Histopathological Patterns of Renal Injury in Gentamicin-Exposed Wistar Rats Following Turmeric Essential Oil and Losartan Treatment

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Abstract

Introduction: Gentamicin-induced nephrotoxicity is characterized by proximal tubular degeneration, necrosis, inflammatory infiltration, vascular congestion, and interstitial changes. Turmeric-derived preparations and losartan have been investigated as potential modifiers of renal injury, although direct histopathological evidence concerning turmeric essential oil remains limited. Aim: To describe and compare renal histopathological patterns in gentamicin-exposed Wistar rats treated with turmeric essential oil or losartan. Methods & Materials: This experimental animal study was conducted at the Department of Pharmacology and Therapeutics, Sir Salimullah Medical College, Dhaka, Bangladesh, from July 2019 to June 2020. Twenty-five adult male Wistar rats were randomly allocated to five groups of five animals. Group A received normal saline. Group B received gentamicin 80 mg/kg/day intraperitoneally on days 8-14. Groups C and D received turmeric essential oil at 50 and 100 mg/kg/day orally, respectively, for 14 days, together with gentamicin on days 8-14. Group E received losartan 10 mg/kg/day orally for 14 days, together with gentamicin on days 8-14. Renal sections were stained with haematoxylin and eosin and assessed for inflammatory-cell infiltration, vascular congestion, proximal tubular injury, epithelial regeneration, and interstitial vacuolation using ordinal grades from 0 to 4. Results: The gentamicin-treated group showed inflammatory-cell infiltration grade 4, vascular congestion grade 3, proximal tubular injury grade 3, and interstitial vacuolation grade 2. Tubular necrosis and desquamation involved more than 50% of proximal tubules, and regeneration was absent. With turmeric essential oil at 50 mg/kg/day, inflammation was grade 2, while congestion, tubular injury, and vacuolation were grade 1. With turmeric essential oil at 100 mg/kg/day and losartan, all four injury domains were grade 1. Tubular epithelial regeneration was present in all three intervention groups. Conclusion: Turmeric essential oil and losartan were associated with lower renal histopathological lesion grades than gentamicin alone. The findings were descriptive and did not establish statistical superiority or equivalence between treatments. Assistant Professor, Department of Pharmacology, Jamalpur Medical College, Jamalpur, Bangladesh mazpramanik@gmail.com (ORCID: 0009-0006-2838-8790) Lecturer, Department of Pharmacology, Jamalpur Medical College, Jamalpur, Bangladesh Assistant Professor (In situ), Department of Pharmacology, Jamalpur Medical College, Jamalpur, Bangladesh INTRODUCTION Despite its recognized nephrotoxicity, gentamicin remains widely used against serious Gram-negative infections because of its rapid bactericidal activity, clinical familiarity, and relatively low cost. Following glomerular filtration, gentamicin is taken up preferentially by proximal tubular epithelial cells, where intracellular accumulation is associated with lysosomal dysfunction, mitochondrial injury, reactive oxygen species generation, inflammatory signalling, apoptosis, and necrosis [1]. Structurally, gentamicin-associated injury is expressed predominantly in the renal cortex and includes tubular epithelial swelling, vacuolation, degeneration, desquamation, necrosis, luminal cellular debris, interstitial inflammatory-cell infiltration, vascular congestion, and subsequent regenerative activity. In Wistar rats, progressive cortical tubular degeneration, necrosis, cytolysis, desquamation, mesangial alterations, and capillary congestion were observed with increasing gentamicin dose and duration [2]. Likewise, progression from focal inflammatory-cell collections and tubular basophilia to more evident degeneration, necrosis, casts, and interstitial inflammation was documented after repeated gentamicin exposure [3]. More recently, tubular degeneration, necrosis, inflammatory infiltration, congestion, and interstitial changes have continued to be used as principal structural indicators of gentamicin-associated injury, often alongside molecular and urinary biomarkers [4,5]. Consequently, histopathological examination provides direct evidence of lesion location, type, and severity and remains central to the evaluation of structural renal injury. Unlike isolated serum measurements, tissue examination can distinguish predominantly tubular, interstitial, vascular, or glomerular involvement and can document coexisting degeneration and regeneration within the same experimental model. Because oxidative stress and inflammation are closely associated with gentamicin-induced tubular damage, turmeric-derived preparations have been investigated as potential modifiers of renal tissue injury. Following turmeric administration in gentamicin-exposed rats, lower histopathological injury grades and less severe tissue disruption were reported than with gentamicin alone [6]. Similarly, reductions in tubular degeneration, necrosis, inflammatory abnormalities, and apoptosis were observed with curcumin treatment in experimental gentamicin nephrotoxicity [7]. Additionally, mitochondrial integrity was preserved, and gentamicin-induced renal cortical injury was attenuated following curcumin administration [8]. More recently, conventional and liposomal curcumin were both associated with improved renal cytoarchitecture, while formulation-dependent differences in histopathological protection were identified [9]. Furthermore, dose-related reductions in renal injury, oxidative stress, endoplasmic reticulum stress, and apoptosis were reported with curcumin pretreatment, although incremental benefits became smaller at higher doses [10]. Nevertheless, turmeric essential oil is chemically distinct from whole turmeric and curcumin-rich preparations. Within the oil, volatile sesquiterpenes, particularly ar-turmerone, α-turmerone, and β-turmerone, predominate, whereas curcuminoids are generally absent or present only in negligible quantities [11]. In a cadmium-induced rat model, turmeric essential oil was associated with improved renal-function indicators and modulation of inflammatory cytokines, supporting possible renal activity of the oil itself, although gentamicin-associated renal histopathology was not examined [12]. Alongside botanical interventions, losartan has been evaluated as a pharmacological comparator. Reduced renal damage was reported when losartan at 10 mg/kg/day was administered with gentamicin at 80 mg/kg/day, whereas worsening nephrotoxicity was reported with a substantially higher losartan dose [13,14]. Thus, evidence concerning losartan appears dependent on dose and experimental conditions. Given the limited preparation-specific evidence for turmeric essential oil and the scarcity of direct histopathological comparisons with losartan, the present study described renal lesion patterns across normal control, gentamicin-exposed, turmeric-essential-oil-treated, and losartan-treated Wistar rats. METHODS & MATERIALS Study design and setting Between July 2019 and June 2020, a controlled experimental animal study was conducted in the Department of Pharmacology and Therapeutics, Sir Salimullah Medical College, Dhaka, in collaboration with the Institute of Nutrition and Food Science, University of Dhaka. Ethical approval was obtained from the Institutional Ethics Committee of Sir Salimullah Medical College, Dhaka. Experimental animals and sample size A total of 25 healthy adult male Wistar albino rats (Rattus norvegicus), aged approximately 10-12 weeks and weighing 150-200 g, were obtained from the animal house of the Department of Pharmacology, Bangabandhu Sheikh Mujib Medical University, Dhaka. By applying the resource-equation method, the error degrees of freedom were calculated as E=N−G=25−5=20, where N represented the total number of animals and G represented the number of experimental groups. Following a seven-day acclimatization period, the animals were randomly allocated to five groups of five rats each. Throughout the experiment, the rats were maintained in labelled metallic cages measuring approximately 38×23×15 cm, with each experimental group housed separately. Under a 12-hour light and 12-hour dark cycle, the animal room was maintained at approximately 26-28°C. Standard laboratory feed and drinking water were provided ad libitum. Regular cage cleaning, waste removal, and daily observation were undertaken throughout the study. Preparation of turmeric essential oil Fresh turmeric (Curcuma longa) rhizomes were collected locally and authenticated by the Institute of Nutrition and Food Science, University of Dhaka. After washing, surface drying, and slicing, 500 g of rhizomes were mixed with 1 L of distilled water and subjected to hydrodistillation at approximately 105°C for four hours. Subsequently, the essential oil was separated from the upper layer of the distillate, transferred to a tightly sealed amber bottle, and stored in a dark place until administration. Experimental interventions All oral interventions were administered once daily through an orogastric tube. To induce renal injury, gentamicin was administered intraperitoneally at 80 mg/kg/day from days 8 to 14 in Groups B, C, D, and E. Details of the experimental groups and treatment schedules are presented in (Table I). Table I: Distribution of experimental rats and treatment schedule Group Experimental designation n Treatment during days 1-7 Treatment during days 8-14 A Normal control 5 Normal saline, 20 mL/kg/day orally Normal saline, 20 mL/kg/day orally B Gentamicin-treated control 5 No active treatment Gentamicin, 80 mg/kg/day intraperitoneally C Turmeric essential oil, 50 mg/kg group 5 Turmeric essential oil, 50 mg/kg/day orally Turmeric essential oil, 50 mg/kg/day orally, plus gentamicin, 80 mg/kg/day intraperitoneally D Turmeric essential oil, 100 mg/kg group 5 Turmeric essential oil, 100 mg/kg/day orally Turmeric essential oil, 100 mg/kg/day orally, plus gentamicin, 80 mg/kg/day intraperitoneally E Losartan-treated group 5 Losartan, 10 mg/kg/day orally Losartan, 10 mg/kg/day orally, plus gentamicin, 80 mg/kg/day intraperitoneally