Wang Kun, Shi Jian, Chen Zhen, Zhuang Yibo, Xue Dong
Department of Surgical Urology, the Third Affiliated Hospital of Soochow University, Changzhou, 213003, Jiangsu, China.
Department of Pediatrics, the Third Affiliated Hospital of Soochow University, Changzhou, Jiangsu, China.
Int Urogynecol J. 2025 May 30. doi: 10.1007/s00192-025-06169-z.
The Cdk1/p53/p21 feedback loop could play an important role in maintaining a consistently high urinary tract epithelial permeability. This study is aimed at elucidating whether this feedback loop is associated with the inadequate autophagic response and the heightened inflammatory response in interstitial cystitis/bladder pain syndrome (IC/BPS).
To construct the IC/BPS cell model, HTB4 cells were first treated with 1% HO for 1 h, followed by a 24-h exposure to TNFα (10 ng/ml). The protein expression levels of LC3-I/II and Beclin 1 were assessed using Western blot analysis. Autophagosome formation was visualized using monodansylcadaverine (MDC) staining. The secretion levels of inflammatory cytokines, including IL-6, IL-8, and TNFα, were quantified using enzyme-linked immunosorbent assay. Additionally, malondialdehyde (MDA) levels and the activities of antioxidant enzymes (GSH and SOD2) were measured.
Compared with normal urothelial HTB4 cells, the number of autophagosomes and the levels of LC3-I/II and Beclin 1 are significantly increased in IC/BPS cells. These cells also exhibit markedly higher levels of inflammatory markers (TNFα, IL-6, IL-8) and MDA, alongside notably reduced levels of the antioxidants GSH and SOD2. Compared with IC/BPS cells or IC/BPS cells co-transfected with p21 siRNA and Cdk1 control, the IC/BPS cells co-transfected with p21 siRNA and Cdk1 inhibitor exhibit a significant increase in terms of autophagosome numbers and the LC3-I/II level. Additionally, the levels of inflammatory factors are significantly decreased in the IC/BPS cells co-transfected with p21 siRNA and Cdk1 inhibitor.
In IC/BPS, the Cdk1/p53/p21 feedback loop could regulate the level of autophagy and inflammatory response. This discovery identifies promising new therapeutic targets for IC/BPS, which may pave the way for innovative clinical approaches to this complex disorder.
细胞周期蛋白依赖性激酶1（Cdk1）/ 抑癌基因p53 / p21反馈回路可能在维持尿路上皮通透性持续处于高水平方面发挥重要作用。本研究旨在阐明该反馈回路是否与间质性膀胱炎/膀胱疼痛综合征（IC / BPS）中自噬反应不足和炎症反应增强有关。
为构建IC / BPS细胞模型，首先用1%过氧化氢（HO）处理人膀胱上皮细胞系HTB4细胞1小时，随后用肿瘤坏死因子α（TNFα，10纳克/毫升）处理24小时。采用蛋白质免疫印迹分析评估微管相关蛋白1轻链3（LC3 - I/II）和Beclin 1的蛋白表达水平。使用单丹磺酰尸胺（MDC）染色观察自噬体形成。采用酶联免疫吸附测定法定量检测包括白细胞介素6（IL - 6）、白细胞介素8（IL - 8）和TNFα在内的炎性细胞因子的分泌水平。此外，还测量了丙二醛（MDA）水平和抗氧化酶（谷胱甘肽（GSH）和超氧化物歧化酶2（SOD2））的活性。
与正常尿路上皮HTB4细胞相比，IC / BPS细胞中的自噬体数量以及LC3 - I/II和Beclin 水平显著增加。这些细胞还表现出炎性标志物（TNFα、IL - 6、IL - 8）和MDA水平明显更高，同时抗氧化剂GSH和SOD2水平显著降低。与IC / BPS细胞或同时转染p21小干扰RNA（siRNA）和Cdk1对照的IC / BPS细胞相比，同时转染p21 siRNA和Cdk1抑制剂的IC / BPS细胞在自噬体数量和LC3 - I/II水平方面显著增加。此外，同时转染p21 siRNA和Cdk1抑制剂的IC / BPS细胞中炎性因子水平显著降低。
在IC / BPS中，Cdk1 / p53 / p21反馈回路可调节自噬水平和炎症反应。这一发现为IC / BPS确定了有前景的新治疗靶点，可能为针对这种复杂疾病的创新临床方法铺平道路。
