Wang Rui, Hong Ming, Huang Jingyi, Zhou Na, Zhang Yao, Xu Siyuan, Liu Jiaye, Yuan Junjie, Zhang Lusiqi, Huang Linyuan, Huang Ping, Tan Bo, Cao Hong-Ying
School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China.
School of Basic Medical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China.
Front Neurosci. 2021 Oct 1;15:715492. doi: 10.3389/fnins.2021.715492. eCollection 2021.
This research aimed to investigate the neurotoxicity of low-dose cyclophosphamide (CYP) on the urinary bladder of rats by and studies. To establish CYP-induced cystitis rat model, rats were treated with three intraperitoneal injections of CYP (25 mg/kg) in a week. During treatment, the up-down method was used to assess the mechanical withdrawal threshold. On day 8, urodynamic test and bladder smooth muscle contractility study, including the contraction of bladder strips to electrical field stimulation (EFS, 2-64 Hz), carbachol (CCh, 10-10 M) and KCl (120 mM), were performed to evaluate the function of bladder function. Body weight and bladder weight were also recorded. Morphometric analysis using an optical microscope and transmission electron microscope was performed to observe the changes of microstructure and submicrostructure of the bladder. The major pelvic neurons were isolated and treated with acrolein (the main CYP metabolite) to assess apoptosis . RT-PCR assays were used to quantify the mRNA expression levels of and in bladder tissues and primary neurons. After CYP injections, the body weights decreased, but the bladder weights increased in the model group. The mechanical withdrawal threshold of the cystitis model remained at a low level. The morphometric analysis suggested bladder inflammation and neuroinflammation in the bladder of the cystitis rat model. Urodynamic test revealed that, the amplitude, the pressure baseline, the peak pressure and pressure threshold of model rats significantly increased after CYP treatment. The muscle strips of model rats exhibited significantly higher contractility caused by EFS and CCh than the controls. Apoptotic cells appeared at the highest concentration group (100 μM acrolein) after 6 h of acrolein incubation in apoptosis assay of primary neurons. The mRNA expression levels of and were significantly increased in the cystitis rat model and in the acrolein-treated neurons. Low-dose CYP treatment was confirmed to induce nerve injury, which leading to bladder pain and overactive bladder in female rats, and the up-regulation of and may contribute to these pathological changes.
本研究旨在通过行为学和功能学研究，探讨低剂量环磷酰胺（CYP）对大鼠膀胱的神经毒性。为建立CYP诱导的膀胱炎大鼠模型，大鼠每周接受3次腹腔注射CYP（25mg/kg）。治疗期间，采用上下法评估机械撤针阈值。在第8天，进行尿动力学测试和膀胱平滑肌收缩性研究，包括膀胱条对电场刺激（EFS，2 - 64Hz）、卡巴胆碱（CCh，10 - 10M）和氯化钾（120mM）的收缩反应，以评估膀胱功能。同时记录体重和膀胱重量。使用光学显微镜和透射电子显微镜进行形态计量分析，以观察膀胱微观结构和亚微观结构的变化。分离主要盆神经节神经元并用丙烯醛（CYP主要代谢产物）处理，以评估细胞凋亡。采用RT-PCR分析定量膀胱组织和原代神经元中相关基因的mRNA表达水平。CYP注射后，模型组体重下降，但膀胱重量增加。膀胱炎模型的机械撤针阈值维持在较低水平。形态计量分析表明膀胱炎大鼠模型膀胱存在炎症和神经炎症。尿动力学测试显示，CYP处理后模型大鼠的振幅、压力基线、峰值压力和压力阈值显著增加。模型大鼠的肌肉条对EFS和CCh的收缩性明显高于对照组。在原代神经元凋亡试验中，丙烯醛孵育6小时后，最高浓度组（100μM丙烯醛）出现凋亡细胞。膀胱炎大鼠模型和丙烯醛处理的神经元中相关基因的mRNA表达水平显著增加。低剂量CYP处理可导致雌性大鼠神经损伤，进而引起膀胱疼痛和膀胱过度活动，相关基因的上调可能促成这些病理变化。
