OBJECTIVE: This study aims to investigate the potential and mechanisms of cyclanoline (Cyc) in overcoming cisplatin resistance.
METHODS: Cisplatin-resistant T24 and BIU-87 cell models (T24/DR and BIU-87/DR) were established by increasing gradual concentration. Western Blot (WB) assessed the phosphorylation of STAT3, JAK2, and JAK3. T24/DR and BIU-87/DR cell lines were treated with selective STAT3 phosphorylation modulators, and cell viability was evaluated by CCK-8. Cells were subjected to cisplatin, Cyc, or their combination. Immunofluorescence (IHC) examined p-STAT3 expression. Protein and mRNA levels of apoptosis-related and cell cycle-related factors were measured. Changes in proliferation, invasion, migration, apoptosis, and cell cycle were monitored. In vivo, subcutaneous tumor transplantation models in nude mice were established, assessing tumor volume and weight. Changes in bladder cancer tissues were observed through HE staining, and the p-STAT3 was assessed via WB and IHC.
RESULTS: Cisplatin-resistant cell lines were successfully established, demonstrating increased phosphorylation of STAT3, JAK2, and JAK3. Cisplatin or Cyc treatment decreased p-STAT3, inhibited invasion and migration, and induced apoptosis and cell cycle arrest in the G0/G1 phase in vitro. In vivo, tumor growth was significantly suppressed, with extensive tumor cell death. IHC and WB consistently showed a substantial downregulation of STAT3 phosphorylation. These changes were more pronounced when cisplatin and Cyc were administered in combination.
CONCLUSIONS: Cyc reverses cisplatin resistance via JAK/STAT3 inhibition in bladder cancer, offering a potential clinical strategy to enhance cisplatin efficacy in treating bladder cancer.
目的:本研究旨在探讨环胆碱(Cyc)克服顺铂耐药的潜力和机制。
方法:通过逐渐增加浓度建立顺铂耐药T24和BIU-87细胞模型(T24/DR和BIU-87/DR)。蛋白质印迹(WB)评估STAT3、JAK2和JAK3的磷酸化。用选择性STAT3磷酸化调节剂处理T24/DR和BIU-87/DR细胞系,并通过CCK-8评估细胞活力。细胞接受顺铂,Cyc,或他们的组合。免疫荧光(IHC)检测了p-STAT3的表达。检测凋亡相关因子和细胞周期相关因子的蛋白和mRNA水平。增殖的变化,入侵,迁移,凋亡,监测细胞周期。在体内,建立裸鼠皮下移植瘤模型,评估肿瘤体积和重量。通过HE染色观察膀胱癌组织的变化,通过WB和IHC评估p-STAT3。
结果:成功建立顺铂耐药细胞系,显示STAT3、JAK2和JAK3的磷酸化增加。顺铂或Cyc治疗降低p-STAT3,抑制侵袭和迁移,并在体外诱导G0/G1期细胞凋亡和细胞周期阻滞。在体内,肿瘤生长被显著抑制,肿瘤细胞广泛死亡。IHC和WB一致显示STAT3磷酸化的显著下调。当顺铂和Cyc联合给药时,这些变化更为明显。
结论:Cyc通过抑制JAK/STAT3逆转膀胱癌顺铂耐药,为提高顺铂治疗膀胱癌的疗效提供了潜在的临床策略。