Gossypetin mitigates doxorubicin-induced nephrotoxicity: A histopathological and biochemical evaluation

1 Introduction

DOX, an efficient chemotherapeutic agent, belongs to the family of anthracycline antibiotics (Ijaz et al., 2023). It is widely used to cure multiple types of malignancies such as lung, ovary, blood and breast cancer (Khames et al., 2019). Despite its major advantages in clinical oncology, it also depicts adverse side effects on healthy cells due to its ability to cause non-targeted toxicity (Moutabian et al., 2022). DOX depicts cellular toxicity by targeting certain molecules in the cell. The anti-cancer property of DOX is due to its intercalation into DNA molecules, leading to inhibition of the synthesis of DNA, RNA and protein (Carvalho et al., 2009). DOX exposure triggers the inhibition of topoisomerase II activity, autophagy dysregulation and redox cycling blockage owing to ROS production (Zilinyi et al., 2018). Moreover, it also instigates pulmonary, testicular, hematological and cardiac damage that restricts its clinical application in the field of oncology (Ayla et al., 2011).

Kidneys are an important organ that regulates homeostasis, acid-base balance, blood pressure, and toxic metabolites (Hertzberg et al., 2017). DOX prompts irreversible damage to the kidney tissues owing to its limited regeneration and healing ability. Although the exact process to trigger nephrotoxicity remains unclear but various studies believed that oxidative (OS) stress mediated by radical formation is the primary reason for kidney damage (Owumi et al., 2021). DOX disrupts cellular activities of the antioxidant profile as well as enhances the MDA level (Wu et al., 2021). Furthermore, escalation in creatinine and urea levels is evidence of progressive kidney damage (Afsar et al., 2020).

Flavonoids, a polyhydroxy phenolic compound, are abundantly found in flowers, wine, grains, fruits and vegetables (Ishtiaq et al., 2022). GTIN is a well-known bioflavonoid that contains 6 OH groups. It can be isolated from Hibiscus sabdariffa and many other flowers. It is evinced to exhibit multiple pharmacological properties such as anti-inflammatory (Lin et al., 2021), antibacterial, antioxidant and anti-cancer (Xie et al., 2019). Hence, the aforementioned therapeutical properties of GTIN open a new avenue for research and impelled us to determine its remedial effects against DOX-instigated renal toxicity.

2 Materials and methods

3 Results

3.5

3.5 Effects of GTIN and DOX on histology

DOX exposure instigated a considerable deterioration as well as detachment of epithelial cells, bowman capsule dilation, disruption of renal tubules and glomerular contraction. Whereas, GTIN + DOX exposure mitigated the above-stated adverse changes in renal morphology in the GTIN + DOX co-administrated group than in DOX group, reflecting the renoprotective role of GTIN. GTIN and control group demonstrated almost the same renal morphology (Fig. 1).

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Fig. 1

Remedial potential of GTIN on DOX instigated renal histological damages. (A) Control group (B) DOX intoxicated group (3 mg/kg) (C) GTIN + DOX co-treated group (30 mg/kg + 3 mg/kg) (D) GTIN supplemented group (30 mg/kg). RT: Renal tubules.

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4 Discussion

According to our results, the biochemical analysis illustrated a profound decline in the profile of GPx, GSR, SOD, CAT, GST, and GSH accompanied by an increment in ROS and MDA levels after DOX administration. The instability in ROS level and antioxidant profile prompts OS that ultimately triggers oxidative impairment (Aziz et al., 2021; Mustafa et al., 2022). Moreover, MDA escalation depicts lipid peroxidation. SOD binds with two oxygen radicals to form hydrogen peroxide, while CAT converts H₂O₂ to oxygen and H₂O by inhibiting the generation of cytotoxic ions, (OH^–) (Ighodaro and Akinloye, 2018). GPx also mitigates OS by reducing hydrogen peroxide and lipid peroxide levels (Safhi, 2018). Hence, it is imperative to alleviate ROS level by regulating the normal antioxidant profile that subsequently declines renal OS. GTIN supplementation significantly escalated the antioxidant enzymes activities accompanied by an alleviation in ROS and MDA levels. The outcomes of our investigation were in compliance with Samant and Gupta (2022), who highlighted that the antioxidant ability and free radical scavenging property of GTIN in neurons that contribute to regaining the enzymatic profile and ROS and MDA level.

According to the current study, DOX intoxication augmented the urea, creatinine, KIM-1, as well as NGAL level, while a phenomenal reduction was detected in creatinine clearance level. Urea and creatinine, are used to monitor renal functions (Sahu et al., 2020). Creatinine is formed by the breakdown of phosphocreatine catalyzed by creatine kinase and it is removed from the kidney through glomerular filtration (Sepulveda, 2019). Any oxidative damage to renal tissues escalated the urea and creatinine level accompanied by lower creatinine clearance. Moreover, KIM-1, a transmembrane protein, does not normally present in urine but its presence implies injury in proximal tubules (Dobrek et al., 2016). NGAL is also seen in urine, kidney tubules, and blood following renal damage (Mori et al., 2005). However, co-administration of GTIN + DOX lowered the urea, creatinine, KIM-1, as well as NGAL levels that is probably associated with an improved glomerular filtration rate reflected by higher creatinine clearance.

DOX administration augmented inflammatory responses that are associated with the increased inflammatory mediator levels (IL-6, NF-κB, IL-1β, TNF-α along with COX-2). NF-κB, a transcription factor, is activated by various stresses (oxidative agents) in the cell leads to the regulation of a variety of proinflammatory mediators i.e., COX-2 and TNF-α. A previous study about DOX exposure on renal tissues depicted a remarkable increase in NF-κB expression, which ultimately boosted the release of other inflammatory markers (Hussain et al., 2021). According to our findings, GTIN treatment suppressed the level of inflammatory markers. Our results are further endorsed by earlier study, which predicted that structural features of flavonoids might be responsible to suppress inflammation (Nile et al., 2018).

DOX intoxication elicited a considerable augmentation in Bax, Caspase-9 along with Caspase-3 levels, while lessened the Bcl-2 level. Bax, Caspase-9 and Caspase-3, (pro-apoptotic markers), are the key indicator of apoptosis compared with Bcl-2, which is an anti-apoptotic marker. Cellular OS stimulates the caspase-3 level in the tissues, that promotes the apoptotic process and results in apoptotic damage (Liu et al., 2012). Moreover, disturbance in Bax as well as Bcl-2 level encourage cytochrome c eviction from mitochondrial membrane into the cytoplasm. Cytochrome c eviction promotes the stimulation of Caspase-9 and Caspase-3, which ultimately prompts apoptosis (Katiyar et al., 2005). GTIN + DOX co-administration mitigated all the above said irregularities by upregulating Bcl-2 and lowering Caspase-9, Bax and Caspase-3 levels owing to its anti-apoptotic potential.

Histomorphological observation of DOX-administrated rats showed deterioration and detachment of epithelial cells, glomerular contraction, degeneration of tubules and dilation of the bowman capsule. These alternations promoted renal impairment associated with the production of free radicals-induced lipid oxidation in renal tissues. Our findings were analogous to Mohamed et al. (2022), who illustrated glomerular contraction, necrosis, vacuolization, epithelial cell detoriations and bowman capsule dilation followed by DOX exposure. However, GTIN supplementation effectively mitigated nephrotoxicity and morphological damages prompted by DOX in renal tissues. The renoprotective properties of GTIN might be directly linked with its free radical scavenging potential and anti-inflammatory potential.

5 Conclusion

In conclusion, GTIN treatment elicited excellent attenuative effects against DOX nephrotoxicity. GTIN holds some promising therapeutic strategies to prevent any changes in the level of renal biomarkers, oxidative markers, apoptotic markers, inflammatory markers, antioxidants activity as well as histological architecture of renal tissues. Collectively, it can be concluded that GTIN may have clinical prospects in future to treat renal dysfunction in humans and other animals.

Funding

The authors extend their appreciation to the Deputyship of Research and Innovation, ‘Ministry of Education’ in Saudi Arabia for funding this research (IFKSUOR3-279-1).

Author contribution

MUI, KA and HAK designed the study, conceived the study, and analyzed the results. KA and MUI conceived an initial part of the study, performed the experiment, histology and helped in compiling the results. SR, AA, MI, HA, and TA helped in writing the results. MUI, SR, KZ, MI, TA, HA, and AA made a substantial contribution in the interpretation of data and revising the manuscript for intellectual content. All authors read and approved the final manuscript.