■微囊藻毒素-亮氨酸精氨酸(MC-LR)是在全球水源中发现的蓝细菌肝毒性毒素,包括在泰国东北部,其中opisthorchiasis相关的胆管癌(CCA)是最普遍的。MC-LR是一种潜在的致癌物;然而,其在肝吸虫相关CCA中的参与仍然不明确。这里,我们旨在评估MC-LR通过Wnt/β-catenin通路对CCA进展的影响。
■细胞分裂,迁移,细胞周期过渡,通过MTT法体外评价MC-LR转运体的表达,伤口愈合试验,流式细胞术,免疫荧光染色,分别。用1、10、100和1,000nMMC-LR处理培养细胞24小时后,使用免疫印迹和qRT-PCR分析研究MC-LR对Wnt/β-catenin信号通路的增殖作用。免疫组织化学用于确定与邻近组织相比的CCA组织中的β-连环蛋白表达。
■人永生化的胆管细胞(MMNK-1)和从opisthorchiasis相关的CCA(KKU-213B)建立的人细胞系表达MC-LR转运蛋白和内化的MC-LR。暴露于10nM和100nM的MC-LR显着增强了两种细胞系中的细胞分裂和迁移(P<0.05),并显着提高了S期细胞的百分比(P<0.05)。MC-LR通过激活Wnt/β-catenin信号通路和抑制磷酸酶活性来提高PP2A的表达。抑制β-连环蛋白破坏复合物基因(Axin1和APC)导致β-连环蛋白及其下游靶基因(CyclinD1和c-Jun)上调。MSAB对Wnt/β-联蛋白信号传导的抑制证实了这些结果。此外,与邻近区域相比,β-catenin在癌组织中显著表达(P<0.001)。
■我们的发现表明MC-LR通过Wnt/β-catenin通路促进细胞增殖和CCA的进展。需要使用体内实验进行进一步评估以证实该观察结果。这一发现可以提高对MC-LR摄入量和CCA风险的健康认识。
UNASSIGNED: Microcystin-leucine arginine (MC-LR) is a cyanobacterial hepatotoxic toxin found in water sources worldwide, including in northeastern Thailand, where opisthorchiasis-associated cholangiocarcinoma (CCA) is most prevalent. MC-LR is a potential carcinogen; however, its involvement in liver fluke-associated CCA remains ambiguous. Here, we aimed to evaluate the effect of MC-LR on the progression of CCA via the Wnt/β-catenin pathway in vitro.
UNASSIGNED: Cell division, migration, cell cycle transition, and MC-LR transporter expression were evaluated in vitro through MTT assay, wound healing assay, flow cytometry, and immunofluorescence staining, respectively. Following a 24-h treatment of cultured cells with 1, 10, 100, and 1,000 nM of MC-LR, the proliferative effect of MC-LR on the Wnt/β-catenin signaling pathway was investigated using immunoblotting and qRT-PCR analysis. Immunohistochemistry was used to determine β-catenin expression in CCA tissue compared to adjacent tissue.
UNASSIGNED: Human immortalized cholangiocyte cells (MMNK-1) and a human cell line established from opisthorchiasis-associated CCA (KKU-213B) expressed the MC-LR transporter and internalized MC-LR. Exposure to 10 nM and 100 nM of MC-LR notably enhanced cells division and migration in both cell lines (P < 0.05) and markedly elevated the percentage of S phase cells (P < 0.05). MC-LR elevated PP2A expression by activating the Wnt/β-catenin signaling pathway and suppressing phosphatase activity. Inhibition of the β-catenin destruction complex genes (Axin1 and APC) led to the upregulation of β-catenin and its downstream target genes (Cyclin D1 and c-Jun). Inhibition of Wnt/β-catenin signaling by MSAB confirmed these results. Additionally, β-catenin was significantly expressed in cancerous tissue compared to adjacent areas (P < 0.001).
UNASSIGNED: Our findings suggest that MC-LR promotes cell proliferation and progression of CCA through Wnt/β-catenin pathway. Further evaluation using invivo experiments is needed to confirm this observation. This finding could promote health awareness regarding MC-LR intake and risk of CCA.