Department of Biology, Faculty of Science, Razi University, Kermanshah, Iran.
Gastroenterohepatology Research Center, Shiraz University of Medical Sciences, 71935-1311, Shiraz, Iran.
Biochem Genet. 2021 Oct;59(5):1203-1224. doi: 10.1007/s10528-021-10057-8. Epub 2021 Mar 16.
Numerous researches have extensively studied factors such as microRNAs that lead to cancer. Thus, the current study's purpose is to investigate the biological consequences of hsa-miR-451b inhibition on the properties and functions of gastric cancer stem-like cells. First, gastric cancer stem-like cells were transfected by hsa-miR-451b inhibitor then we used real-time RT-PCR to evaluate its effect on the expression of hsa-miR-451b and two of its direct target genes, Stemness markers such as KLF4, SOX2, CD44, OCT3/4 and NANOG genes and finally Akt, PI3K, Bcl-2, Bax, CASP3 and PCNA genes involved in apoptosis. Here, we conducted a DNA Laddering assay to investigate apoptosis. The level of the MMP-2 and -9 Activities and Migration were examined by Zymography and Transwell invasion assay. HUVEC cells were used to investigate angiogenesis. The outcomes revealed that the level of the MMP-2 and -9 Activities, migration and angiogenesis decreased, but apoptosis was induced by inhibiting hsa-miR-451b. Evaluating KREMEN1 and CASK expression showed that the former increased, and the latter dropped under hsa-miR-451b inhibition. Also, upregulation of the KLF4 and SOX2 and downregulation of the CD44, OCT3/4, and NANOG decreased Self-renewal ability of gastric cancer stem cells under hsa-miR-451b inhibition. Even, under hsa-miR-451b inhibition, downregulation of Akt, PI3K, Bcl-2 and PCNA as well as upregulation of Bax and CASP3 revealed a movement towards apoptosis in MKN-45 stem-like cells. In summary, hsa-miR-451b is an oncomir in the carcinogenesis of gastric cancer stem-like cells and may be suggested as an appropriate therapeutic target for future gastric cancer treatment.
大量研究广泛研究了导致癌症的因素，如 microRNAs。因此，本研究的目的是研究 hsa-miR-451b 抑制对胃癌干细胞样细胞特性和功能的生物学后果。首先，通过 hsa-miR-451b 抑制剂转染胃癌干细胞样细胞，然后使用实时 RT-PCR 评估其对 hsa-miR-451b 及其两个直接靶基因表达的影响，如干性标志物 KLF4、SOX2、CD44、OCT3/4 和 NANOG 基因，最后研究参与凋亡的 Akt、PI3K、Bcl-2、Bax、CASP3 和 PCNA 基因。在这里，我们进行 DNA 梯状电泳分析来研究凋亡。通过明胶酶谱和 Transwell 侵袭实验检测 MMP-2 和 MMP-9 活性和迁移。使用 HUVEC 细胞研究血管生成。结果表明，抑制 hsa-miR-451b 可降低 MMP-2 和 MMP-9 活性、迁移和血管生成，但诱导凋亡。评估 KREMEN1 和 CASK 的表达显示，前者增加，后者在 hsa-miR-451b 抑制时减少。此外，hsa-miR-451b 抑制后，KLF4 和 SOX2 上调，CD44、OCT3/4 和 NANOG 下调，降低了胃癌干细胞自我更新能力。即使在 hsa-miR-451b 抑制下，下调 Akt、PI3K、Bcl-2 和 PCNA 以及上调 Bax 和 CASP3 也显示出 MKN-45 干细胞样细胞向凋亡的转移。总之，hsa-miR-451b 是胃癌干细胞样细胞发生癌变的致癌 miRNA，可作为未来胃癌治疗的合适治疗靶点。
