Chronic renal failure-induced multiple-organ injury in rats is alleviated by the selective CysLT1 receptor antagonist montelukast

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Abstract

Chronic renal failure (CRF) is associated with oxidative stress that promotes production of reactive oxygen species and cytokine release. We aimed to investigate the possible protective effect of montelukast, a CysLT1 receptor antagonist, against oxidative damage in a rat model of CRF, induced by 5/6 reduction of renal mass. Male Wistar albino rats were randomly assigned to either the CRF group or the sham-operated control group, which received saline or montelukast (10 mg/kg, i.p.) for 4 weeks. At the end of the 4 weeks, rats were decapitated and trunk blood was collected. Creatinine, blood urea nitrogen and lactate dehydrogenase (LDH) activity were measured in the serum samples, while leukotriene B4, TNF-α, IL-1β, IL-6, total antioxidant capacity (AOC) and leukocyte apoptosis were assayed in plasma samples. Kidney, lung, heart and brain tissue samples were taken for the determination of tissue malondialdehyde (MDA), glutathione (GSH) levels, and myeloperoxidase (MPO) activity. Oxidant-induced tissue fibrosis was determined by tissue collagen contents, and the extent of tissue injuries was analyzed microscopically. CRF caused significant decreases in tissue GSH and plasma AOC, which were accompanied with significant increases in MDA levels, MPO activities, and collagen contents of all the studied tissues, while the circulating levels of the pro-inflammatory mediators, LDH activity, creatinine and BUN were elevated. Montelukast treatment reversed all these biochemical indices, as well as histopathological alterations induced by CRF. Similarly, flow cytometric measurements revealed that leukocyte apoptosis was increased in CRF group, while montelukast reversed this effect. In conclusion, CRF-induced oxidative tissue injury occurs via the activation of pro-inflammatory mediators and by neutrophil infiltration into tissues, and that protective effects of montelukast on CRF-induced injury can be attributed to its ability to inhibit neutrophil infiltration and apoptosis, to balance oxidant-antioxidant status and to regulate the generation of pro-inflammatory mediators.

Introduction

Chronic inflammation is a highly prevalent co-morbid condition that predicts poor clinical outcome in end-stage renal disease (ESRD) patients [1], [2], [3], enhancing mainly cardiovascular risk and mortality [4]. Evidence for oxidative stress in chronic renal failure (CRF) was based on the elevation of toxic lipid peroxidation products, which cause destruction and damage to cell membranes [5], [6], [7], [8]. Several earlier studies have revealed that these toxic products cause an inflammatory burden in CRF through the generation of an imbalance between increased production of reactive oxygen species (ROS) and limited or decreased antioxidant capacity [9].

A wide array of inflammatory biomarkers, such as C-reactive protein, interleukin (IL)-6 and white blood cell count, has been proven to be robust predictors of poor outcome in the ESRD patients [9], [10]. It is well documented that the inflammatory response in CRF is characterized by activation of neutrophils and collective action of chemotactic mediators [11]. The cellular sources of inflammatory cytokines enhanced in ESRD patients are mainly neutrophils and monocytes. Once neutrophils migrate into the tissue, they release reactive oxygen species, proteases, elastase, myeloperoxidase (MPO), cytokines and various other mediators [12]. Along with the various pro-inflammatory chemokines, cysteinyl leukotrienes (CysLTs), the 5-lipoxygenase metabolites of arachidonic acid, are also proven to be potent inflammatory mediators that cause tissue injury.

Antileukotriene drugs, i.e. leukotriene receptor antagonists and synthesis inhibitors, are a new class of anti-inflammatory drugs that have shown efficacy in the treatment of several inflammatory models. A selective reversible CysLT1 receptor antagonist, montelukast (MK-0476), was used in the treatment of asthma and is reported to reduce eosinophilic inflammation in the airways [13], [14], while CysLT1 receptor antagonists or biosynthesis inhibitors have been reported to ameliorate ethanol-induced gastric mucosal damage [15], [16] and experimental colitis [17]. Recently, we have shown that the CysLT1 receptor antagonist, montelukast, ameliorates burn- and sepsis-induced multiple organ damage [18], [19]. Furthermore, we have reported that montelukast improved microscopic damage and renal function and protected against renal ischemia/reperfusion injury in rats [20]. These data suggest that limiting the inflammatory cascade by blocking the CysLT1 receptors may also limit the oxidative damage induced by CRF. Therefore, using a rat model of chronic renal failure, the present study was designed to elucidate the putative protective effect of montelukast on renal functions and oxidant/antioxidant status of the kidney and the affected organs, lung, heart and brain.

Section snippets

Animals

Male Wistar albino rats (200–250 g) were housed in a room at a constant temperature of 22 ± 2 °C with a 12 h light–dark cycle, and had free access to standard pellet chow and water. All experimental protocols were approved by the Marmara University School of Medicine Animal Care and Use Committee.

Surgery and experimental protocol

The animals (n = 32) were randomly assigned to either the CRF group or the sham-operated control group. Under ketamine anesthesia (100 mg/kg ketamine and 0.75 mg/kg chlorpromazine; i.p.) and aseptic conditions,

Results

The BUN and creatinine levels in the saline-treated CRF group were found to be significantly higher than those in the sham-operated control rats treated with either montelukast or saline (p < 0.001; Table 1). Montelukast treatment in the CRF group reversed the serum creatinine level back to control levels, and significantly reduced the BUN level (p < 0.001). Serum LDH activity showed a significant increase in the saline-treated CRF group, indicating a generalized tissue damage (p < 0.01), while

Discussion

As observed by increased lipid peroxidation, collagen content and myeloperoxidase activity and decreased GSH level, the present results demonstrate that CRF in rats yields to oxidative injury in the renal tissue, as well as in the lung, heart and brain tissues. These observed tissue damages were accompanied by elevated serum levels of pro-inflammatory mediators (LTB4, TNF-α, IL-1β and IL-6), while histological analyses verified the severity of CRF-induced systemic inflammatory response in all

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