Immunomodulatory, antioxidant, and anti-apoptotic effect of Solidago virgaurea extract against experimental Doxorubicin-induced nephrotoxicity

Authors

Abstract

Aim DOX is a widely used chemotherapeutic agent, but its severe nephrotoxic effects limit its clinical use. This study evaluates the potential protective effects
of Solidago virgaurea extract against DOX-induced kidney damage in rats.
Materials and Methods Animals were classified into four groups, with 8 rats in each group: control, extract-only, DOX, and DOX plus extract (DOX + Solidago virgaurea extract).
Results After 14 days, DOX administration led to significant renal impairment, demonstrated by elevated serum Cr and BUN levels, along with histopathological disruption. DOX also induced oxidative stress by increasing MDA levels and reducing TAC. Furthermore, it triggered renal inflammation by upregulating pro-inflammatory cytokines such as TNF-α, IL-6, IL-1β, while downregulating anti-inflammatory cytokines (IL-10, TGF-β, IL-35). Additionally, DOX promoted renal apoptosis by increasing apoptotic markers (Caspase-3, BAX), accompanied by a decrease in anti-apoptotic BCL-2 protein levels. Co- treatment with Solidago virgaurea extract effectively mitigated these adverse effects by reversing the previous parameters. Histopathological analysis further confirmed preserved renal architecture compared to the DOX-only group.
Discussion These results concluded that Solidago virgaurea extract exhibits strong nephroprotective potential, making it a promising adjunctive therapy for preventing chemotherapy-induced kidney injury.

Keywords

Introduction

The therapeutic use of chemotherapeutic agents frequently leads to systemic toxicity, affecting multiple organ systems, particularly renal impairment [1]. Drug-induced renal damage manifests clinically as nephrotoxicity or nephrotic syndrome, impairing renal functions such as filtration, reabsorption, and excretion [2]. Current data indicate that approximately 60% of chemotherapy patients suffer from nephrotoxicity, contributing to drug-associated morbidity and mortality [3].
Among chemotherapeutic agents, DOX, a widely used anthracycline, remains crucial in treating multiple malignancies [4]. However, its clinical efficacy is limited by two major drawbacks: non-selective cytotoxicity, which affects healthy tissues, and the development of drug resistance in tumor cells [4, 5]. The renal system is particularly susceptible to DOX- induced toxicity, with multiple studies confirming its nephrotoxic potential [5, 6]. The underlying mechanisms include oxidative stress, apoptotic pathway activation, and inflammatory cascade induction, with key mediators such as NF-κB, TNF-α, and IL-1β playing pivotal roles in initiating and exacerbating renal injury [7]. These findings highlight the urgent need for adjunctive protective agents to mitigate DOX-induced renal damage during cancer treatment [5].
Solidago virgaurea L., commonly known as European goldenrod, is a perennial medicinal plant belonging to the Asteraceae family, with a broad geographical distribution across Europe, North Africa, and parts of Asia, including China and India [8]. Ethnopharmacological studies report its traditional use in treating urological conditions, such as kidney stones and prostate disorders [8, 9]. Recent pharmacological research has identified several biological activities, including antimicrobial, anti-inflammatory, analgesic, and anticancer effects [10]. This study aimed to comprehensively investigate its nephroprotective potential against DOX-induced renal injury. Using an experimental model, we specifically evaluated the extract’s ability to preserve renal function and architecture while mechanistically targeting oxidative stress, inflammatory mediators (NF-κB, TNF-α, IL-1β), and apoptotic pathways. To our knowledge, this constitutes the first report elucidating S. virgaurea’s multi-targeted renoprotective mechanisms against DOX toxicity, thereby bridging its ethnopharmacological use with evidence-based therapeutic potential.

Materials and Methods

Experimental Animals
Thirty-two male Wistar rats (160 ± 30 g) were housed in standard laboratory cages under controlled environmental conditions, with a temperature of 22 ± 2°C, humidity levels of 50–60%, and a 12-hour light/dark cycle. Throughout the study, the animals had ad libitum access to standard rodent chow and water. Following a 14-day acclimatization period.
Experimental Design
The rats were randomly divided into four equal groups (n = 8 per group). Group 1 (Control) received oral saline administration for 10 days, followed by 4 additional days after a single Intraperitoneal (i.p.) injection of saline. Group 2 (Extract Only) was administered Solidago virgaurea hydroalcoholic extract (250 mg/kg/day orally; purchased from Atos Pharma, Cairo, Egypt, ready for use) for 14 consecutive days, with the dose selected based on previous research [11]. Group 3 (DOX Group) received a single nephrotoxic dose of DOX (20 mg/kg i.p.; Sigma-Aldrich Co., St. Louis, MO, USA) to induce kidney damage according to established protocols [12]. Group 4 (Extract + DOX) received pretreatment with the extract (250 mg/kg/day orally) for 10 days, followed by DOX administration (20 mg/kg i.p.) on day 11, and then continued extract treatment for an additional 4 days.
Specimen Sampling
Following an overnight fast, the animals were anesthetized with thiopental sodium (75 mg/kg body weight, i.p.). Blood samples were collected via retro-orbital puncture and allowed to clot at room temperature before centrifugation (3,000 rpm for 10 minutes). The serum was separated, aliquoted, and stored at -20°C for subsequent analysis of BUN and Cr levels. Laparotomy was then performed to excise both kidneys. The right kidneys were immediately fixed in 10% neutral buffered formalin for 2 hours for histopathological examination, while the left kidneys were homogenized (10% w/v) in ice-cold Tris- HCl buffer (50 mM, pH 7.4) containing 300 mM sucrose. The homogenates were centrifuged (4,000 rpm for 15 minutes at 4°C), and the resulting supernatants were stored at -80°C for subsequent biochemical analyses.
Measurement of Serum Renal Function Markers
The collected serum samples were stored at -20°C until analysis. BUN and Cr levels were quantified using standardized commercial assay kits (SPECTRUM Diagnostics, Egyptian Company for Biotechnology, Cairo, Egypt) according to the manufacturer’s protocols.
Oxidative Stress Biomarker Analysis
Lipid peroxidation was assessed by measuring MDA levels using the method of Ohkawa et al. [13]. Briefly, TBARS were quantified spectrophotometrically at 534 nm following reaction at 95°C in an acidic medium. TAC was evaluated using Bio- diagnostics kits according to the method of Koracevic et al. [14], which measures residual hydrogen peroxide after its reaction with sample antioxidants through enzymatic conversion to a chromophore (absorbance measured at 505 nm).
Analysis of Inflammatory Cytokines and Apoptotic Markers Renal tissue homogenates were analyzed for proinflammatory cytokines TNF-α (Cat# ab46070, Abcam, Cambridge, UK), IL-6 (Cat# ab100772, Abcam, Cambridge, UK), IL-1β (Cat# ab255730, Abcam, Cambridge, UK), anti-inflammatory cytokines IL-10 (Cat# ab100765, Abcam, Cambridge, UK), IL-35 (Cat# MBS022464, MyBioSource, San Diego, CA, USA), TGF-β (Cat# BMS623-3, Thermo Fisher Scientific, Waltham, MA, USA), and apoptotic markers caspase-3 (Cat# ER0143, Wuhan Fine Biotech Co.), BAX (Cat# MBS2512405, MyBioSource, San Diego, CA, USA), BCL-2 (Cat# CSB-E08854r, Cusabio, Houston, TX, USA) using commercially available ELISA kits according to the manufacturers’ protocols.
Histopathological Examination
Fixed right kidney samples (preserved in 10% neutral buffered formalin) were processed using standard paraffin embedding. Serial 5-μm sections were stained with Hematoxylin and Eosin (H&E) for light microscopic evaluation of renal architecture and pathological alterations.
Statistical Analysis
All data are expressed as mean ± SD. Intergroup comparisons were analyzed using one-way ANOVA followed by Tukey’s post hoc test (GraphPad Prism software, version 8; GraphPad Software, San Diego, CA, USA). Statistical significance was defined as p < 0.05.
Ethical Approval
This study was approved by the Ethics Committee of Umm AL- Qura University, Makka, Saudi Arabia (Date: 2025-01-19, No: HAPO-02-K-012-2025-06-2819).

Results

Nephroprotective Effect of Solidago Virgaurea Extract on DOX- Induced Renal Impairment and Histological Injury
As illustrated in Figure 1, a single i.p. injection of DOX led to a significant increase (p < 0.05) in renal function markers Cr and BUN compared to the control group (Fig. 1A, B). These biochemical changes were associated with distinct histological alterations, including severe interstitial inflammation and proximal tubular degeneration, characterized by cast formation and epithelial cell damage, leading to tubular lumen obstruction (Fig. 2E). In contrast, renal sections from control animals exhibited normal glomerular and tubular morphology (Fig. 1C, D).
However, co-administration of Solidago virgaurea extract alongside DOX significantly mitigated renal dysfunction and histological damage. Treated animals showed notable improvements in renal function tests, accompanied by only mild interstitial inflammation (Fig. 1F). The extract provided nephroprotective benefits by restoring renal architecture and minimizing DOX-induced nephrotoxicity.
Antioxidant Effect of Solidago Virgaurea Extract Against DOX- Induced Renal Oxidation
Compared to the control group, i.p. injection of DOX significantly increased the lipid peroxidation indicator (MDA) while decreasing the TAC in the renal supernatant (Fig. 2A, B). Conversely, co-treatment with Solidago virgaurea extract significantly reduced MDA levels and elevated TAC levels. These findings demonstrate that Solidago virgaurea possesses potent antioxidant properties.
Immunomodulatory Impact of Extract Against DOX-Induced Renal Inflammation
As shown in Figure 4, the DOX-treated rats revealed a marked elevation (p < 0.05) in proinflammatory cytokines (TNF-α, IL-6, IL-1β; Fig. 2C–E) alongside a marked reduction in anti-inflammatory mediators (IL-10, IL-35, TGF-β; Fig. 2F–H) compared to the control group. In contrast, co-treatment with Solidago virgaurea extract demonstrated immunomodulatory effects by suppressing proinflammatory markers and elevating anti-inflammatory cytokine levels relative to rats treated with DOX alone. These results highlight the potent anti-inflammatory properties of Solidago virgaurea in mitigating DOX-induced renal inflammation.
Anti-Apoptotic Effect of Solidago Virgaurea Extract
Figure 6 illustrates the impact of DOX treatment on apoptotic protein expression in renal tissue. Compared to the control rats, DOX-treated rats exhibited an elevation in apoptotic markers caspase-3 and BAX, accompanied by a notable decline in the anti-apoptotic protein BCL-2. However, co-administration of Solidago virgaurea extract alongside DOX significantly mitigated apoptosis by reducing caspase-3 and BAX levels while promoting the expression of BCL-2. These findings suggest that Solidago virgaurea extract exerts a protective effect against DOX-induced renal tubular apoptosis, reinforcing its potential therapeutic benefits.

Discussion

Nephrotoxicity is considered one of the main side effects of DOX administration, primarily attributed to oxidative damage [15], as well as its direct degenerative effects due to accumulation in renal tissues [16]. The main objective of our study was to examine the protective role of Solidago virgaurea as a natural extract against DOX-induced renal injury, elucidating its mechanism of action through the mitigation of oxidative stress, renal inflammation, and tubular apoptosis, with a focus on histological improvement in renal tissue.
DOX-related nephrotoxicity was confirmed biochemically by elevated serum levels of Cr and BUN, reflecting renal injury and glomerular dysfunction. This can be attributed to impaired glomerular filtration caused by DOX administration, as supported by multiple studies [1, 17]. These studies documented that DOX metabolites exert harmful effects on the nephron, directly impacting glomerular filtration or indirectly through oxidative stress induced by DOX.
Histological examination further confirmed nephrotoxicity, revealing tubular injury and vascular congestion. These findings align with the results of our study, in which a single i.p. injection of DOX on day 11 caused a significant rise in renal dysfunction markers and altered renal morphology.
Conversely, pretreatment with the Solidago virgaurea extract before DOX injection, followed by continued administration for 14 days, markedly reduced renal injury indicators and improved renal histopathology. This is consistent with the findings of Rahimi et al. [18], who reported that Solidago canadensis L. hydroalcoholic extract mitigated paracetamol- induced renal toxicity by improving renal function and reducing histopathological damage, including hyperemia, vacuolation, and inflammatory cell infiltration.
In our study, DOX injection induced significant oxidative stress in renal tissues, as evidenced by increased MDA and a marked reduction in TAC. These findings align with a previous study [19]. DOX metabolism generates free radicals, leading to lipid peroxidation of the glomerular epithelial lining. This process damages glomerular membranes, impairing their function and disrupting podocyte activity [19].
In contrast, concurrent oral administration of Solidago virgaurea extract with DOX significantly reduced oxidative stress markers (such as MDA) while increasing TAC. These results are consistent with the findings of El-Tantawy et al. [11], who demonstrated the protective effects of S. virgaurea extract against isoproterenol-induced cardiotoxicity in a rat model. The extract reduced renal oxidative stress by decreasing MDA and nitric oxide levels while elevating GSH, SOD, and vitamin C. Cytokines are well-known proteins that play a crucial role in immune responses and are involved in the pathogenesis of DOX toxicity [20]. This work demonstrated the role of proinflammatory cytokines in the progressive course of DOX- induced nephrotoxicity. In contrast, anti-inflammatory cytokines contribute to immune modulation during nephrotoxicity [21]. Our results revealed that DOX significantly increased proinflammatory cytokine levels, consistent with Ibrahim et al. [20], while decreasing anti-inflammatory cytokines, aligning with Atta et al. [22], who reported reduced IL-10 and TGF-β in a rat model of nephrotoxicity. However, co-administration with S. virgaurea extract modulated the immune response, reversing cytokine levels by elevating IL-35, IL-10, TGF-β, and suppressing IL-6, IL-1β, and TNF-α.
These findings agree with a previous study by Sanad et al. [23], which highlighted the antidiabetic role of S. virgaurea extract through its reduction of serum TNF-α. The anti-inflammatory activity of S. virgaurea may be attributed to its caffeoylquinic acid derivatives, which display powerful anti-inflammatory effects against carrageenan-induced ankle edema by inhibiting IL-1β and TNF-α [24].
DOX has been shown to play a significant role in stimulating renal tubular apoptosis by upregulating caspase-3 in proximal tubular epithelial cells [25]. The activation of caspase-3 in renal tissue following DOX treatment indicates caspase-dependent apoptosis, as evidenced by its presence in damaged glomeruli, tubules, and interstitial cells [19]. Our findings revealed that the DOX group exhibited a marked increase in caspase-3 and BAX expression, along with a decline in BCL-2 levels, consistent with a previous study [25]. However, co-administration of Solidago virgaurea extract with DOX significantly reduced renal apoptosis by modulating these apoptotic markers. The anti- apoptotic effect of S. virgaurea extract can be attributed to its potent antioxidant and anti-inflammatory properties.

Limitations

One of the primary limitations of this study is the focus on the short-term effects of Solidago virgaurea extract against DOX-induced nephrotoxicity. While the results indicate promising protective effects, long-term evaluations, including chronic administration and delayed outcomes, are necessary to confirm sustained benefits. Additionally, the availability of primary immune antibodies for staining inflammatory and apoptotic markers in renal tissue was limited, which may have constrained a more detailed immunohistochemical analysis. Furthermore, the restricted access to gene expression analysis via PCR for apoptotic markers due to resource limitations posed a challenge in fully elucidating the molecular mechanisms underlying renal protection. Future studies addressing these constraints with extended experimental timelines, enhanced molecular investigations, and improved access to analytical tools will provide deeper insights into the nephroprotective potential of Solidago virgaurea.

Conclusion

Our study demonstrates that Solidago virgaurea extract effectively protects against DOX-induced nephrotoxicity through multiple mechanisms. DOX administration caused significant renal dysfunction, oxidative stress (elevated MDA, reduced TAC), inflammation (increased TNF-α, IL-6, IL-1β; decreased IL-10, TGF-β, IL-35), and apoptosis (upregulated caspase-3/ BAX, downregulated BCL-2). Co-treatment with S. virgaurea extract reversed these effects by improving renal function tests, reducing oxidative damage, restoring cytokine balance, and inhibiting apoptotic pathways. Histopathological findings confirmed the extract’s protective effects, showing preserved renal architecture and reduced tubular damage. Overall, your study highlights S. virgaurea’s potential as a therapeutic agent against chemotherapy-induced kidney injury, mediated by its antioxidant, anti-inflammatory, and anti-apoptotic properties. The extract’s bioactive compounds may offer a promising adjunct therapy to mitigate DOX nephrotoxicity.

References

1. Alibakhshi T, Khodayar MJ, Khorsandi L, Rashno M, Zeidooni L. Protective effects of zingerone on oxidative stress and inflammation in cisplatin-induced rat nephrotoxicity. Biomed Pharmacother. 2018;105:225-32.
   Article PubMed Google Scholar
2. Nussbaum EZ, Perazella MA. Update on the nephrotoxicity of novel anticancer agents. Clin Nephrol. 2018;89(3):149-65.
   Article PubMed Google Scholar
3. Fukasawa H, Furuya R, Yasuda H, Yamamoto T, Hishida A, Kitagawa M. Anti-cancer agent-induced nephrotoxicity. Anticancer Agents Med Chem. 2014;14(7):921-7.
   Article PubMed Google Scholar
4. Sritharan S, Sivalingam N. A comprehensive review on time-tested anticancer drug doxorubicin. Life Sci. 2021;278:119527.
   Article PubMed Google Scholar
5. Prša P, Karademir B, Biçim G, et al. The potential use of natural products to negate hepatic, renal and neuronal toxicity induced by cancer therapeutics. Biochem Pharmacol. 2020;173:113551.
   Article PubMed Google Scholar
6. Rafiee Z, Moaiedi MZ, Gorji AV, Mansouri E. P-Coumaric Acid Mitigates Doxorubicin-Induced Nephrotoxicity Through Suppression of Oxidative Stress, Inflammation and Apoptosis. Arch Med Res. 2020;51(1):32-40.
   Article PubMed Google Scholar
7. Shams S, Lubbad LI, Simjee SU, Jabeen A. N-(2-hydroxy phenyl) acetamide ameliorate inflammation and doxorubicin-induced nephrotoxicity in rats. Int Immunopharmacol. 2023;123:110741.
   Article PubMed Google Scholar
8. Demir H, Acik L, Bali EB, Koç LY, Kaynak G. Antioxidant and antimicrobial activities of Solidago virgaurea extracts. Afr J Biotechnol. 2009;8(2):274-9.
   PubMed Google Scholar
9. Gross SC, Goodarzi G, Watabe M, Bandyopadhyay S, Pai SK, Watabe K. Antineoplastic activity of Solidago virgaurea on prostatic tumor cells in an SCID mouse model. Nutr Cancer. 2002;43(1):76-81.
   Article PubMed Google Scholar
10. Thiem B, Goślińska O. Antimicrobial activity of Solidago virgaurea L. from in vitro cultures. Fitoterapia. 2002;73(6):514-6.
    Article PubMed Google Scholar
11. El-Tantawy WH. Biochemical effects of Solidago virgaurea extract on experimental cardiotoxicity. J Physiol Biochem. 2014;70(1):33-42.
    Article PubMed Google Scholar
12. Ayla S, Seckin I, Tanriverdi G, et al. Doxorubicin induced nephrotoxicity: protective effect of nicotinamide. Int J Cell Biol. 2011;2011:390238.
    Article PubMed Google Scholar
13. Ohkawa H, Ohishi N, Yagi K. Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Anal Biochem. 1979;95(2):351-8.
    Article PubMed Google Scholar
14. Koracevic D, Koracevic G, Djordjevic V, Andrejevic S, Cosic V. Method for the measurement of antioxidant activity in human fluids. J Clin Pathol. 2001;54(5):356-61.
    Article PubMed Google Scholar
15. Afsar T, Razak S, Almajwal A, Al-Disi D. Doxorubicin-induced alterations in kidney functioning, oxidative stress, DNA damage, and renal tissue morphology; improvement by Acacia hydaspica tannin-rich ethyl acetate fraction. Saudi J Biol Sci. 2020;27(9):2251-60.
    Article PubMed Google Scholar
16. Injac R, Boskovic M, Perse M, et al. Acute doxorubicin nephrotoxicity in rats with malignant neoplasm can be successfully treated with fullerenol C60(OH)24 via suppression of oxidative stress. Pharmacol Rep. 2008;60(5):742-9.
    PubMed Google Scholar
17. Kuzu M, Yıldırım S, Kandemir FM, et al. Protective effect of morin on doxorubicin-induced hepatorenal toxicity in rats. Chem Biol Interact. 2019;308:89-100.
    Article PubMed Google Scholar
18. Rahimi O, Asadi Louie N, Salehi A, Faed Maleki F. Hepatorenal protective effects of hydroalcoholic extract of Solidago canadensis L. against paracetamol- induced toxicity in mice. J Toxicol. 2022;2022:9091605.
    Article PubMed Google Scholar
19. Asci H, Saygin M, Cankara FN, et al. The impact of alpha-lipoic acid on amikacin-induced nephrotoxicity. Ren Fail. 2015;37(1):117-21.
    Article PubMed Google Scholar
20. Ibrahim Fouad G, Ahmed KA. The protective impact of berberine against doxorubicin-induced nephrotoxicity in rats. Tissue Cell. 2021;73:101612.
    Article PubMed Google Scholar
21. Albukhari TA, Bagadood RM, Bokhari BT, et al. Chrysin attenuates gentamicin-induced renal injury in rats through modulation of oxidative damage and inflammation via regulation of Nrf2/AKT and NF-kB/KIM-1 pathways. Biomedicines. 2025;13(2):271.
    Article PubMed Google Scholar
22. Atta AH, Atta SA, Khattab MS, et al. Ceratonia siliqua pods (Carob) methanol extract alleviates doxorubicin-induced nephrotoxicity via antioxidant, anti- inflammatory and anti-apoptotic pathways in rats. Environ Sci Pollut Res Int. 2023;30(35):83421-38.
    Article PubMed Google Scholar
23. Sanad FA, Ahmed SF, El-Tantawy WH. Antidiabetic and hypolipidemic potentials of Solidago virgaurea extract in alloxan-induced diabetes type 1. Arch Physiol Biochem. 2022;128(3):716-23.
    Article PubMed Google Scholar
24. Abdel Motaal A, Ezzat SM, Tadros MG, El-Askary HI. In vivo anti-inflammatory activity of caffeoylquinic acid derivatives from Solidago virgaurea in rats. Pharm Biol. 2016;54(12):2864-70.
    Article PubMed Google Scholar
25. Su Z, Ye J, Qin Z, Ding X. Protective effects of madecassoside against doxorubicin induced nephrotoxicity in vivo and in vitro. Sci Rep. 2015;5:18314.
    Article PubMed Google Scholar

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