Huang Denggao, Guo Na, Peng Yanling, Nie Zhenyu, Gao Yuanhui, Cao Hui, Wang Shunlan, Chen Mei, Wen Xiaohong, Wang Zhaojun, Zheng Linlin, He Haowei, Sun Yong, Chen Yang, Zhang Shufang
Central Laboratory, Haikou Affiliated Hospital of Central South University Xiangya School of Medical, Haikou, Hainan, 570208, P. R. China.
Haikou City Key Laboratory of Clinical Medicine, Haikou, Hainan, 570208, P. R. China.
Sci Rep. 2025 Jul 2;15(1):23049. doi: 10.1038/s41598-025-07387-7.
The role of adipose-derived stromal stem cells (ADSCs) in BLCA progression is unclear. We investigated the effects of invasion, stemness, Epithelial-mesenchymal transition (EMT), and drug resistance of BLCA cells co-cultured with ADSCs for a long period of time. Cells were divided into six groups: ADSCs group, ADSCs: T24 group (10:1, 3:1 and 1:1 groups), ADSCs-derived conditioned medium group (CM) and T24 cell group (T24), and cells in each group were cultured to 14 days, and puromycin (puro) was added to the co-cultured cell to remove ADSCs cells without puro resistance, and then the function of T24 cells (10:1-COC, 3:1-COC and 1:1-COC) after co-culture was studied; CCK-8 assay, Transwell, Wound healing, Flow cytometry, RNA-sequencing, qRT-PCR and Western Bloting assay were used to detect cell proliferation, invasion, migration, apoptosis and cellular mRNA and protein expression levels, respectively. We unexpectedly found enhanced stemness and drug resistance of BLCA cells after prolonged contact culture with ADSCs. T24 cells after co-culture mediated cell proliferation, invasion, EMT, stemness, drug resistance and immune escape by up-regulating MDM2, mt-P53 and PD-L1, compared to CM and T24 groups. The inhibitor Atezo and CP-31,398 eliminated mt-P53 and PD-L1-mediated T24 cell drug resistance and stemness, respectively. This study demonstrated that after prolonged co-culture of BLCA cells with ADSCs, the stemness, drug resistance, and immune evasion of T24 cells were dramatically enhanced, suggesting that long-term resident ADSCs in the bladder cancer tumor microenvironment play a procarcinogenic role.
脂肪来源的间充质干细胞（ADSCs）在膀胱癌（BLCA）进展中的作用尚不清楚。我们研究了长时间与ADSCs共培养的BLCA细胞的侵袭、干性、上皮-间质转化（EMT）和耐药性。细胞分为六组：ADSCs组、ADSCs:T24组（10:1、3:1和1:1组）、ADSCs来源的条件培养基组（CM）和T24细胞组（T24），每组细胞培养至14天，向共培养细胞中加入嘌呤霉素（puro）以去除无嘌呤霉素抗性的ADSCs细胞，然后研究共培养后T24细胞（10:1-COC、3:1-COC和1:1-COC）的功能；分别使用CCK-8检测、Transwell实验、伤口愈合实验、流式细胞术、RNA测序、qRT-PCR和蛋白质免疫印迹实验来检测细胞增殖、侵袭、迁移、凋亡以及细胞mRNA和蛋白质表达水平。我们意外地发现，与ADSCs长时间接触培养后，BLCA细胞的干性和耐药性增强。与CM组和T24组相比，共培养后的T24细胞通过上调MDM2、线粒体P53（mt-P53）和程序性死亡受体配体1（PD-L1）介导细胞增殖、侵袭、EMT、干性、耐药性和免疫逃逸。抑制剂阿替利珠单抗（Atezo）和CP-31,398分别消除了mt-P53和PD-L1介导的T24细胞耐药性和干性。本研究表明，BLCA细胞与ADSCs长期共培养后，T24细胞的干性、耐药性和免疫逃逸显著增强，提示长期驻留在膀胱癌肿瘤微环境中的ADSCs发挥促癌作用。
