Cyclophosphamide-induced Urotoxicity Research Articles

Protective effects of asperuloside against cyclophosphamide-induced urotoxicity and hematotoxicity in rats

Abstract Cyclophosphamide (CP) is a highly efficacious chemotherapy drug for treating cancers and autoimmune disorders, but it is also notable for its deleterious side effects including urotoxicity in cancer patients, which has been extensively linked to CP-induced oxidative/inflammatory cascades. Herein, we investigated the protective effects of asperuloside (ASP) against CP-induced urotoxicity. Rats received oral administration of ASP (20 and 40 mg/kg bw/day) for 35 days and were injected with weekly CP (100 mg/kg bw, i.p.) for 4 weeks to induce acute bladder toxicity. CP acutely altered haematological parameters and significantly reduced body weight gain, bladder glutathione peroxidase, reduced glutathione, catalase, and superoxide dismutase activities. Furthermore, CP caused an upward surge in bladder malondialdehyde, nuclear factor-kappa B, tumour necrosis factor-α, interleukin-1β, and interleukin 6 concentrations. ASP supplementation ameliorated CP-induced haematological derangement and bladder urotoxicity through the restoration of oxidative and inflammatory parameters in CP-treated rats. These findings suggested that ASP could be valorised as a possible therapeutic agent against chemotherapy-related toxicities as well as oxidative damage disorders.

Effect of a Low Dose of Carvedilol on Cyclophosphamide-Induced Urinary Toxicity in Rats-A Comparison with Mesna.

One of the major side effects of cyclophosphamide (CPX)—an alkylating anticancer drug that is still clinically used—is urotoxicity with hemorrhagic cystitis. The present study was designed to evaluate the ability of carvedilol to protect rats from cyclophosphamide-induced urotoxicity. Rats were injected intraperitoneally (i.p.) with CPX (200 mg/kg) and administered carvedilol (2 mg/kg) intragastrically a day before, at the day and a day after a single i.p. injection of CPX, with or without mesna (40, 80, and 80 mg/kg i.p. 20 min before, 4 h and 8 h after CPX administration, respectively). Pretreatment with carvedilol partly prevented the CPX-induced increase in urinary bladder and kidney index, and completely protects from CPX-evoked alterations in serum potassium and creatinine level, but did not prevent histological alterations in the urinary bladder and hematuria. However, carvedilol administration resulted in significant restoration of kidney glutathione (GSH) level and a decrease in kidney interleukin 1β (IL-1β) and plasma asymmetric dimethylarginine (ADMA) concentrations. Not only did mesna improve kidney function, but it also completely reversed histological abnormalities in bladders and prevented hematuria. In most cases, no significant interaction of carvedilol with mesna was observed, although the effect of both drugs together was better than mesna given alone regarding plasma ADMA level and kidney IL-1β concentration. In conclusion, carvedilol did not counteract the injury caused in the urinary bladders but restored kidney function, presumably via its antioxidant and anti-inflammatory properties.

Protective effect of Spirulina against cyclophosphamide-induced urotoxicity in mice

Cyclophosphamide (CP) is an anti-neoplastic drug, which is widely used for treating cancer and non-malignant tumors. One of the major side effects of CP is hemorrhagic cystitis. Spirulina (Arthrospira platensis; Sp) is a blue-green algae with the ability to attenuate oxidative stress, which may be utilized for alleviating side effects of chemotherapeutic drugs in the clinic. The aim of the present study was to evaluate the ability of Sp, to protect mice from cyclophosphamide-induced urotoxicity and hemorrhagic cystitis due to its antioxidant properties. Adult female mice were orally administered Sp (600 g/kg body weight/day) over nine days as well as a single dose of CP (40 mg/kg body weight) intraperitoneally either four days previously (CP + Sp group) or four days after the start of Sp intake (Sp + CP group); two further groups were treated with either Sp or CP only, respectively. The results showed that CP induced hemorrhagic cystitis in mice, with levels of malondialdehyde (MAD) significantly increased and those of glutathione (GSH) decreased compared with the control group (P < 0.05), while the opposite effects were observed in the mice who received Sp only (P < 0.05). Furthermore, in the CP + Sp group, MAD and GSH levels were improved compared with those in the CP only group, and in the Sp + CP group, the effects of CP were reversed. In addition, histomorphological alterations of the urinary bladder were significantly lower than those in the CP group. In conclusion, pre-treatment with Sp protected mice from CP-induced urotoxicity, probably via its anti-oxidant and anti-apoptotic properties.

Uroprotective mechanism of quercetin against cyclophosphamide-induced urotoxicity: Effect on oxidative stress and inflammatory markers.

The urotoxicity is a common complication associated with patients receiving cyclophosphamide (CYP). This study was designed to investigate the uroprotective mechanism of quercetin (Quer) flavonoid against CYP induced urotoxicity via determination of oxidative stress markers as well as inflammatory mediators in bladder tissue. Forty male Wistar rats were divided into four groups; Normal group: received saline for 10 days. Quer control group: received quercetin 50 mg/kg/day for 10 days. CYP group: received saline for 10 days and injected with a single dose of 150 mg/kg CYP intraperitoneal (i.p) at day 8. The Quer + CYP group: received Quer 50 mg/kg/day for 10 days plus CYP 150 mg/kg i.p. injection at day 8. The CYP injection produced a significant elevation in bladder contents of malondialdehyde (MDA), and nitric oxide (NO), and bladder protein levels and expressions of tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6) in addition to the upregulation of cyclooxygenase-2 (COX-2) bladder gene expression. Also, CYP injection showed a marked reduction in bladder levels of catalase, superoxide dismutase (SOD), and IL-10 when compared with normal group. Moreover, histopathological examination of the bladder showed degenerative alterations, severe edema, and inflammation following CYP injection. Quer attenuated the biochemical markers and histopathological changes induced by CYP. The uroprotective effect of Quer was exerted by restoring the balance between oxidative/antioxidative status and pro-/anti-inflammatory cytokines via its antioxidant and anti-inflammatory activities.

Incidence of Cyclophosphamide-induced Urotoxicity and Protective Effect of Mesna in Rheumatic Diseases.

To assess bladder toxicity of cyclophosphamide (CYC) and uroprotective effect of mesna in rheumatic diseases. Data of 1018 patients (725 women/293 men) treated with CYC were evaluated in this retrospective study. All of the following information was obtained: the cumulative CYC dose, route of CYC administration, duration of therapy, concomitant mesna usage, and hemorrhagic cystitis. Cox proportional hazard model was used for statistics. We identified 17 patients (1.67%) with hemorrhagic cystitis and 2 patients (0.19%) with bladder cancer in 4224 patient-years. The median time for diagnosis to hemorrhagic cystitis was 10 months (4-48) and bladder cancer was 8 years (6-10.9). There were 583 patients (57.2%) who received mesna with intravenous CYC therapy. We observed similar incidence rate for hemorrhagic cystitis in both patient groups concomitantly treated with or without mesna [9/583 (1.5%) vs 8/425 (1.8%) respectively, p = 0.08]. Cumulative CYC dose (HR for 10-g increments 1.24, p < 0.001) was associated with hemorrhagic cystitis. Cumulative dose was the only risk factor for hemorrhagic cystitis in patients treated with CYC. No proof was obtained for the uroprotective effect of mesna in our cohort.

Role of Nicotinic and Estrogen Signaling during Experimental Acute and Chronic Bladder Inflammation

Inflammation is a physiological process that characterizes many bladder diseases. We hypothesized that nicotinic and estrogen signaling could down-regulate bladder inflammation. Cyclophosphamide was used to induce acute and chronic bladder inflammation. Changes in bladder inflammation were measured histologically and by inflammatory gene expression. Antagonizing nicotinic signaling with mecamylamine further aggravated acute and chronic inflammatory changes resulting from cyclophosphamide treatment. Estrogen and nicotinic signaling independently attenuated acute bladder inflammation by decreasing neutrophil recruitment and down-regulating elevated lipocalin-2 and cathepsin D expression. However, the combined signaling by the estrogen and nicotinic pathways, as measured by macrophage infiltration and up-regulation of interleukin-6 expression in the bladder, synergistically reduced chronic bladder inflammation. The elevated expression of p65 nuclear localization in bladders treated with cyclophosphamide or cyclophosphamide with mecamylamine suggested nuclear factor-kappa B activation in the chronic inflammatory process. The complementary treatment of 17 beta-estradiol and the nicotinic agonist anabasine resulted in the translocation of p65 to the cytoplasm, again greater than either alone. Activation of nuclear factor-kappaB can result in macrophage activation and/or elevation in epithelial proliferation. These data suggest that 17 beta-estradiol and anabasine reduce chronic bladder inflammation through reduction of nuclear translocation of p65 to suppress cytokine expression.

In vitro antioxidant activity of Juglans regia L. bark extract and its protective effect on cyclophosphamide-induced urotoxicity in mice

Walnut (Juglans regia L.) bark has been claimed to possess anti-inflammatory, blood purifying, anticancer, depurative, diuretic and laxative activities. It contains several therapeutically active constituents, especially polyphenols. We studied the antioxidant potential of aqueous extract of walnut bark and its modulatory effect on cyclophosphamide (CP)-induced urotoxicity in Swiss albino male mice. Free radical-scavenging activity of extract was assessed in four in vitro assays. The phenolic and flavonolic contents of the extract were also measured. Walnut bark extract treatment (150 mg/kg p.o. × 10 days) resulted in protective restoration of decreased antioxidants in CP-treated (18 mg/kg i.p. × 10 days) animals. CP treatment caused decreases in the activities of catalase (CAT), glutathione peroxidase (GP), glutathione reductase (GR) and glutathione S-transferase (GST) and in the glutathione (GSH) content in urinary bladder and a significant concomitant increase in lipid peroxidation (LPO). Administration of extract restored all the antioxidants significantly and lowered the elevated LPO in the bladder. A correlation between radical scavenging capacities of the extract with phenolic content was observed thus justifying its antioxidant potential against oxidative stress-mediated urotoxicity in mice. Walnut is reported to possess antiproliferative activity. Its protective effect on CP-induced toxicity in bladder is a promising activity, which warrants possible clinical investigations on this medicinal plant.

Effect of naturally occurring isothiocyanates in the inhibition of cyclophosphamide-induced urotoxicity

Cyclophosphamide-induced urotoxiciy was reduced in Swiss albino mice by the treatment of naturally occurring isothiocyanates such as AITC or PITC (25 microg/dose/animal, i.p.) for 5 days along with CTX (1.5 mmol/kg body wt.; i.p.). Severely inflamed and dark coloured urinary bladders of the CTX alone treated animals were found to be normalized on morphological analysis by the treatment of AITC or PITC. Urine protein levels were reduced by the treatment with AITC (6.2 +/- 0.37 g/l) and PITC (6.56 +/- 1.56 g/l), which was elevated by CTX administration (8.66 +/- 0.47 g/l). Urine urea N2 that was enhanced significantly by CTX administration (26.87 +/- 1.86 g/l) was reduced by treatment with both AITC (17.38 +/- 0.06 g/l) and PITC (15.85 +/- 1.56 g/l). GSH content, which was drastically reduced by CTX administration in both bladder (0.87 +/- 0.1 nmol/mg protein) and liver (2.47 +/- 0.6 nmol/mg protein) was enhanced by treatment with AITC and PITC both in bladder (AITC- 3.65 +/- 0.18 nmol/mg protein; PITC- 2.8 +/- 0.15 nmol/mg protein) and in liver (AITC- 4.10 +/- 0.81 nmol/mg protein; PITC- 4.70 +/- 0.44 nmol/mg protein). Histopathology of the bladders of CTX alone treated group showed severe necrosis of the tissue whereas AITC and PITC treated group showed normal bladder pathology.

Effect of Withania somnifera on cyclophosphamide-induced urotoxicity

Administration of an extract from the plant Withania somnifera (Family: Solanaceae) (20 mg/dose/animal; i.p.) for five days along with cyclophosphamide (CTX) (1.5 mmol/kg body wt. i.p.) reduced the CTX induced urotoxicity. Morphological analysis of the bladders of the CTX-treated group showed severe inflammation and dark coloration whereas CTX along with the Withania-treated group showed normal bladder morphology. The extract was found to reduce the protein level in the serum (7.92 g/l) after 4 h of CTX treatment, which was higher in the CTX alone-administered group (11.44 g/l). Blood urea N 2 level which was drastically enhanced (136.78 mg/100 ml) 2 after the CTX treatment was significantly reduced (52.08 mg/100 ml) when the animals were treated with Withania extract. Similarly the glutathione (GSH) content in both bladder (1.55 μmol/mg protein) and liver (3.76 μmol/mg protein) was enhanced significantly ( P<0.001) in the Withania-treated group compared with the CTX alone-treated animals (bladder 0.5 μmol/mg protein; liver 1.2 μmol/mg protein) Histopathological analysis of the bladder of CTX alone-treated group showed severe necrotic damage where as the Withania somnifera-treated group showed normal bladder architecture.

Drug interaction effects on antitumour drugs (XV): Disulfiram as protective agent against cyclophosphamide-induced urotoxicity without compromising antitumour activity in mice.

The prevention of cyclophosphamide-induced urotoxicity by disulfiram was studied in mice. A single dose of cyclophosphamide (100-400 mg/kg, intraperitoneally) produced a significant dose-dependent increase in urinary bladder weight within 48 hr of treatment. Disulfiram prevented cyclophosphamide-induced bladder damage in a dose-dependent manner in mice when orally administered simultaneously with antitumour agents, but failed to diminish the acute toxicity, leukocytotoxicity and immunotoxicity of cyclophosphamide. The protective effect of disulfiram on the bladder was critically dependent on administration timing. Oral administration of disulfiram between 60 min. before and 60 min. after the injection of cyclophosphamide was found to be effective. The optimum time was simultaneous administration of both drugs. Diethyldithiocarbamate and carbon disulfide, metabolites of disulfiram, prevented cyclophosphamide-induced bladder damage when administered simultaneously with cyclophosphamide 1 to, 3 or 5 hr afterwards. Disulfiram slightly potentiated the antitumour activity of cyclophosphamide against Sarcoma 180 or EL-4 leukaemia in vivo when administered simultaneously with cyclophosphamide. In contrast, diethyldithiocarbamate or carbon disulfide did not interfere with cyclophosphamide antitumour activity when administered 3 hr after cyclophosphamide. From these preliminary studies, disulfiram appears to be a likely candidate for protection against cyclophosphamide-induced urotoxicity without compromising the therapeutic utility of the alkylating agent.

A preliminary clinical study of cyclophosphamide with reduced glutathione as uroprotector.

Reduced glutathione (GSH) has been reported to be an effective protector against cyclophosphamide-induced urotoxicity in experimental models, providing protection comparable to that of mesna. This paper describes our preliminary results of the clinical use of GSH in combination with cyclophosphamide. GSH was administered i.v. in two divided doses of 2.5 g, 15 min before and 30 min after escalating doses of cyclophosphamide ranging from 1.2 up to 1.6 g/m2 (1-h infusion). GSH was well tolerated and did not produce unexpected toxicity. The lack of bladder damage, including microscopic hematuria, supports the protective role of this thiol compound.

Comparison of reduced glutathione with 2-mercaptoethane sulfonate to prevent cyclophosphamide-induced urotoxicity

Since the usefulness of high-dose cyclophosphamide is often limited by hemorrhagic cystitis, and the use of sulfhydryl-containing compounds prevents this side effect, this study was carried out to compare the uroprotective efficacy of 2-mercaptoethane sulfonate (MESNA) with that of reduced glutathione. In experimentally cyclophosphamide-induced urotoxicity in mice, the protective efficacy and potency of these thiols were similar, since both agents afforded complete protection in the same dose range. This finding suggests that these compounds are suitable sources of urinary thiols. Although the rationale for the clinical use of these protectors is similar, glutathione may have therapeutic advantages over MESNA because of a wider margin of safety and multiple protective actions displayed by the tripeptide thiol.

Prevention of cyclophosphamide-induced urotoxicity by reduced glutathione and its effect on acute toxicity and antitumor activity of the alkylating agent.

The effect of reduced glutathione on acute lethal toxicity and urotoxicity induced by cyclophosphamide was studied on both mice and rats. The results of this investigation indicate that reduced glutathione is an effective protective agent against bladder damage from treatment with the alkylating agent. The timing of glutathione administration (IV) with respect to cyclophosphamide treatment influenced the uroprotective efficacy of the thiol compound. A schedule-dependent protective effect of glutathione against acute lethal toxicity of the antitumor drug was also observed. This partial protection was accompanied by a reduction in body weight loss following cyclophosphamide treatment. The therapeutic activity of cyclophosphamide on two experimental tumor systems (L1210 and Gross leukemia) was not impaired by combined treatment with glutathione, even at a relatively high dose of glutathione compared with cyclophosphamide.