In vitro RNA interference against beta-catenin inhibits the proliferation of pediatric hepatic tumors.
In vitro RNA interference against beta-catenin inhibits the proliferation of pediatric hepatic tumors.
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DOI：
10.3892/ijo.28.3.715
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发表时间：
2006-03
影响因子：
5.2
通讯作者：
S. Sangkhathat;T. Kusafuka;J. Miao;A. Yoneda;Keigo Nara;Seiji Yamamoto;Y. Kaneda;M. Fukuzawa
中科院分区：
文献类型：
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作者：
S. Sangkhathat;T. Kusafuka;J. Miao;A. Yoneda;Keigo Nara;Seiji Yamamoto;Y. Kaneda;M. Fukuzawa
β-连环蛋白突变已在大多数儿童肝脏恶性肿瘤中被发现，包括肝母细胞瘤（HB）和肝细胞癌（HCC），表明其在该年龄组的肝脏肿瘤发生中发挥重要作用。然而，β-连环蛋白/经典Wnt信号通路在癌细胞肿瘤生长中的作用尚未得到直接研究。为了解决 β-连环蛋白在已建立的儿科 HB 和 HCC 细胞系 HuH-6 和 HepG2（含有突变和过度表达的 β-连环蛋白）中维持恶性表型的能力，我们通过转染短干扰 RNA (siRNA) 进行了一系列体外分析，以生成功能丧失模型。 HuH-7 是另一种源自儿科患者、没有稳定突变的 HB 细胞系，用于比较。 RNA干扰成功地操纵了过度表达的β-连环蛋白的降解。在所有细胞系中，转染 48 小时后，β-连环蛋白 mRNA 被抑制 80-90%，并且其蛋白表达也减少。在HuH-6和HepG2中，治疗后预先存在的β-连环蛋白核积累消失，并且β-连环蛋白下游靶基因c-myc和cyclinD1减少。两种细胞系的体外增殖均受到短暂抑制。相比之下，HuH-7中β-连环蛋白的抑制并没有导致靶基因表达或细胞增殖的显着变化。我们的数据表明，β-连环蛋白可以被认为是该基因突变和过度表达的儿童肝脏肿瘤基因治疗的特定靶点。
Mutations of beta-catenin have been identified in the majority of pediatric hepatic malignancies, including hepatoblastoma (HB) and hepatocellular carcinoma (HCC), suggesting its important contribution in hepatic tumorigenesis in this age group. However, the role of beta-catenin/canonical Wnt signaling pathway in the neoplastic growth of cancer cells has not been directly studied. To address beta-catenin's capability in maintaining the malignant phenotype in established pediatric HB and HCC cell lines, HuH-6 and HepG2, harboring mutated and overexpressed beta-catenin, we carried out a series of in vitro analyses through a transfection of short interfering RNAs (siRNAs) to generate a loss-of-function model. HuH-7, another HB cell line derived from a pediatric patient without a stabilizing mutation was used for comparison. RNA interference successfully manipulated the degradation of overexpressed beta-catenin. In all cell lines, beta-catenin mRNA was suppressed by 80-90% after 48 h of transfection, and a reduction of its protein expression was demonstrated. In HuH-6 and HepG2, the pre-existing beta-catenin nuclear accumulation disappeared and reductions of beta-catenin downstream target genes, c-myc and cyclinD1, were also evidenced after the treatment. The in vitro proliferation of both cell lines was transiently inhibited. In contrast, the suppression of beta-catenin in HuH-7 did not lead to a significant change in the expression of target genes or cellular proliferation. Our data indicate that beta-catenin can be considered a specific target for gene therapy in pediatric hepatic tumors with mutations and overexpression of this gene.