, ns, no significance.

Subsequently, we examined whether reserpine possessed a similar ferroptosis-inhibiting function in vitro. We employed ferroptosis inducers, RSL3 and imidazole ketone erastin (IKE), targeting GPX4 or SLC7A11. The experimental results showed that reserpine significantly inhibited ferroptosis induced by RSL3 or IKE in human kidney 2 (HK-2) cells, with similar effects to the ferroptosis inhibitor Lip1(Figure 6C, D). Furthermore, we explored whether reserpine’s ferroptosis-inhibiting function would affect the clinical application of cisplatin in antitumor therapy. Our results showed that as the concentration of cisplatin increased, the viability of Caov3 cells and HT1080 cells gradually decreased, while the combined treatment with reserpine did not affect cell viability (Figure 6E, F), indicating that reserpine does not interfere with the antitumor efficacy of cisplatin.

3.5. Reserpine inhibits cGAS/STING signaling and inflammatory gene expression

Cisplatin is known to cause mitochondrial damage, leading to mitochondrial DNA release that activates the cGAS/STING signaling pathway and subsequent inflammatory gene expression, exacerbating inflammation. Our results indicate that cisplatin treatment promotes the activation of cGAS/STING signaling cascade, and reserpine was found to attenuate the activation of the cGAS/STING signaling and reduce phosphorylation of the downstream target TBK1, IRF3 (Figure 7A,B). Importantly, according to the results of RNA-seq, reserpine also suppressed the expression of type I interferon-regulated genes related to antiviral immune response down-stream of the cGAS/STING pathway (Figure 8). Collectively, these results support reserpine’s role in mitigating cisplatin mediated inflammation during AKI.

Figure 7.

Figure 7.

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Reserpine prevents cGAS/STING pathway activation after cisplatin treatment. Western blot analysis of cGAS, STING, TBK1, p-TBK1, p-IRF3 in vehicle, cisplatin and cisplatin + reserpine groups. Quantitative analysis of proteins is shown on the side (n ≥ 3). Data are presented as mean ± SD. *p < 0.05, **p < 0.01.

Figure 8.

Figure 8.

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Reserpine reduces inflammatory gene expression induced by cisplatin. Heatmap shows cGAS/STING downstream IFN-related inflammatory gene expression in vehicle, cisplatin and cisplatin + reserpine groups analyzed by RNA-seq (n = 3).

4. Discussion

Cisplatin stands as a cornerstone in the arsenal of chemotherapeutic agents due to its broad efficacy against various cancers [37]. However, its clinical utility is often overshadowed by significant adverse effects, notably AKI [38–40]. A key pathological event in cisplatin-induced AKI is the injury to the proximal tubules [12,41]. In clinical settings, cisplatin is frequently administered alongside protocols that employ short-term, low-dose hydration to mitigate such side effects [15,42]. Thus, identifying medications capable of protect renal tubular cells from damage holds substantial clinical value. This study, utilizing cisplatin-induced AKI mouse models, demonstrated that cisplatin significantly impairs renal function and induces renal tubular damage. Treatment with reserpine was found to lower serum BUN and SCr levels, mitigate renal tubular damage, and reduce the expression of the renal injury marker NGAL, indicating reserpine’s protective effect against cisplatin-induced AKI.

Ferroptosis, a recently identified form of programmed cell death characterized by iron-dependent lipid peroxidation, plays a crucial role in cisplatin-induced renal injury [43,44]. This process is primarily driven by glutathione (GSH) depletion, reduced activity of GSH peroxidase 4 (GPX4), and the iron-catalyzed Fenton reaction of Polyunsaturated fatty acids [45–47]. Previous research has highlighted the protective effects of lipid peroxidation inhibitors like Fer-1 and iron chelators such as Deferoxamine in cisplatin-induced AKI [43,48,49], underscoring the significance of ferroptosis in this context. Our findings reveal that reserpine counteracts the cisplatin-induced downregulation of SLC7A11/GPX4, a critical component in cell defense against ferroptosis, suggesting reserpine’s potential to antagonize ferroptosis in vivo. In vitro, reserpine can also effectively antagonize ferroptosis caused by SLC7A11/GPX4 inhibition in HK2 cells.

Moreover, studies have reported the activation of the cGAS/STING signaling pathway and the expression of downstream interferon (IFN)-related inflammatory genes following cisplatin treatment. Notably, reserpine treatment alleviated the activation of cGAS/STING signaling, which correlated with reduced infiltration of neutrophils and macrophages. Given that cGAS/STING can be triggered by mitochondrial damage and that ferroptosis involves mitochondrial dysfunction [29,50,51], it is plausible that cisplatin-induced mitochondrial damage initiates cGAS/STING signaling and oxidative stress, leading to ferroptosis in renal tubular cells [43]. Reserpine’s mechanism of action may involve inhibiting cisplatin-induced ferroptosis via SLC7A11/GPX4, thereby protecting renal tubular cells from death and reducing cGAS/STING activation.

Furthermore, emerging evidence suggests that mitochondrial dysfunction and the activation of the cGAS/STING pathway are prominent features of AKI. Apart from cisplatin-induced AKI, ischemia–reperfusion injury and LPS-induced AKI also involve the activation of cGAS/STING and the occurrence of ferroptosis [52–54]. Additionally, chronic kidney diseases such as diabetic nephropathy and Alport syndrome are also accompanied by the activation of cGAS/STING in the diseases progression [55]. Reserpine exhibited effects of inhibiting cGAS/STING activation and ferroptosis, however, its potential application in the treatment of other specific types of kidney diseases remains to be further explored and validated. Nonetheless, there persist a few constraints and matters that remain to be further studied. Our study demonstrates that pretreatment with reserpine for 1 h can prevent cisplatin-induced kidney injury. However, the question of whether reserpine can still effectively alleviate kidney injury after cisplatin treatment deserves further investigation. Reserpine, while exhibiting potential in alleviating renal injury, also presents potential side effects such as bradycardia and hypotension, which must be taken into account during clinical translation.

In summary, this study demonstrates that reserpine mitigates the cytotoxic effects of cisplatin on renal tubular cells, improves renal function, and decreases kidney inflammation in a mouse model. Reserpine has the potential to serve as a therapeutic adjunct, reducing nephrotoxicity associated with cisplatin chemotherapy while maintaining its anti-tumor efficacy.

Funding Statement

This work was sponsored by Natural Science Foundation of Chongqing, China (CSTB2023NSCQ-MSX0663), National Natural Science Foundation of China (81972564) and Talents Cultivation Project of the Third Military Medical University (XZ-2019-505-010).

Author contributions

Nahua Xu, Hui Li, and Qi Yao designed the research, analyzed and interpreted the acquired data. Nahua Xu, Rong Mu conducted the major experiments, Siyuan Deng and Ye Han performed IHC staining and participated in HK2 cell analysis. Yanyun Shi and Xuemei Fu involved in RNA-seq analysis. Qi Yao, Hui Li, Nahua Xu collaboratively wrote the manuscript. All authors approved the final paper.

Disclosure statement

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

Data availability Statement

The data used to support the findings of this study are included within the article.

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