Protective effects of lupeol in rats with renal ischemia‑reperfusion injury

Abstract

Acute kidney injury (AKI) caused by ischemia and, exogenous or endogenous nephrotoxic agents poses a serious health issue. AKI is seen in 1% of all hospital admissions, 2-5% of hospitalizations and 67% of intensive care unit (ICU) patients. The in-hospital mortality rates for AKI is 40-50, and >50% for ICU patients. Ischemia-reperfusion (I/R) injury in the kidney can activate inflammatory responses and oxidative stress, resulting in AKI. The common endpoint in acute tubular necrosis is a cellular insult secondary to ischemia or direct toxins, which results in effacement of brush border, cell death and decreased function of tubular cells. The aim of the present study was to assess if the reported antioxidant and anti-inflammatory agent lupeol can exert any effects against renal I/R damage. In total, 24 Wistar Albino rats were randomly assigned into four groups of 6, namely Sham, lupeol, ischemia and therapy groups. In the lupeol group, intraperitoneal administration of 100 mg/kg lupeol was given 1 h before laparotomy, whilst only laparotomy was conducted in the sham group. The renal arteries of both kidneys were clamped for 45 min, 1 h after either intraperitoneal saline injection (in the ischemia group) or 100 mg/kg lupeol application (in the therapy group). The blood samples and renal tissues of all rats were collected after 24 h. In blood samples, blood urea nitrogen (BUN) was measured by the urease enzymatic method, and creatinine was measured by the kinetic Jaffe method. Using ELISA method, TNF-α and IL-6 levels were measured in the blood samples, whereas malondialdehyde (MDA), glutathione (GSH), caspase-3 levels were measured in kidney tissues. In addition, kidney histopathological analysis was performed by evaluating the degree of degeneration, tubular dilatation, interstitial lymphocyte infiltration, protein cylinders, necrosis and loss of brush borders. It was determined that renal damage occurred due to higher BUN, creatinine, MDA, TNF-α and caspase-3 values observed in the kidney tissues and blood samples of rats in ischemia group compared with the Sham group. Compared with those in the ischemia group, rats in the therapy group exhibited increased levels of GSH and reduced levels of BUN, TNF-α, MDA. Furthermore, the ischemia group also had reduced histopathological damage scores. Although differences in creatinine, IL-6 and caspase-3 levels were not statistically significant, they were markedly reduced in the treatment group. Taken together, these findings suggest that lupeol can prevent kidney damage as mainly evidenced by the reduced histopathological damage scores, decreased levels of oxidative stress and reduced levels of inflammatory markers. These properties may allow lupeol to be used in the treatment of AKI.

Keywords: experimental study; ischemia/reperfusion injury; lupeol; renal.

Figure 1

Image of the right renal pedicle being obstructed with an atraumatic microvascular clamp. Blue arrow showing the right kidney and red arrow showing the intestines.

Figure 2

Representative histopathological images of each of the five criteria. (A) Arrow showing lymphocytic infiltration of the tubular structures. Magnification, x200. (B) All arrows indicate brush borders of the preserved cilia of proximal tubules. Magnification, x400. (C) Both arrows showing eosinophilic hyaline casts (protein cylinders) in the tubules. Magnification, x400. (D) Both arrows showing necrosis and degeneration of the tubular epithelium and the lower arrow pointing tubular dilatation. Magnification, x400.

Figure 3

Representative histopathological image of a damaged kidney from the ischemia group. Necrosis, degeneration, tubular dilatation and protein cylinders can all be seen in the circle. Lack of an epithelium on the basal membrane is shown by the arrow. Magnification, x400.

Figure 4

Representative histopathological image from each group. (A) Group I, score 25. Prominent hyalin casts and dilatation of tubules with degeneration, necrosis and loss of brush borders are indicated. (B) Group L, score 2. Locations of degeneration and loss of brush borders are shown. (C) Group S, score 12. Locations of degeneration, loss of brush borders, necrosis, and dilatation are indicated. (D) Group T, score 12. Locations of degeneration, loss of brush borders, necrosis, and dilatation are indicated. Magnification, x200. Ovals, prominent hyalin casts; asterisks, dilatation of tubules with degeneration; arrowhead, necrosis; arrows, loss of brush borders.

Figure 5

Representative histopathological image of a sparsely damaged kidney from the therapy group. Untreated group animals usually had more severe injury. Arrows indicate the location of tubular dilatation and focal protein cylinders. Star in the tubule represent the location of the partial loss of brush borders. Magnification, x400.