PDF(Pubmed)
Abstract：
CMC is the most frequently diagnosed cancer and one of the leading causes of death in non-spayed female dogs. Exploring novel therapeutic agents is necessary to increase the survival rate of dogs with CMC. MPOBA is a BZOP derivative that has a significant anticancer effect in a human cell line. The main goal of this study was to investigate the anticancer properties of MPOBA against two CMC cell lines (REM134 and CMGT071020) using a 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, a wound healing assay, a transwell migration assay, an Annexin V-FITC apoptosis assay with a flow cytometry analysis, a mRNA expression analysis using quantitative real-time PCR (qRT-PCR), and an immunohistochemistry (IHC). According to the accumulated studies, MPOBA caused significant concentration- and time-dependent reductions in cell proliferation and cell migration and induced apoptosis in both CMC cell lines. In gene expression analysis, nine canine genes, including TP53, BCL-2, BAX, epidermal growth factor receptor (EGFR), snail transcription factor (SNAIL), snail-related zinc-finger transcription factor (SLUG), TWIST, E-cadherin, and N-cadherin, were investigated. The mRNA expression results revealed that MPOBA induced upregulation of TP53 and overexpression of the pro-apoptotic gene BAX, together with an inhibition of BCL-2. Moreover, MPOBA also suppressed the mRNA expression levels of SNAIL, EGFR, and N-cadherin and induced upregulation of E-cadherin, crucial genes related to the epithelial-to-mesenchymal transition (EMT). However, there was no significant difference in the IHC results of the expression patterns of vimentin (VT) and cytokeratin (CK) between MPOBA-treated and control CMC cells. In conclusion, the results of the present study suggested that MPOBA exhibited significant anticancer activity by inducing apoptosis in both CMCs via upregulation of TP53 and BAX and downregulation of BCL-2 relative mRNA expression. MPOBA may prove to be a potential candidate drug to be further investigated as a therapeutic agent for CMC.
摘要：
CMC是最常被诊断出的癌症，也是非绝育雌性狗死亡的主要原因之一。探索新的治疗剂对于提高CMC犬的存活率是必要的。MPOBA是在人细胞系中具有显著抗癌作用的BZOP衍生物。本研究的主要目的是使用3-（4,5-二甲基-2-噻唑基）-2,5-二苯基-2H-四唑溴化物（MTT）测定，研究MPOBA对两种CMC细胞系（REM134和CMGT071020）的抗癌特性，伤口愈合试验,Transwell迁移试验，膜联蛋白V-FITC凋亡检测与流式细胞术分析，使用定量实时PCR（qRT-PCR）的mRNA表达分析，和免疫组织化学（IHC）。根据积累的研究，MPOBA引起两种CMC细胞系中细胞增殖和细胞迁移的显着浓度和时间依赖性降低，并诱导细胞凋亡。在基因表达分析中，9个犬基因,包括TP53，BCL-2，BAX，表皮生长因子受体（EGFR），蜗牛转录因子（SNAIL），蜗牛相关锌指转录因子（SLUG），扭曲,E-cadherin,和N-钙黏着蛋白,被调查了。mRNA表达结果显示MPOBA诱导TP53上调和促凋亡基因BAX过表达,与BCL-2的抑制一起。此外，MPOBA还抑制了SNAIL的mRNA表达水平，EGFR,和N-cadherin和诱导的E-cadherin上调，与上皮间质转化（EMT）相关的关键基因。然而,MPOBA处理和对照CMC细胞之间波形蛋白（VT）和细胞角蛋白（CK）表达模式的IHC结果没有显着差异。总之,本研究的结果表明，MPOBA通过上调TP53和BAX以及下调BCL-2相对mRNA表达来诱导两个CMCs的凋亡，从而表现出明显的抗癌活性。MPOBA可能被证明是作为CMC治疗剂进一步研究的潜在候选药物。
