Ameliorative impact of ezetimibe on gentamicin-induced kidney damage; A histopathological perspective study Journal of Nephropathology

Introduction: Gentamicin, despite its beneficial effects, has significant nephrotoxic effects that are observed in 20% of the patients. Ezetimibe is recognized as an inhibitor of cholesterol absorption. Objectives: The present study aimed to investigate the histopathological effects of ezetimibe on gentamicin-induced kidney damage. Materials and Methods: Twenty male Wistar rats were randomly divided into five groups and treated as the following; group 1 (normal group), group 2 [gentamicin group, daily 80 mg/kg, intra-peritoneal (i.p.) for seven days], group 3-5 (gentamicin 80 + ezetimibe at doses of 2.5, 12.5, and 37.5 mg/kg, respectively). Kidney sections were examined for histopathological parameters including vacuolization of the tubular renal cells, degeneration, necrosis, flattening of the tubular cells and debris in the tubular lumen. Results: Gentamicin injection significantly induced histopathological alterations ( P < 0.05). Ezetimibe therapy significantly decreased the levels of vacuolization, degeneration, necrosis, flattening of the tubular cells and debris in the nephrotoxic rats ( P < 0.05). Conclusion: The results illustrated that treatment with ezetimibe can improve kidney damage caused by gentamicin injection. ABSTRACT Implication for health policy/practice/research/medical education: In an experimental study, we found that treatment with ezetimibe can improve kidney damage caused by gentamicin injection. Please cite this paper as: Emami E, Hasanpour Dehkordi A, Maghsoudi A, Nasri H, Vahedi A. Ameliorative impact of ezetimibe on gentamicin-induced kidney damage; A histopathological perspective study. Nephropathol.


Introduction
Kidney damage can be acute or chronic and is assessed with serum markers such as blood urea nitrogen (BUN) and creatinine levels as well as other biomarkers (1). Serum creatinine and blood urea have been reported to increase 7-10 days after the initiation of treatment with aminoglycosides (2).
Aminoglycoside antibiotics, such as gentamicin are administered to treat gram-negative bacterial infections (3). Gentamicin, despite its beneficial effects, has significant nephrotoxic effects that are observed in 20% of the patients. The toxicity of gentamicin is associated with its accumulation in the renal cortex, especially in the proximal tubules, which induces apoptosis and necrosis in the epithelial cells (4,5). Despite the introduction of less nephrotoxic antibiotics against gram-negative microorganisms, gentamicin is still widely administered due to its low-cost and effectiveness (6). Therefore, a potential therapeutic approach to protect or reverse the effects of gentamicin-induced kidney damage would have Journal of Nephropathology, Vol 11, No 2, April 2022 www.nephropathol.com 2 significant clinical benefits.
The exact mechanism of gentamicin in the development of renal toxicity is unknown; however, reactive oxygen species (ROS) appear to be involved (7)(8)(9)(10)(11). The production of the renal cortical lipoperoxidation and hydrogen peroxide (H 2 O 2 ) has been demonstrated to increase in gentamicin-treated mice (7,8). According to studies, administration of antioxidant enzymes such as superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR) and catalase (CAT) recuperates the severity of gentamicin-induced renal damage. Kidney damage is also reduced by consuming dietary antioxidants, vitamin E and selenium (12,13).
Ezetimibe is an inhibitor of cholesterol absorption. It blocks intestinal absorption of cholesterol and phytosterols through its interaction with sterol transporters, detected as Niemann-Pick C1-like1 protein (NPC1L1) (14)(15)(16). In animal models, the uptake inhibition of intestinal cholesterol by ezetimibe leads to a decrease in cholesterolretaining fat in chylomicrons; therefore, the amount of cholesterol that is transferred by chylomicron to the liver reduces (17). This leads to the rearrangement of hepatic low-density lipoprotein (LDL) receptors which, in turn, has been accompanied by an elevation in LDL-receptor presentation and to a reduction in LDL-cholesterol (18)(19)(20).

Objectives
The present study aimed to investigate the histopathological effects of ezetimibe on gentamicin-induced kidney damage.

Animals and study design
Twenty male Wistar rats weighing 200-250 g were used in the present study. The animals were kept in the Research Center of Medicinal Plants, Shahrekord University of Medical Sciences in a standard condition (temperature 21-25°C and 12-hour cycle of darkness and light) with access to water and food.
The rats were divided into five groups (n=4); group 1(normal group); received no drug. Group 2 received gentamicin at a dose of 80 mg/kg. Group 3 to 5 received gentamicin at a dose of 80 mg/kg and after a one-hour interval, ezetimibe at doses of 2.5, 12.5 and 37.5 mg/kg, respectively. Injections were administered intraperitoneally for seven days.

Histopathological examinations
Ultimately, the kidneys of the animals were isolated and kept in formalin for 12 hours and processed for histopathological examination. The 3 μm-thick paraffin tissues were stained with hematoxylin and eosin (H&E).
All samples were examined for histopathological parameters including vacuolization of the tubular renal cells, degeneration, necrosis, flattening of the tubular cells and debris in the tubular lumen.

Statistical analysis
All data were analyzed through one-way ANOVA and then the post hoc Tukey test using GraphPad Prism version 4.03 and expressed as the mean ± standard error (SE). P < 0.05 was considered statistically significant.

Results
According to the results demonstrated in Figure 1 (A-D), vacuolization, flattening, degeneration and necrosis in the group receiving gentamicin enhanced remarkably compared to the normal group (P < 0.001) The groups receiving ezetimibe had a significant decrease of the above histopathological parameters in all doses compared to the gentamicin group (P < 0.001).
Regarding the debris parameter ( Figure 1E), a significant difference between the gentamicin group and the normal group was detected (P < 0.01). In addition, a significant decrease of debris parameter in the groups receiving

Conclusion
Gentamicin causes kidney damage and toxicity and ezetimibe appears to counteract the effects of gentamicin.
Based on the results, ezetimibe exerts its effects by lowering cholesterol, reducing free radicals and increasing the activity of antioxidants. It seems that the use of ezetimibe in combination with other drugs such as statins can increase its effects on improving renal failure.

Acknowledgments
Authors would like to thank Elham Bijad for statistical analysis.

Authors' contribution
Proposal preparing; HN. Animal lab supervision and headlining; AHD. First edit; EE and AV. ARM and HN both are corresponding authors who edited and finalized the paper equally. All authors read and signed the final paper.

Conflicts of interest
As one of the contributing authors to this study, HN acts as the Editor-in-Chief of the journal. It should be noted that his contribution to this journal has not influenced the peer-review process.

Ethical issues
This study was approved by the Ethics Committee