PRP-1 significantly decreases the ALDHhigh cancer stem cell population and regulates the aberrant Wnt/β-catenin pathway in human chondrosarcoma JJ012 cells
PRP-1 significantly decreases the ALDHhigh cancer stem cell population and regulates the aberrant Wnt/β-catenin pathway in human chondrosarcoma JJ012 cells
复制标题
DOI：
10.3892/or.2019.7172
复制
发表时间：
2019-07-01
期刊：
影响因子：
4.2
通讯作者：
Galoian, K. A.
中科院分区：
文献类型：
--
作者：
Hoyt, A. K.;Moran, A.;Galoian, K. A.
软骨肉瘤是化疗和放疗难治的恶性骨肿瘤，因此，需要新的治疗策略。富含脯氨酸的多肽1（PRP-1）先前已证明在软骨肉瘤的抗肿瘤特性。为了进一步研究PRP-1在软骨肉瘤细胞中的作用，通过分析醛脱氢酶（ALDH）活性（一种确定的CSC标志物）来确定其对癌症干细胞（CSC）群体的影响，该活性与Wnt/β-连环蛋白信号传导的调节相关。用PRP-1处理软骨肉瘤JJ 012细胞后，观察到ALDH（高）CSC显著减少。对于Wnt/β-连环蛋白信号传导基因的RT 2分析仪PCR阵列分析，将细胞分选为：i）大量JJ 012细胞; ii）从未处理的JJ 012细胞分选的ALDH（高）细胞（ALDH（高-未处理））;和iii）从PRP-1处理的JJ 012细胞分选的ALDH（低）细胞（ALDH（低-PRP-1））。与大量JJ 012细胞相比，Wnt/β-连环蛋白信号传导基因的表达水平在ALDH（高未处理）细胞中被确定为下调，而在ALDH（低PRP-1）细胞中被确定为上调。此外，发现参与该途径的两个重要癌基因MMP 7和CCND 2在ALDH（低PRP-1）细胞中下调。免疫细胞化学显示β-连环蛋白定位于PRP-1处理的细胞核中。蛋白质印迹法显示ALDH（low-PRP-1）细胞中β-连环蛋白的表达较大量JJ 012细胞增加。用递增浓度的PRP-1和β-连环蛋白核易位抑制剂CGP 57380处理的细胞的细胞质和细胞核部分的分析表明，PRP-1处理后β-连环蛋白的核易位。此外，用特异性ALDH抑制剂二乙氨基苯甲醛和PRP-1处理JJ 012细胞导致细胞质β-连环蛋白表达显著降低。这表明ALDH失活可能与β-catenin的核转位有关。先前已经记录了通过Wnt途径的失活从间充质干细胞衍生肉瘤。本研究的发现支持了Wnt/β-连环蛋白激活可能在肉瘤中发挥不同作用的观点，限制了与CSC活性降低相关的肿瘤进展。
Chondrosarcomas are malignant bone tumors refractory to chemotherapy and radiation treatment; thus, novel therapeutic strategies are required. Proline-rich polypeptide 1 (PRP-1) has previously demonstrated antitumor properties in chondrosarcoma. To further investigate the role of PRP-1 in chondrosarcoma cells, its effects on cancer stem cell (CSC) populations were determined by analyzing aldehyde dehydrogenase (ALDH) activity, an established marker of CSCs, in association with regulation of the Wnt/beta-catenin signaling. A significant decrease in ALDH(high) CSCs was observed following treatment of chondrosarcoma JJ012 cells with PRP-1. For RT2 profiler PCR array analysis of Wnt/beta-catenin signaling genes, cells were sorted into: i) Bulk JJ012 cells; ii) ALDH(high) cells sorted from untreated JJ012 cells (ALDH(high-untreated)); and iii) ALDH(low) cells sorted from PRP-1-treated JJ012 cells (ALDH(low-PRP-1)). The expression levels of Wnt/beta-catenin signaling genes were determined to be downregulated in the ALDH(high-untreated) cells and upregulated in ALDH(low-PRP-1) cells when compared to the bulk JJ012 cells. Additionally, two important oncogenes involved in this pathway, MMP7 and CCND2, were found to be downregulated in the ALDH(low-PRP-1) cells. Immunocytochemistry demonstrated the localization of beta-catenin in the nuclei of the PRP-1-treated cells. Western blotting indicated increased beta-catenin expression in the ALDH(low-PRP-1) cells compared with the bulk JJ012 cells. Analysis of the cytoplasmic and nuclear fractions of cells treated with increasing concentrations of PRP-1 and beta-catenin nuclear translocation inhibitor CGP57380, suggested the nuclear translocation of beta-catenin following PRP-1 treatment. In addition, treatment of JJ012 cells with a specific ALDH inhibitor, diethylaminobenzaldehyde, and PRP-1 resulted in a significant decrease in cytoplasmic beta-catenin protein expression. This indicated that ALDH inactivation may be associated with the nuclear translocation of beta-catenin. Derivation of sarcomas from mesenchymal stem cells via inactivation of the Wnt pathway has been previously documented. The findings of the present study support the notion that Wnt/beta-catenin activation may serve a differential role in sarcomas, limiting tumor progression in association with decreased CSC activity.