Hemorrhagic Cystitis In Rats Research Articles

Involvement of Interleukin-6-Regulated Nitric Oxide Synthase in Hemorrhagic Cystitis and Impaired Bladder Contractions in Young Rats Induced by Acrolein, a Urinary Metabolite of Cyclophosphamide

Hemorrhagic cystitis is a common complication in children receiving cyclophosphamide, a chemotherapeutic alkylating agent. Acrolein is a urinary metabolite from cyclophosphamide and can induce hemorrhagic cystitis. Here, we investigated the effects and mechanisms of acrolein by intravesical instillation on urinary bladder muscle contractions and pathological alterations in rats. Acrolein instillation significantly increased the muscle contractions of rat bladder detrusor after 1 and 6 h but markedly decreased detrusor contractions after 24 h. Acrolein increased phosphorylated protein kinase C (pan-PKC) expressions in bladders after 1 and 6 h but inhibited it after 24 h. Inducible nitric oxide (NO) synthase (iNOS) protein expressions were markedly induced in bladders 24 h after acrolein treatment. Twenty-four-hour acrolein instillation increased the levels of nitrite/nitrate and interleukin-6 (IL-6) in the urinary bladder. The iNOS inhibitors significantly inhibited the acrolein-increased nitrite/nitrate levels, but not IL-6 levels. IL-6-neutralizing antibodies effectively inhibited the acrolein-increased NOx levels. The increased detrusor contractions by 1-h acrolein treatment were significantly reversed by the PKC inhibitor RO32-0432, and the decreased detrusor contractions by 24-h acrolein treatment were significantly reversed by the iNOS inhibitor and IL-6-neutralizing antibody. Both the iNOS inhibitor and IL-6-neutralizing antibody effectively reversed the increased iNOS expression, decreased PKC phosphorylation, increased bladder weight, and hemorrhagic cystitis in rats 24 h after acrolein treatment. Taken together, these results suggest that an IL-6-regulated iNOS/NO signaling pathway participates in the acrolein-triggered detrusor contraction inhibition and hemorrhagic cystitis. These findings may help us to find a new strategy to treat cyclophosphamide-induced hemorrhagic cystitis.

Oral Glutamine Attenuates Cyclophosphamide-Induced Oxidative Stress in the Bladder but Does Not Prevent Hemorrhagic Cystitis in Rats

Cyclophosphamide (CP) is widely used in the treatment of cancer and non-malignant disease states such as rheumatoid arthritis. Hemorrhagic cystitis is a major dose-limiting side effect of CP. The incidence of this side effect is related to the dosage and can be as high as 75%. Elimination of the side effects of CP can lead to better tolerance of the drug, and a more efficient therapy can be achieved for patients in need of CP treatment. Several studies have demonstrated that oxidative stress and neutrophil infiltration play important roles in CP-induced bladder damage. Glutamine is utilized under clinical conditions for preventing chemotherapeutic drug-induced side effects, based on its ability to attenuate oxidative stress. The aim of the study is to verify whether glutamine prevents CP-induced oxidative stress and bladder damage using a rat model. Adult male rats were administered 150mg/kg body weight of CP intraperitoneally. Glutamine pretreated rats were administered 1g/kg body weight of glutamine orally 2h before the administration of CP. Vehicle/glutamine-treated rats served as controls. All the rats were killed 16h after the dose of CP/vehicle. The urinary bladders were removed and used for light microscopic and biochemical studies. The markers of oxidative stress including malondialdehyde content, protein carbonyl content, protein thiol, and myeloperoxidase activity, a marker of neutrophil infiltration, were measured in bladder homogenates. CP treatment induced hemorrhagic cystitis in the rats. Pretreatment with glutamine significantly reduced CP-induced lipid peroxidation (p < 0.01), protein oxidation (p < 0.01), and increase in myeloperoxidase activity (p < 0.05). However, it did not prevent CP-induced bladder damage. The results of the present study show that glutamine pretreatment does not attenuate CP-induced hemorrhagic cystitis, although it prevents CP-induced oxidative stress and neutrophil infiltration significantly. It is therefore necessary to clarify the utility of glutamine as a chemoprotective agent before it is recommended in the market as a nutrient supplement.

Protective Effects of Bilirubin Against Cyclophosphamide Induced Hemorrhagic Cystitis in Rats

The end product of the heme oxygenase pathway, bilirubin, is the most abundant endogenous antioxidant in mammals. We report the heme oxygenase-1 mediated production of bilirubin and its cytoprotective roles in cyclophosphamide induced hemorrhagic cystitis in rats. Female Sprague-Dawley rats received intraperitoneal administration of cyclophosphamide. In the first experiment hemin (an inducer of heme oxygenase-1) with or without zinc protoporphyrin IX (an inhibitor of heme oxygenase activity) was given before cyclophosphamide injection. Endogenous bilirubin production was analyzed in bladder tissues immunohistochemically. In another experiment bilirubin solution was administered before the cyclophosphamide injection. Changes in bladder weight, microscopic feature and expression levels of inducible nitric oxide synthase, proinflammatory cytokines and heme oxygenase were evaluated using polymerase chain reaction and immunostaining. Bilirubin was generated in bladders with cyclophosphamide induced cystitis, especially in the urothelium and suburothelium. Hemin pretreatment provided increased production of endogenous bilirubin, which was decreased by zinc protoporphyrin IX. In an evaluation of the roles of bilirubin exogenous bilirubin administration ameliorated cyclophosphamide induced inflammatory changes and reduced the increase in bladder weight. The elevated expression of inducible nitric oxide synthase and interleukin-1beta in cyclophosphamide induced cystitis was significantly down-regulated by exogenously applied bilirubin. The expression of heme oxygenase-1 and 2 was not modified by bilirubin administration. Cyclophosphamide induced hemorrhagic cystitis is accompanied by endogenous bilirubin production through heme oxygenase-1 induction in the bladder. Bilirubin has cytoprotective roles in association with the down-regulation of inducible nitric oxide synthase expression. Our results suggest that bilirubin may have therapeutic potential against bladder inflammatory insults such as cyclophosphamide induced cystitis.

Lidocaine 2% Gel Versus Plain Lubricating Gel for Pain Reduction During Flexible Cystoscopy: A Meta-Analysis of Prospective, Randomized, Controlled Trials

The current literature shows mixed results for the effectiveness of topical intraurethral lidocaine gel as local anesthesia during flexible cystoscopy. We performed a meta-analysis of randomized, controlled trials of the efficacy of 2% lidocaine vs plain gel for decreasing the pain that male patients incur during flexible cystoscopy. A search of the literature from 1950 to September 2006 yielded 46 applicable articles. Search terms included cystoscopy and pain. Study selection included randomized controlled trials, flexible cystoscopy, males, control groups receiving plain gel and treatment groups receiving 2% lidocaine before cystoscopy. Data extraction was done by 2 of us (ARP and JSJ) who independently reviewed each study and were blinded to identifying features. The primary outcome measured was pain incurred by the patient throughout the entire cystoscopy procedure, as measured using a visual analog score. Data from 9 eligible trials on a total of 817 patients in 7 publications were included in the meta-analysis. Using a random effects model the difference between visual analog scale pain scores in patients receiving 2% lidocaine and plain gel was estimated to be -4.61 (approximate 95% CI -9.6, 0.385), indicating no statistically significant difference. Based on a meta-analysis of 9 randomized controlled trials there is no evidence to suggest a statistically significant difference in the efficacy of pain control between lidocaine gel and plain gel lubrication in men during flexible cystoscopy. This supports the conclusion that its benefit is limited to lubrication and any other perceived benefit is consistent with placebo.

Cyclooxygenase-2 expression on ifosfamide-induced hemorrhagic cystitis in rats

Hemorrhagic cystitis (HC) is a limiting side effect of chemotherapy with ifosfamide (IFS). In this study, we investigated the participation of cyclooxygenase-2 (COX-2) upon ifosfamide-induced HC. Male Wistar rats (150-200 g; six rats per group) were treated with saline, IFS (400 mg/kg, i.p.) and analyzed by changes in bladder wet weight, macroscopic and microscopic parameters, and COX-2 expression. In other groups etoricoxib (selective COX-2 inhibitor), indomethacin (non-selective COX inhibitor), thalidomide (selective TNF-alpha inhibitor), pentoxifyllin (non-selective TNF-alpha inhibitor) were added 1 h before IFS administration. The classical protocol using three doses of Mesna was also evaluated and compared with two extra doses of etoricoxib or indomethacin. COX-2 was expressed significantly 24 h after IFS administration mainly in myofibroblasts and mast cells evaluated by immunohistochemistry. Treatment 1 h before IFS injection with etoricoxib, indomethacin, thalidomide, and pentoxifylline reduced COX-2 expression and some macroscopic and microscopic parameters in IFS-induced HC. Moreover, addition of etoricoxib or indomethacin with the last two doses of Mesna was more efficient than three doses of Mesna alone when evaluated microscopically. COX-2 participates in the pathogenesis of IFS-induced HC and the treatment with COX and TNF-alpha inhibitors reduced COX-2 expression. The addition of COX-inhibitors to the last two doses of Mesna represents a new therapeutic strategy of preventing HC.

Comparison of uroprotective efficacy of mesna and amifostine in Cyclophosphamide- induced hemorrhagic cystitis in rats

Hemorrhagic cystitis (HC) is a dose limiting side effect of cyclophosphamide (CYP). In this study, we aimed to investigate the role of amifostine in the protection of CYP-induced HC and compare its efficacy with mesna. This animal study was conducted in the Experimental Animals Breeding and Research Center of the Medical Faculty of Uludag University. Male Wistar rats (150-200 g; 10 rats per group) were randomly assigned to four groups. Group I (control group) received no drugs, group II received CYP (200 mg/kg, i.p.) alone, group III received amifostine (200 mg/kg, i.p.) and CYP, and group IV received CYP and mesna (40 mg/kg, i.p.) immediately and 4 and 8 h after administration of CYP. Bladders of animals were assessed macroscopically and histologically 24 h later. Gross assessment for presence of edema and hemorrhage and histological evaluation of damage to the bladder were scored according to Gray's criteria. For macroscopic and microscopic data, we used statistical evaluation by Kruskal-Wallis nonparametric analysis of variance followed by the Mann-Whitney U-test. All the animals in group II had evidence of HC. Significant histological damage and macroscopic changes were present in this group compared to control group (P<0.001). The median scores for bladder damage in group III and IV were significantly lower compared to group II (P<0.001). When the median scores for bladder damage of group I, III, and IV were compared, there was no significant difference among these groups. This study demonstrated the efficacy of amifostine in prevention of cyclophosphamide-induced hemorrhagic cystitis.

Contribution of antioxidants to preventive effect of mesna in cyclophosphamide-induced hemorrhagic cystitis in rats.

The aim of this study was to evaluate whether combination of antioxidants and mesna may prevent cystitis induced by cyclophosphamide better than mesna alone. A total of 46 male Sprague-Dawley rats were divided into six groups. Five groups received single dose of cyclophosphamide (CP, 100 mg/kg) intraperitoneally with the same time intervals: group 2 received CP only, group 3 received mesna (21.5 mg/kg for three times), group 4 beta-carotene (20 mg/kg for two times) and mesna, group 5 received alpha-tocopherol (20 mg/kg for two times) and mesna, and group 6 received melatonin (5 mg/kg for two times) and mesna on the day of CP injection. Group 1 served as control. CP injection resulted in severe cystitis. Mesna has showed meaningful but not full protection against CP toxicity. Although beta-carotene did not show any additional beneficial effect when combined with mesna, alpha-tocopherol and especially melatonin with mesna resulted full protection that the pathologist, blinded to the slides, could not differ from sham control. Oxidants may be important in the pathogenesis of CP-induced cystitis. Melatonin and alpha-tocopherol may help to ameliorate bladder damage along with other drugs such as mesna and diuretics.

Ternatin, a flavonoid, prevents cyclophosphamide and ifosfamide-induced hemorrhagic cystitis in rats.

To compare the classical uroprotective efficacy of mesna (2-mercaptoethanesulfonic acid) with ternatin (flavonoid isolated from Egletes viscosa Less.) in cyclophosphamide (CYP) and ifosfamide (IFS) induced hemorrhagic cystitis (HC). Male Wistar rats received an intraperitoneal injection of saline, CYP or IFS and were treated with saline or mesna, 5 min before, 4 and 8 h after CYP or IFS administration. In other animals, 1, 2 or 3 doses of mesna were replaced with ternatin or 3 doses of mesna were replaced with dimethylsulphoxide (DMSO), ternatin diluent. In an additional group, the last 2 doses of mesna were replaced with saline. HC was evaluated 24 h after CYP or IFS administration. CYP or IFS treatment induced marked changes in macroscopic and microscopic evaluation and in bladder wet weight (BWW), and these alterations were significantly inhibited by treatment with 3 doses of mesna, as well as by the replacement of 1 or 2 doses of mesna with ternatin. The replacement of 2 doses of mesna with saline or all doses of mesna with ternatin or DMSO did not prevent HC. In conclusion, the replacement of 1 or 2 doses of mesna with ternatin efficiently blocked CYP- or IFS-induced HC, however mesna is necessary for initial uroprotection.

Inducible nitric oxide synthase inhibition in cyclophosphamide induced hemorrhagic cystitis in rats.

Cyclophosphamide (CP) is an antineoplastic agent used alone or in combination with other chemotherapeutic agents for the treatment of many neoplastic diseases. Hemorrhagic cystitis (HC) is a major potential toxicity and dose limiting side effect of CP. Recently, it has been shown that endogenous inflammatory mediators are involved in cystitis by increasing nitric oxide (NO) production in target tissue. The aim of this study was to evaluate the relationship between NO and CP induced hemorrhagic cystitis HC in rats. A total of 30 female Spraque-Dawley rats were divided into 4 groups. Group 1 served as control, three groups received single dose of CP (100 mg/kg) intraperitoneally (i.p.): group 2 received CP only. Group 3 received the NO precursor L-arginine (80 mg/kg/day), and group 4 received the selective inducible NO synthase (iNOS) inhibitor S-methylisothiourea (SMT; 20 mg/kg/day) before and the day after cyclophosphamide injection. CP injection resulted in severe cystitis. SMT but not L-arginine produced marked inhibition of CP induced bladder damage. We concluded that NO produced by iNOS, is an important mediator in the pathogenesis of CP induced cystitis.

Involvement of Nitric Oxide and Hyperbaric Oxygen in the Pathogenesis of Cyclophosphamide Induced Hemorrhagic Cystitis in Rats

We evaluated the relationship between nitric oxide and hyperbaric oxygenation in the pathogenesis and treatment of cyclophosphamide induced hemorrhagic cystitis in rats. Cyclophosphamide (100 mg/kg) was injected in male Sprague-Dawley rats for cystitis induction. Animals were treated before and the day after cyclophosphamide injection with 100 mg/kg of the nitric oxide substrate L-arginine, 20 mg/kg of the nonselective nitric oxide synthase inhibitor L-NG-nitroarginine methyl ester and 20 mg/kg of the selective inducible nitric oxide synthase inhibitor S-methylisothiourea. Animals were exposed to hyperbaric oxygen (2.8 atmospheres absolute for 90 minutes twice daily) with or without the administration of L-arginine and nitric oxide synthase inhibitors. Cyclophosphamide injection resulted in severe cystitis. S-methylisothiourea produced marked inhibition of cyclophosphamide induced bladder tissue damage. L-arginine and L-NG-nitroarginine methyl ester failed to a show meaningful protective effect. Hyperbaric oxygen protected the bladder only against ulceration. Moreover, hyperbaric oxygen did not contribute to the protective effects of L-arginine, L-NG-nitroarginine methyl ester or S-methylisothiourea. Nitric oxide produced by inducible nitric oxide synthase is an important mediator in the pathogenesis of cyclophosphamide induced cystitis. Hyperbaric oxygen has a beneficial effect on repairing bladder damage rather than on bladder protection.

The Use of Prostaglandin F2 Alpha for the Prophylaxis of Cyclophosphamide Induced Cystitis in Rats

It is well-established that cystitis is a significant cause of morbidity after cyclophosphamide administration in clinical populations. We induced hemorrhagic cystitis in rats using cyclophosphamide and compared controls to those pretreated with prostaglandin F2 alpha. Rats were then evalutated for differences in bladder weights, gross edema, gross bleeding, and histologic changes. The weights of the bladders which had been treated with cyclophosphamide were 94% greater than the controls. The weights of the bladders which were pretreated with prostaglandin F2 alpha before cyclophosphamide were only 19% greater than controls. Significant differences were found between cyclophosphamide controls and prostaglandin F2 alpha pretreated groups for gross weight (p < .0005), gross edema (p < .0005), and histology (.0005 < p < .005). We conclude that prostaglandin F2 alpha may be helpful in preventing cyclophosphamide induced cystitis.