Sung Koangyong, Hosoya Kenji, Murase Yusuke, Deguchi Tatsuya, Kim Sangho, Sunaga Takafumi, Okumura Masahiro
Laboratory of Veterinary Surgery, Department of Veterinary Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo, Japan.
Veterinary Teaching Hospital, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo, Japan.
Vet Comp Oncol. 2022 Mar;20(1):324-335. doi: 10.1111/vco.12779. Epub 2021 Nov 12.
Cancer stem-like cells (CSCs) cause treatment failure in various tumours; however, establishing CSC-targeted therapies has been hampered by difficulties in the identification and isolation of this small sub-population of cells. Recent studies have revealed that tumour cells with low proteasome activity display a CSC phenotype that can be utilized to image CSCs in canines. This study visualizes and reveals the CSC-like properties of tumour cells with low proteasome activity in HMPOS (osteosarcoma) and MegTCC (transitional cell carcinoma), which are canine cell lines. The parent cells were genetically engineered to express ZsGreen1, a fluorescent protein connected to the carboxyl-terminal degron of canine ornithine decarboxylase that accumulates with low proteasome activity (ZsG cells). ZsG cells were imaged and the mode of action of this system was confirmed using a proteasome inhibitor (MG-132), which increased the ZsGreen1 fluorescence intensity. The CSC-like properties of ZsG cells were evaluated on the basis of cell divisions, cell cycle, the expression of CSC markers and tumourigenicity. ZsG cells underwent asymmetric divisions and had a low percentage of G0/G1 phase cells; moreover, ZsG cells expressed CSC markers such as CD133 and showed a large tumourigenic capability. In histopathological analysis, ZsG cells were widely distributed in the tumour samples derived from ZsG cells and in the proliferative regions of the tumours. The results of this study indicate that visualized canine tumour cells with low proteasome activity have a CSC-like phenotype and that this visualization system can be utilized to identify and isolate canine CSCs.
癌症干细胞（CSCs）导致各种肿瘤的治疗失败；然而，由于难以鉴定和分离这一小部分细胞，建立针对CSCs的疗法受到了阻碍。最近的研究表明，蛋白酶体活性低的肿瘤细胞表现出一种CSC表型，可用于对犬类的CSCs进行成像。本研究对犬类细胞系骨肉瘤（HMPOS）和移行细胞癌（MegTCC）中蛋白酶体活性低的肿瘤细胞的CSC样特性进行了可视化和揭示。亲代细胞经过基因工程改造，以表达ZsGreen1，这是一种与犬鸟氨酸脱羧酶的羧基末端降解子相连的荧光蛋白，其会随着蛋白酶体活性降低而积累（ZsG细胞）。对ZsG细胞进行成像，并使用蛋白酶体抑制剂（MG - 132）确认该系统的作用模式，该抑制剂增加了ZsGreen1荧光强度。基于细胞分裂、细胞周期、CSC标志物的表达和致瘤性对ZsG细胞的CSC样特性进行了评估。ZsG细胞进行不对称分裂，G0/G1期细胞百分比低；此外，ZsG细胞表达CD133等CSC标志物，并表现出较大的致瘤能力。在组织病理学分析中，ZsG细胞广泛分布于源自ZsG细胞的肿瘤样本以及肿瘤的增殖区域。本研究结果表明，可视化的蛋白酶体活性低的犬类肿瘤细胞具有CSC样表型，并且该可视化系统可用于鉴定和分离犬类CSCs。
