Mai M. Helmy

Montelukast abrogates rhabdomyolysis-induced acute renal failure via rectifying detrimental changes in antioxidant profile and systemic cytokines and apoptotic factors production.

In addition to antiasthmatic effect, the cysteinyl leukotriene receptor 1 (CysLT₁) antagonist montelukast shows renoprotective effect during ischemia/reperfusion and cyclosporine-induced renal damage. Here, we proposed that montelukast protects against rhabdomyolysis-induced acute renal failure. Compared with saline-treated rats, at 48 h following the induction of rhabdomyolysis using intramuscular glycerol (10 ml 50% glycerol/kg), significant elevations in serum levels of urea, creatinine, phosphate and acute renal tubular necrosis were observed. This was associated with elevations in serum Fas, interleukin-10, tumor necrotic factor-alpha, and transforming growth factor-beta1 and renal malondialdehyde and nitrite and detrimental reductions in renal catalase and superoxide dismutase activities. The effects of rhabdomyolysis on renal functional, biochemical and structural integrity and the associated changes in cytokines and Fas levels were abolished upon concurrent administration of montelukast (10 mg/kg i.p.) for 3 days (1 day before and 2 days after induction of rhabdomyolysis). Alternatively, administration of the anti-oxidant, α-tocopherol (400 mg/kg i.m.) for 3 days, succeeded in alleviating renal oxidative stress, but had no significant effect on the circulating levels of most cytokines and partially restored kidney functional and structural damage. Serum level of interleukin-6 was not altered by rhabdomyolysis but showed significant elevations in rats treated with montelukast or α-tocopherol. Collectively, motelukast abrogated functional and structural renal damage induced by rhabdomyolysis via ameliorating renal oxidative stress and modulation of systemic cytokines and apoptotic factors production. The results of this work are expected to open new avenues for early prevention of rhabdomyolysis-induced acute renal failure using selective CysLT₁ antagonists such as montelukast.

Redox imbalances incite the hypertensive, baroreflex, and autonomic effects of cyclosporine in rats.

Previous studies including ours showed that cyclosporine (CSA) causes baroreflex dysfunction and hypertension. Here we tested the hypothesis that oxidative damage in central and peripheral tissues underlies the hypertensive, baroreflex and autonomic actions elicited by CSA in rats. We investigated the effects of individual and combined 7-day treatments with CSA (25 mg/kg/day, n=7) and 4-hydroxy-2,2,6,6-tetramethyl piperidinoxyl (tempol, superoxide dismutase mimetic, 100 mg/kg/day, n=7) on blood pressure, reflex heart rate responses to peripherally mediated pressor and depressor responses, and biomarkers of oxidative stress. CSA elevated blood pressure and reduced reflex bradycardic (phenylephrine) and tachycardic (sodium nitroptrusside) responses. The ability of muscarinic (atropine, 1 mg/kg i.v.) or β-adrenoceptor blockade (propranolol, 1 mg/kg i.v.) to reduce reflex heart rate responses was reduced in CSA-treated rats, suggesting the impairment by CSA of reflex cardiac autonomic control. Concurrent administration of tempol abolished CSA-induced hypertension and normalized the associated impairment in baroreflex gain and cardiac autonomic control. Tempol also reversed the CSA-induced increases in aortic and brainstem nitrite/nitrate and malondialdehyde (MDA) and decreases in aortic superoxide dismutase (SOD). These findings implicate oxidative stress in peripheral and central cardiovascular sites in the deleterious actions of CSA on blood pressure and baroreceptor control of heart rate.

Evaluation of vitamin E in the treatment of erectile dysfunction in aged rats

AIMS The present study aims to elucidate the role of oxidative stress in erectile dysfunction associated with aging and to investigate the effect of treatment with vitamin E in this respect. MAIN METHODS Rats were divided into four groups: young (3-month-old), aged rats (18-month-old), aged rats given 80 IU of vitamin E/rat/day for 21-days, aged rats given 5mg/kg of sildenafil/day for 21-days. Intracavernosal pressure/mean arterial pressure (ICP/MAP), nitric oxide production, TBARS, GSH levels and SOD activity in corpus cavernosum were measured. KEY FINDINGS Significant decrease in ICP/MAP was observed in aged rats at both low and high frequency of stimulation. Significant increase in ICP/MAP was observed in aged rats treated with vitamin E over the range of 0.8 to 5 Hz but young control values were not restored. Percentage potentiation of ICP/MAP than aged group at 0.8 Hz was 326±41.3% and 897±72.2% for vitamin E and sildenafil respectively. Decreased levels of NO(2)/NO(3) and SOD activity in the penile tissue observed with aging were elevated back to control by either vitamin E or sildenafil. Penile concentration of TBARS was 20.86±0.83 for aged rats vs. 11.39±0.79 nmol/g tissue for young controls. Both vitamin E and sildenafil reduced penile TBARS in aged rats. SIGNIFICANCE This study proves that antioxidant therapy with vitamin E ameliorates the age-associated erectile dysfunction. Sildenafil may exert some antioxidant properties which add to the advantages of its long-term use. The effect of combinations of low-dose sildenafil and vitamin E on age-associated erectile dysfunction merits to be studied.

Potential hepato-protective effect of α-tocopherol or simvastatin in aged rats

BACKGROUND The effect of α-tocopherol or simvastatin treatment on antioxidant defense in liver of old rats was investigated. METHODS Endogenous thiobarbituric acid reactive substances (TBARS) and total nitrite/nitrate (NO(2)/NO(3)) levels as well as non-enzymatic glutathione (GSH) and enzymatic antioxidants [glutathione-S-transferase (GST), superoxide dismutase (SOD), glutathione peroxidase (GPX) and catalase (CAT) activities] were determined in the livers of young (3 months), aged (22 months), α-tocopherol- or simvastatin-treated aged rats. Serum lipid profile and liver function parameters were also assessed in these 4 groups. RESULTS Both α-tocopherol and simvastatin almost equally restored the age-induced changes in liver TBARS and CAT activity, serum aspartate aminotransferase (GOT), alanine aminotransferase (GPT) and alkaline phosphatase (ALP). α-Tocopherol, but not simvastatin, tended to restore GST and GPX activities in livers of aged rats. Simvastatin, on the other hand, counteracted age-induced increases in serum cholesterol, TG, LDL, total hepatic NO(2)/NO(3) level, and preserved a normal liver function during aging. CONCLUSION Thus, either drug may be beneficial, in spite of a mechanistic difference in the antioxidant effect of both of them, in alleviating age-induced liver injury.

Estrogen Provokes the Depressant Effect of Chronic Nicotine on Vagally Mediated Reflex Chronotropism in Female Rats

We recently reported that acute nicotine impairs reflex tachycardic activity in estrogen-depleted, but not estrogen-repleted, female rats, suggesting a restraining influence for estrogen against the nicotine effect. In this study, we tested whether the baroreflex-protective effect of estrogen can be replicated when nicotine was administered chronically. We also report on the dose dependence and autonomic modulation of the nicotine-baroreflex interaction. The effects of nicotine (0.5, 1, or 2 mg/kg/day for 14 days) on baroreflex curves relating changes in heart rate to increases [phenylephrine (PE)] or decreases [sodium nitroprusside (SNP)] in blood pressure were evaluated in sham-operated (SO), ovariectomized (OVX), and estrogen-replaced OVX (OVXE2) rats. Slopes of the curves were taken as a measure of baroreflex sensitivity (BRSPE and BRSSNP). In SO rats, both reflex bradycardic and tachycardic responses were attenuated by nicotine in a dose-related fashion. In nicotine-treated rats, blockade of β-adrenergic (propranolol), but not muscarinic (atropine), receptors caused additional reductions in reflex chronotropic responses, implying that nicotine selectively impairs reflex vagal activity. OVX selectively decreased BRSPE but not BRSSNP and abolished the nicotine-induced impairment of either response. These effects of OVX were reversed after treatment with estrogen or the estrogen receptor modulator raloxifene. In atropine-treated rats, comparable BRS values were demonstrated in all rat preparations regardless of the estrogen or nicotine milieu. Collectively, the inhibition of vagal activity accounts for the depressant effect of chronic nicotine on baroreflex activity. Furthermore, contrary to nicotines acute effects, the baroreflex-attenuating effect of chronic nicotine is exacerbated by estrogen.

Interruption of central neuronal pathway of imidazoline I1 receptor mediates the hypertensive effect of cyclosporine in rats.

Increased central sympathetic outflow secondary to afferent sympathetic excitation has been implicated in the hypertensive effect of the immunosuppressant drug cyclosporine (CSA). The present study investigated the roles of central alpha(2)-adrenoceptors and I(1)-imidazoline receptors in modulating the hypertensive action of CSA. The blood pressure (BP) response to CSA in conscious rats was assessed in the absence and presence of peripherally or centrally acting sympatholytic drugs. Also, the effect of selective pharmacologic blockade of alpha(2) or I(1) receptors by yohimbine and efaroxan, respectively, on the pressor response to CSA was evaluated. CSA (20 mg/kg i.v.) produced a rapid increase in BP that peaked (25+/-4 mm Hg) after approximately 4 min and continued for the 45 min study duration. Ganglionic (hexamethonium 20 mg/kg) or alpha(1)-adrenoceptor (prazosin 1 mg/kg) blockade reduced the pressor effect of CSA. Pressor responses to phenylephrine (alpha(1)-adrenoceptor agonist) were not affected by CSA, thereby eliminating a possible role for alterations of vascular alpha(1)-adrenoceptor responsiveness in CSA hypertension. CSA hypertension was attenuated in rats pretreated intravenously with drugs that reduce central sympathetic tone including clonidine (mixed alpha(2)/I(1)-receptor agonist, 30 microg/kg) or moxonidine (selective I(1)-receptor agonist, 100 microg/kg) in contrast to no effect for guanabenz (selective alpha(2)-receptor agonist, 30 microg/kg). Intracisternal (i.c.) administration of moxonidine also reduced CSA hypertension. Selective blockade of central I(1) (efaroxan, 0.15 microg/rat, i.c.) but not alpha(2) (yohimbine, 25 microg/5 microl/rat, i.c.) receptors abolished the hypertensive response to CSA. Together, these findings highlight that CSA elicits its hypertensive effect via disruption of central sympathoinhibitory pathways which include I(1)-imidazoline receptors.

Additive Renoprotection by Pioglitazone and Fenofibrate against Inflammatory, Oxidative and Apoptotic Manifestations of Cisplatin Nephrotoxicity: Modulation by PPARs

Nephrotoxicity is a major side effect for the antineoplastic drug cisplatin. Here, we employed pharmacological, biochemical, and molecular studies to investigate the role of peroxisome proliferator-activated receptors (PPARs) in cisplatin nephrotoxicity. Rats were treated with a single i.p. dose of cisplatin (5 mg/kg) alone or combined with pioglitazone (PPARγ agonist), fenofibrate (PPARα agonist), pioglitazone plus fenofibrate, or thalidomide (Tumor necrosis factor-α inhibitor; TNF-α). Cisplatin nephrotoxicity was evidenced by rises in renal indices of functional (blood urea nitrogen, BUN, and creatinine), inflammatory (TNF-α, interleukin 6, IL-6), oxidative (increased malondialdehyde, MDA, and decreased superoxide dismutase, SOD and nitric oxide metabolites, NOx), apoptotic (caspase 3), and histological (glomerular atrophy, acute tubular necrosis and vacuolation) profiles. Cisplatin effects were partly abolished upon concurrent exposure to pioglitazone, fenofibrate, or thalidomide; more renoprotection was observed in rats treated with pioglitazaone plus fenofibrate. Immunostaining showed that renal expressions of PPARα and PPARγ were reduced by cisplatin and restored to vehicle-treated values after simultaneous treatment with pioglitazone or fenofibrate. Fenofibrate or pioglitazone renoprotection remained unaltered after concurrent blockade of PPARα (GW6471) and PPARγ (GW9662), respectively. To complement the rat studies, we also report that in human embryonic kidney cells (HEK293 cells), increases caused by cisplatin in inflammatory, apoptotic, and oxidative biomarkers were (i) partly improved after exposure to pioglitazone, fenofibrate, or thalidomide, and (ii) completely disappeared in cells treated with a combination of all three drugs. These data establish that the combined use of pioglitazone and fenofibrate additively improved manifestations of cisplatin nephrotoxicity through perhaps GW6471/GW9662-insensitive mechanisms.

Pioglitazone ameliorates methotrexate-induced renal endothelial dysfunction via amending detrimental changes in some antioxidant parameters, systemic cytokines and Fas production

Methotrexate (MTX) is widely used in treatment of cancers and autoimmune diseases. However, nephrotoxicity is one of its most important side effects. The peroxisome proliferator-activated receptor gamma agonist, pioglitazone, is known to exert antiinflammatory and reno-protective effects in various kidney injuries. The purpose of this study was to investigate the potential involvement of endothelial damage in MTX-induced renal injury and to elaborate the possible protective effect of pioglitazone against MTX-induced endothelial impairment. Compared with saline-treated rats, treatment with MTX (7mg/kg for 3days) caused significant elevations in serum levels of urea and creatinine, increased renal nitrate/nitrite level and impaired renovascular responsiveness of isolated perfused kidney to endothelium-dependent vasodilations induced by acetylcholine (0.01-2.43nmol) and isoprenaline (1μmol). These effects were abolished by concurrent treatment with pioglitazone (2.5mg/kg, for 5days starting 2days before MTX). Alternatively, MTX treatment did not affect endothelium-independent renovascular relaxation induced by sodium nitroprusside (0.001-10μmole). The possibility that alterations in renal antioxidants, circulating cytokine and apoptotic factor (Fas) levels contributed to MTX-pioglitazone interaction was assessed. Pioglitazone treatment abrogated renal oxidative stress (decreased reduced glutathione and catalase activity and increased malondialdehyde), elevated serum cytokine (interleukin-6, interleukin-10, tumor necrosis factor-alpha and transforming growth factor-beta1) and Fas induced by MTX. Histologically, MTX caused defused tubular cells swelling and vacuolization associated with endothelial damage in renal arterioles. These effects disappeared upon co-treated with pioglitazone. Collectively, pioglitazone abolished MTX-induced endothelium dysfunction and nephrotoxicity via ameliorating oxidative stress and rectifying cytokines and Fas abnormalities caused by MTX.

Zileuton alleviates acute cisplatin nephrotoxicity: Inhibition of lipoxygenase pathway favorably modulates the renal oxidative/inflammatory/caspase-3 axis

OBJECTIVE The current study investigated for the first time the possible beneficial effect of zileuton, a selective 5-lipoxygenase inhibitor (5-LOX), against cisplatin-induced acute renal failure. METHODOLOGY Male Sprague-Dawley rats (180-200 g) were administered cisplatin once (5 mg/kg, i.p.) alone or combined with oral zileuton (10 mg/kg, given twice; 1 h before and 12 h after cisplatin). RESULTS Compared with control rats, acute cisplatin administration caused significant increases of BUN (33.76 ± 7.74 vs 61.88 ± 11.35 mg/dl), serum creatinine (0.61 ± 0.21 vs 1.56 ± 0.28 mg/dl), renal levels of MDA (6.40 ± 1.04 vs 20.52 ± 2.18 nmol/g tissue), NOx (3.45 ± 1.20 vs 17.70 ± 2.27 nmol/g tissue), TNF-α (6.71 ± 0.66 vs 23.71 ± 3.41 pg/g tissue), MPO (0.87 ± 0.09 vs 3.12 ± 0.41 U/mg tissue protein) and renal caspase-3 activity (2.81 ± 0.37 vs 12.70 ± 2.94 U/mg tissue protein). Whereas, total SOD activity (1.99 ± 0.53 vs 0.79 ± 0.06 U/mg tissue protein) and IL-10 (110.98 ± 19.70 vs 62.34 ± 14.42 pg/g tissue) were significantly decreased. Cisplatin-induced nephrotoxicity was further confirmed histopathologically (tubular necrosis, cystic dilatation of renal tubules, vacuolar degeneration of renal tubular epithelium with perivascular oedema, and interstitial fibrosis). These changes were accompanied by alteration in 5-LOX pathway manifested as elevated renal levels of 5-LOX, LTD4 and LTB4. Simultaneous administration of zileuton to the cisplatin-treated rats reversed the deleterious renal insults and restored the measured parameters near to control values. CONCLUSIONS These data establish the first experimental evidence that zileuton abrogates cisplatin nephrotoxicity in rats probably via the inhibition of detrimental actions of 5-LOX products, thus favorably affecting renal oxidative/inflammatory/caspase-3 axis. Based on current findings, the therapeutic prospect of zileuton for this purpose is recommended.

Enhanced lipoxygenase/LTD4 signaling accounts for the exaggerated hypertensive and nephrotoxic effects of cyclosporine plus indomethacin in rats

The combined use of cyclosporine (CSA) and nonsteroidal antiinflammatory drugs (NSAIDs) causes exaggerated rises in systolic blood pressure (SBP) and nephrotoxicity. We examined whether these influences relate to the arachidonate/5-lipoxygenase (LOX) pathway. Rats were treated with CSA (20 mg kg-1 day-1), indomethacin (5 mg kg-1 day-1), or their combination for 10 days. Changes in SBP and renal biochemical/histopathological characteristics along with leukotriene levels were determined in rats treated with or without LT receptor antagonists. CSA or indomethacin caused: (i) renal tubular atrophy and interstitial fibrosis, (ii) increases in serum creatinine, blood urea nitrogen (BUN), and renal LTD4, LTB4, TNF-α, TGF-β1, and caspase-3, and (iii) decreases in renal PGE2 and total antioxidant capacity (TAC). SBP measured by tail-cuff plethysmography was increased by CSA but not indomethacin. These effects were mostly intensified in rats treated with CSA plus indomethacin. The co-treatment with montelukast (cysteinyl LT receptor blocker), but not ONO-4057 (LTB4 receptor blocker), ameliorated CSA/indomethacin-evoked hypertension, renal structural/biochemical deterioration, and LTD4 levels. Moreover, montelukast exhibited a greater capacity in reversing inflammatory, oxidative, apoptotic, and fibrotic abnormalities induced by CSA/indomethacin. Overall, lipoxygenase/LTD4 upregulation contributes to the exaggerated hypertension and nephrotoxicity caused by CSA/indomethacin. The therapeutic potential of cysteinyl LT receptor antagonism in rectifying CSA/NSAIDs-evoked anomalies is warranted.