Effects of anulus fibrosus and experimentally degenerated nucleus pulposus on nerve root conduction velocity: relevance of previous experimental investigations using normal nucleus pulposus

STUDY DESIGN: Nerve conduction velocity was measured in the pig cauda equina after local application of anulus fibrosus or in vitro/postmortem degenerated nucleus pulposus from the same pig. OBJECTIVES: To analyze the effects of anulus fibrosus and degenerated nucleus pulposus on nerve conduction velocity. SUMMARY OF BACKGROUND DATA: Previous studies on nucleus pulposus-induced effects on nerve roots have used normal, nondegenerated nucleus pulposus. Because both anulus fibrosus and degenerated nucleus pulposus are commonly seen in the clinical situation of disc herniation, the value of the previous work could be questioned. METHODS: Anulus fibrosus and nucleus pulposus were harvested using a retroperitoneal approach. The nucleus pulposus was degenerated artificially either by addition of sodium lactate with HCl added to form a pH of either 6.0 or 3.5 (in vitro degeneration), or by storing the nucleus pulposus at 4 C until a pH of 6.0 (postmortem degeneration) was reached. After epidural application, the nerve conduction velocity was determined at 7 days (anulus fibrosus) or 3 days (degenerated nucleus pulposus). RESULTS: Application of anulus fibrosus did not induce any reduction of nerve conduction velocity. In vitro and postmortem degenerated nucleus pulposus induced a reduction of nerve conduction velocity similar to that of normal nucleus pulposus. CONCLUSIONS: Although only nerve function and not pain was assessed, it seems likely that previous experiments using normal nucleus pulposus may be relevant for evaluating the pathophysiologic mechanisms behind the nucleus pulposus-induced nerve root injury, also in a clinical perspective.     

一氧化氮在神经根性疼痛中的作用

目的：探索一氧化氮(NO)在髓核突出所致的神经根性疼痛中的作用。方法：取大鼠自体尾椎髓核无压迫下放置在L4和L5神经根表面，分别在术后3d及1、2、3、4周时观察大鼠后足机械刺激和热刺激敏感性的变化，并用免疫组化方法对移植髓核中的一氧化氮合酶(NOS)进行检测．探索NO在疼痛中的作用：结果：在无明显机械压迫情况下，大鼠腰神经根上放置自体髓核可产生痛觉过敏，移植髓核组织中NOS染色阳性一结论：髓核自身是引起腰腿痛的重要原因，NO可能参与疼痛的产生.     

The effects of a 5-HT2A receptor antagonist on blood flow in lumbar disc herniation : application of nucleus pulposus in a canine model

Blood vessel clots are found around the nerve root in patients with lumbar disc herniation. Thrombosis formation in the experimental application of nucleus pulposus to the nerve root has been shown in histological studies. In addition, reduction of blood flow and nerve conduction velocity are induced by the application of nucleus pulposus, which mimics lumbar disc herniation. In patients with lumbar disc herniation, nerve root block, which is thought to increase nerve blood flow, improves radiculopathy. 5-HT_2A receptor antagonists are used in chronic arterial occlusive diseases to improve blood flow and have been reported to work as well as nonsteroidal anti-inflammatory drugs in improving radiculopathy due to lumbar disc herniation in clinical studies. This study investigated the effects of a 5-HT_2A receptor antagonist on blood vessel diameter and blood flow in a canine experimental model of lumbar disc herniation. A total of 13 dogs were used. The animals were divided into three experimental groups and surgery was performed 1 week before measurements. In the nucleus pulposus group (NP; n = 5), the nucleus pulposus was applied to the nerve roots from the ventral side. In the sham group (n = 5), nucleus pulposus was not applied. In the naïve group (n = 3), the animals did not undergo surgery. Measurements of vessel diameter and blood flow were done before and after administration of saline and drugs. The diameters and blood flow volume of the observed blood vessels were measured on video-recordings every 10 min for 65 min. In all groups, vessel diameter and blood flow did not change before or after administration of saline. In the NP and sham groups, vessel diameter and blood flow increased significantly after administration of 5-HTRA compared with the naïve group. 5-HTRA improved blood vessel diameter and blood flow in the nerve roots inflamed by the application of nucleus pulposus but not in the intact nerve roots. 5-HTRA might be a potential agent to improve blood flow in the nerve roots of patients with lumbar disc herniation.     

临床症状体征与影像学检查分离的腰椎间盘突出症的发生机制研究进展

临床会出现少数症状体征与影像学检查结果不相符的腰椎间盘突出症患者,而单纯用传统的突出髓核直接机械压迫刺激神经根的理论不能解释这种反常的腰椎间盘突出症。腰椎间盘髓核的突出与患者临床症状体征的出现受多因素、多环节的影响,脊神经根的间接性机械压迫与神经根牵张效应为主要因素,而反常症状体征的产生往往与突出的髓核自身位置的迁移、神经系统对信息的传递以及髓核与硬膜囊或神经根的相互作用密切相关。此外,突出的髓核组织所继发的局部微循环、炎症改变,相应节段的骨质增生退变和腰椎应力姿势改变诱发此类反常腰椎间盘突出症患者出现多样性的症状体征。同时,一些患者还存在神经或椎体的先天性发育异常,并可能出现影像学检查上的误诊或漏诊。突出髓核对硬膜囊以及周围神经根之间的确切相互作用机制及其继发的局部病理生理、生物力学改变,病变责任节段的确定以及如何克服影像学检查的局限性需进一步研究。     

Selective inhibition of tumor necrosis factor-alpha prevents nucleus pulposus-induced thrombus formation, intraneural edema, and reduction of nerve conduction velocity: possible implications for future pharmacologic treatment strategies of sciatica

STUDY DESIGN: The possibility to prevent nucleus pulposus-induced functional and structural nerve root injury by selective tumor necrosis factor-alpha inhibition was assessed in an experimental model in the pig spine. OBJECTIVE: The objective of the study was to evaluate the role of tumor necrosis factor-alpha in the mediation of nucleus pulposus-induced nerve injury by using selective inhibition. SUMMARY OF BACKGROUND DATA: The cytokine tumor necrosis factor-alpha has been suggested to play a key role in the nerve root injury induced by local application of nucleus pulposus. However, previous studies have not been able to distinguish the effects between tumor necrosis factor-alpha and other disc-related cytokines because of the use of nonspecific cytokine inhibition. METHODS: Autologous nucleus pulposus was harvested from a lumbar disc and applied to the porcine sacrococcygeal cauda equina. The pigs were simultaneously treated with two selective tumor necrosis factor-alpha inhibitors (etanercept n = 8 and infliximab n = 5), a heparin analogue (enoxaparin n = 5) or saline for control (n = 5). After 7 days the nerve conduction velocity over the application zone was determined and samples of the exposed nerve roots were collected for light microscopic evaluation. RESULTS: The two tumor necrosis factor-alpha inhibitors prevented the reduction of nerve conduction velocity and also seemed to limit the nerve fiber injury, the intracapillary thrombus formation, and the intraneural edema formation. However, treatment with enoxaparin did not seem to be different from control regarding reduction of nerve conduction velocity or histologic changes. CONCLUSIONS: The data clearly indicate that tumor necrosis factor-alpha is involved in the basic pathophysiologic events leading to nerve root structural and functional changes after local application of nucleus pulposus. The study therefore provides a basic scientific platform with potential clinical implications regarding the use of anti-tumor necrosis factor-alpha medication as treatment in patients with disc herniation and sciatica.     

破裂型腰椎间盘突出症的诊断与治疗：（附32例报告）

介绍３２例因腰椎间盘纤维环破裂而引起的髓核直接突入椎管内压迫马尾神经和神经根所出现的临床症状、脊髓造影及术中所见、按术中所见，髓核突破纤维环时对硬膜囊及神经根压迫类型，结合术前脊髓造影及临床表现将其分为四种类型：大块型、碎块型、部分破裂型及硬膜囊内型。对破裂型椎间盘突出症的诊断、预防、治疗及预后提出了讨论。     

Mechanical and biochemical injury of spinal nerve roots: a morphological and neurophysiological study

Compression may induce morphological and neurophysiological changes in nerve roots. However, it has also been demonstrated experimentally that nucleus pulposus, without any compression, may induce similar changes when applied epidurally. The present study was undertaken to examine the morphological and functional effects of autologous nucleus pulposus and the combination of nucleus pulposus and compression in a pig model. Nucleus pulposus from a lumbar disc in the same animal was applied epidurally around the first sacral nerve root in the pig, with or without a specially designed constrictor. After 1 week, nerve root conduction velocity was determined in the exposed and in the contralateral control nerve root by local electrical stimulation and EMG recordings in the back muscles. Nerve root specimens were processed for blinded light-microscopic evaluation. There was a significant reduction in nerve conduction velocity for all exposed nerve roots as well as contralateral control nerve roots when nucleus pulposus had been applied. There were no statistically significant differences between the nerve conduction velocities recorded following the combined application of nucleus pulposus and compression and those recorded after application of nucleus pulposus alone. The reductions were similar to the reduction induced by the constrictor per se, as seen in a previous study. In all series there was also a decrease in conduction velocity in the control nerve roots, in contrast to previous studies. Light microscopy demonstrated axonal changes only in nerve roots exposed to the constrictor. In conclusion, both epidural nucleus pulposus and compression may induce a significant reduction in nerve conduction velocity. The combination, however, of these two agents does not increase the magnitude of such dysfunction. The potency of nucleus pulposus to induce changes in nerve roots after epidural application was further indicated by the fact that reduction in nerve conduction velocity also occurred in the contralateral control nerve roots in this series. The histological data suggest that axonal injury can not alone explain the reduction in nerve conduction velocity, and that the morphological basis for the functional changes must be sought at the subcellular level.     

Improvement of rejection-induced diastolic abnormalities in rat cardiac allografts with inducible nitric oxide synthase inhibition

OBJECTIVE: Inhibition of inducible nitric oxide synthase (nitric oxide II) activity has been proposed as a method to attenuate capillary leak and edema during rejection of heterotopically transplanted rat hearts. Myocardial edema has previously been implicated in diastolic dysfunction during allograft rejection. Accordingly, we tested the hypothesis that inducible nitric oxide synthase inhibition with aminoguanidine would alleviate left ventricular stiffening and myocardial edema formation in 4-day heterotopic rat heart allografts. METHODS: Passive left ventricular filling was studied in American Cancer Institute Lewis rats receiving heterotopic heart transplants receiving either aminoguanidine, a selective nitric oxide synthase inhibitor (n = 6); dexamethasone (1 mg. kg(-1). d(-1) administered subcutaneously) for 4 days after transplantation (n = 6); or intravenous saline solution (n = 6). American Cancer Institute-to-American Cancer Institute isografts (n = 6) were used as controls. RESULTS: Serum nitrite/nitrate levels in the aminoguanidine group (18 +/- 3 mmol/L) and dexamethasone group (22 +/- 4 mmol/L) were reduced versus the intravenous saline group (144 +/- 36 mmol/L [SEM]) to levels seen in controls (25 +/- 9 mmol/L). Left ventricular volume at 15 mm Hg for the aminoguanidine group was increased versus that for the intravenous saline solution group, similar to that for controls, and reduced versus dexamethasone-treated animals. Myocardial water content for the aminoguanidine-treated animals (78.3% +/- 0.4%) was similar to those of intravenous saline-treated animals (78.0% +/- 0. 3%) but greater than those of controls (77.1% +/- 0.2%) and dexamethasone-treated animals (76.7% +/- 0.3%). CONCLUSIONS: Nitric oxide II inhibition with aminoguanidine minimizes the reduction in left ventricular filling that is seen with allograft rejection through a mechanism that is not associated with attenuation of myocardial edema.     

大鼠非压迫性髓核突出模型的建立

目的:设计一种新的非压迫性腰椎间盘髓核突出动物模型。方法:16只SD雄性大鼠随机分对照组和实验组,分别将生理盐水和大鼠自身尾椎髓核混悬液注射到腰椎硬膜外腔,对其马尾神经传导速度和神经根组织形态学进行观察。结果:无明显机械压迫情况下,大鼠硬膜外移植自体髓核能使马尾神经根传导速度和组织形态产生明显改变。结论:本动物模型简单、可靠、费用低廉,为进一步研究腰椎间盘突出症提供了一种动物模型。     

临床症状与髓核突出左右不一致的腰椎间盘突出症的诊治

目的探讨腰椎间盘突出症导致非对称性下肢放射痛的可能原因及术式的选择。方法25例经SCT、MRI检查证实为突出侧与临床症状侧别左右不一致的腰椎间盘突出症患者，均行手术治疗，其中14例行双侧开窗减压探查髓核摘除术，11例仅行突出侧开窗术。结果影像学上髓核突出侧别与术中所见相吻合，但该侧神经根未见到明显压迫或炎性水肿等病理表现；而症状侧无髓核突出，5例神经根未发现异常表现，9例存在不同程度的炎性水肿，其中6例探查发现神经根与对侧髓核不同程度粘连。术后所有患者症状均得到缓解，经过1～5年（平均2．4年）的随访，均无复发。结论SCT结合MRI检查有助于此类腰椎间盘突出症的明确诊断。纤维环无破裂的突出型腰椎间盘突出症，单纯突出侧减压可以获得较好的治疗效果；纤维环破裂的游离型及脱出型腰椎间盘突出症，宜同时行对侧开窗探查。     

借助3D-MRI扫描探讨脊柱(定点)旋转复位法治疗腰椎间盘突出症的临床研究

目的:观察脊柱(定点)旋转复位法治疗腰椎间盘突出症前后突出髓核与神经根的三维空间位置变化,探索该手法治疗腰椎间盘突出症的机制。方法:2009年4月至2011年6月采用脊柱(定点)旋转复位法治疗L5S1腰椎间盘突出症52例,男33例,女19例;年龄19～55岁,平均34.6岁。治疗前后均进行3D-MRI扫描,观察突出髓核与受累神经根解剖关系及核磁冠状位脊柱-骨盆构形变化。结果:MRI显示患者突出髓核与受累神经根解剖关系分为腋侧、肩侧、前方、包围4种类型。手法治疗后触诊患椎椎体位移消失,腰腿痛锐减。所有患者获随访,时间2～28个月,平均12个月,疾病未复发,所有患者恢复原工作或学习。3D-MRI轴位与入院时对照所有病例突出髓核大小形态无显著变化,患者腰椎-骨盆的曲线均有改变。结论:腰椎间盘突出症存在单(多)个椎体位移。椎体位移致腰椎-骨盆构形改变造成髓核及受累神经根生物力学特性的改变,脊柱(定点)旋转复位法纠正患椎椎体位移,恢复了脊柱内外因素平衡,从而达到治疗目的。     

老年腰椎间盘突出症合并腰椎管狭窄症特点与治疗

目的：研究老年腰椎间盘突出症合并椎管狭窄症的临床表现、影像学表现、病理解剖特点及外科治疗方法。方法：回顾分析了147例(男89例，女58例)老年腰椎间盘突出症合并椎管狭窄症患者临床表现、CT、MRI特点及病理解剖特点，病程2周～15年。病变阶段：L4.5 53例，L5S1 42例，L2,4 5例，L2,3 3例，L4,5和L5S1并存44例。全部病例采用椎板减压髓核摘除术，并对治疗方法进行分析。结果：本组92例获得随访，随访时间3个月～3年，平均8个月。术后功能按我们自己制定方法进行评定，优63例，良17例，可10例，差2例，优良率为87％。结论：老年腰椎间盘突出症合并椎管狭窄症具有临床表现不典型、症状与体征不完全一致的特点，并有其特殊的影像学表现。在治疗上应行全椎板减压，髓核摘除，扩大椎管及侧隐窝，使患者获得良好的功能恢复。     

一氧化氮在突出腰椎间盘中的表达及其意义

目的 :研究一氧化氮 (NO)在突出腰椎间盘组织中的含量及组织学定位 ,并对其意义进行探讨。方法 :对 32例腰椎间盘突出患者的突出间盘组织采取两种方法进行研究 :(1) 12例做体外培养 ,用分光光度法测定培养液上清中NO含量 ;(2 ) 2 0例用免疫组化方法对产生NO的细胞类型及组织学定位进行研究。同时对取自 4具新鲜尸体的 12个正常椎间盘采用相同方法做为对照。结果 :突出腰椎间盘组织产生NO的量为 2 0 0 70± 6 5 5 5nmol/g ,正常对照组的NO量为 76 31± 19 49nmol/ g ,两者统计学有显著性差异 (P <0 0 0 1)。免疫组化结果发现 ,2 0例患者椎间盘组织中一氧化氮合成酶表达阳性 16例 ,12个正常椎间盘组织中无表达阳性细胞。结论 :诱导型一氧化氮合成酶主要由突出椎间盘周围的肉芽组织产生 ,阳性细胞主要以成纤维细胞、软骨细胞及淋巴细胞为主。腰椎间盘可自身合成NO ,NO可能在椎间盘退变中起重要作用 ,突出腰椎间盘中的NO主要由突出腰椎间盘周围的肉芽组织产生。     

Extraperitoneal anterolateral discectomy for lumbar disc herniation: indications, techniques, and time-related clinical results.

We analyzed 80 consecutive cases of lumbar disc herniation who underwent an extraperitoneal anterolateral discectomy according to clinical and radiologic parameters. The average follow-up period was 5 1/2 years (range, 1-10 years). The results of 71 patients (89%) were satisfactory through the mid- and long-term follow-up periods, although a very slight decrease in subjective/objective scores and ADL score was found. Satisfactory results were obtained for adolescent and younger male adults and in isolated central or centrolateral herniations with a broad base. From an analysis of the nine reoperated cases, we determined that the contraindications of this technique are a sequestrated or extruded nucleus pulposus and disc protrusion accompanied by posterior dislocation of a vertebral edge fragment. The relative indications include a small, hard posterolateral herniation located just beneath the nerve roots, double-level herniation, disc herniation accompanied by narrow spinal canal, and bulged disc in elder patients. The absolute indications for this surgery are a centrally or centrolaterally protruded disc with a relatively broad base of herniation at single-disc level that is not accompanied by spinal stenosis in adolescent and younger male adults.     

Bone healing regulated by nitric oxide: an experimental study in rats

Nitric oxide has many functions in wound healing and metabolism of bone. In the current study the role of nitric oxide on bone healing was investigated. Thirty-six young adult male Sprague-Dawley rats were divided into three groups: control, nitroso-bovine serum albumin, and aminoguanidine. Five millimeter segmental defects were created in the middle of the right femora. A polyethylene plate and screw posts were used for rigid fixation. Demineralized bone matrix served as the graft material in all groups. Nitroso-bovine serum albumin (an active nitric oxide congener) carried by demineralized bone matrix was applied locally at the defect in the nitroso-bovine serum albumin group. Aminoguanidine (an inducible nitric oxide synthase inhibitor) group received oral aminoguanidine treatment. Formation and healing of bone were determined by radiographic and histologic analyses. In comparison to the control group the healing rate was faster in both experimental groups as indicated by radiographic and histologic data. If accompanied by bone graft with a suitable delivery system, nitric oxide may be useful as a therapeutic adjuvant in clinical situations when local formation of bone is needed. Moreover, when combined appropriately, treatment with orthotopic nitric oxide supplementation and systemic inducible nitric oxide synthase inhibition may enhance bone healing.     

Effect of nucleus pulposus on the neural activity of dorsal root ganglion

STUDY DESIGN: This study was designed to investigate, using neurophysiologic techniques in an in vivo rat model, the effect of application of nucleus pulposus to the nerve root on the neural activity of the dorsal root ganglion and the corresponding receptive fields. OBJECTIVES: To assess a further role of the dorsal root ganglion in mechanisms of radicular pain in lumbar disc herniation. SUMMARY OF BACKGROUND DATA: It has been suggested that the epidural application of autologous nucleus pulposus without mechanical compression causes nerve root inflammation and related radicular pain in lumbar disc herniation. Concerning the dorsal root ganglion, its mechanical hypersensitivity and potential for generating ectopic discharges have been reported. However, the effect of autologous nucleus pulposus on the dorsal root ganglion is uncertain. METHODS: In adult Sprague-Dawley rats spontaneous neural activity was recorded from the surgically exposed L5 dorsal root using electrophysiologic techniques, and the mechanosensitivity of L5 dorsal root ganglia and corresponding receptive fields on the hind paw were measured using calibrated nylon filaments. Autologous nucleus pulposus from the tail or fat was implanted at the L5 nerve root. Neural activity was monitored for 6 hours. RESULTS: Spontaneous neural activity in the nucleus pulposus group gradually increased and showed significant differences compared with the fat group from 2.5 to 6 hours after exposure. The mechanosensitivity of the dorsal root ganglia showed significant increases compared with the fat group. CONCLUSIONS: After application of nucleus pulposus to the nerve root, the dorsal root ganglion demonstrated increased excitability and mechanical hypersensitivity. These results suggest that nucleus pulposus causes excitatory changes in the dorsal root ganglion.     

Does nitric oxide mediate autoimmune destruction of beta-cells? Possible therapeutic interventions in IDDM.

Cytokines have been implicated as immunological effector molecules that induce dysfunction and destruction of the pancreatic beta-cell. The mechanisms of cytokine action on the beta-cell are unknown; however, nitric oxide, resulting from cytokine-induced expression of nitric oxide synthase, has been implicated as the cellular effector molecule mediating beta-cell dysfunction. Nitric oxide is a free radical that targets intracellular iron-containing enzymes, which results in the loss of their function. The cytokine IL-1 beta induces the formation of nitric oxide in isolated rat islets and the insulinoma cell line, Rin-m5F. NMMA and NAME, both inhibitors of nitric oxide synthase, completely protect islets from the deleterious effects of IL-1 beta. These inhibitors are competitive in nature and inhibit both the cytokine-inducible and constitutive isoforms of nitric oxide synthase with nearly identical kinetics. This may preclude their use as therapeutic agents because of increases in blood pressure which result from the inhibition of constitutive nitric oxide synthase activity. Aminoguanidine, an inhibitor of nonenzymatic glycosylation of cellular and extracellular constituents associated with diabetic complications, recently has been reported to inhibit nitric oxide synthase. Aminoguanidine is approximately 40-fold more effective in inhibiting the inducible isoform of nitric oxide synthase, suggesting that aminoguanidine or analogues may serve as potential therapeutic agents to block diseases associated with nitric oxide production by the inducible isoform of nitric oxide synthase. In vivo administration of TNF IL-1 has been shown to induce anti-diabetogenic effects in the NOD mouse. This anti-diabetogenic effect of cytokines appears to conflict with evidence suggesting that cytokines mediate beta-cell dysfunction.(ABSTRACT TRUNCATED AT 250 WORDS)     

经皮内窥镜治疗腰椎间盘突出症的并发症及其处理

目的 总结经皮内窥镜腰椎间盘切除术( PELD)治疗腰椎间盘突出症中出现的并发症,探讨其处理对策.方法 2002年7月至2010年10月采用PELD治疗腰椎间盘突出症患者689例,男性448例,女性241例;年龄13 ～84岁,平均39.8岁.单间隙椎间盘突出669例,双间隙椎间盘突出19例,三间隙椎间盘突出l例.中央型突出66例,旁中央型365例,外侧型242例,极外侧型10例,游离型6例.观察术中和术后并发症及其处理.结果 术中髓核部分残留压迫神经根5例,2例术中改行开窗髓核切除术,2例二期行开窗髓核切除术,1例二期行经椎间孔腰椎体间融合术(TLIF);神经根纤维束部分损伤2例,术后3～6个月内完全恢复;硬脊膜破裂2例,给予缝合皮肤伤口后痊愈.689例患者随访6～96个月,平均33个月.出现椎间隙感染7例,1例保守治疗,4例给予经皮穿刺置管冲洗引流持续局部应用抗生素,2例行后路开窗感染腰椎间盘清除术,均痊愈;术后复发6例,4例患者再次行PELD术,2例患者采用TLIF治疗,术后症状缓解;术后神经根性痛觉过敏和灼样神经根痛19例,经过止痛药物、神经营养药及物理治疗后好转;腰椎管狭窄症行单个节段的PELD术,效果不佳,二期行多节段TLIF治疗10例.结论 术中主要并发症有髓核部分残留压迫神经根、神经根纤维束部分损伤、硬脊膜破裂;术后主要并发症有椎间隙感染、复发、神经根性痛觉过敏和灼样神经根痛等.严格的适应证选择、无菌、熟练操作及术后康复锻炼可以减少并发症的发生.     

Tumor necrosis factor-alpha and interleukin-1beta mediate the production of nitric oxide involved in the pathogenesis of ifosfamide induced hemorrhagic cystitis in mice

PURPOSE: We investigated the participation of nitric oxide in ifosfamide induced hemorrhagic cystitis in mice, and the involvement of tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta in the induction of nitric oxide production in this model. MATERIALS AND METHODS: Hemorrhagic cystitis was induced in mice by 100 to 400 mg./kg. ifosfamide and evaluated 6, 12, 24 or 48 hours thereafter by certain parameters, including vesical edema measurements, microscopic analysis and immunohistochemical testing for inducible nitric oxide synthase. Ifosfamide injected mice were pretreated with 10 to 40 mg./kg. of the nitric oxide synthesis inhibitor L-NG-nitroarginine methyl ester, 80 mg./kg. of mesna, a chemical antagonist of acrolein and the urotoxic metabolite of ifosfamide, 50 microl. antiserum against TNF-alpha and IL-1beta per mouse, 45 mg./kg. of the selective TNF-alpha synthesis inhibitor thalidomide or 200 mg./kg. of the TNF-alpha and IL-1beta synthesis inhibitor pentoxifylline. RESULTS: Ifosfamide induced vesical edema, which peaked 12 hours after ifosfamide injection. Microscopic analysis revealed vascular congestion, edema, hemorrhage, fibrin deposition, neutrophil infiltration and epithelial denudation. Inducible nitric oxide synthase immunolocalization demonstrated intense reactivity to inducible nitric oxide synthase in the cytoplasm of bladder epithelial cells, which showed diffuse necrosis. Pretreatment with mesna reduced the increases in vesical edema, while treatment with L-NG-nitroarginine methyl ester, antiserum to TNF-alpha or IL-1beta, thalidomide or pentoxifylline inhibited vesical edema and microscopic alterations. Antiserum treatments also inhibited the expression of inducible nitric oxide synthase in the urothelium. CONCLUSIONS: Nitric oxide produced by inducible nitric oxide synthase is involved in urothelial damage and in the inflammatory events leading to hemorrhagic cystitis after ifosfamide administration in mice. The induction of inducible nitric oxide synthase in the urothelium appears to depend on the synergistic effect of IL-1beta and TNF-alpha.