Effects of long non-coding RNA KIAA0125 on proliferation and apoptosis of acute myeloid leukemia U937 cells

doi:10.12307/2025.510

Abstract

Abstract: BACKGROUND: U937 cells can be used as a cell model for studying the biological characteristics, signaling pathways, and therapeutic targets of acute myeloid leukemia. Although it has been reported that long non-coding RNA KIAA0125 is highly expressed in acute myeloid leukemia, its biological function in U937 cells remains unclear, and its mechanism of action in the occurrence and development of acute myeloid leukemia needs to be further clarified.
OBJECTIVE: To investigate the expression level of long non-coding RNA KIAA0125 in peripheral blood of patients with acute myeloid leukemia and its effect on the proliferation and apoptosis of U937 cells.
METHODS: RNA-sequencing was used to analyze the bone marrow monocyte samples from acute myeloid leukemia patients, and the differentially expressed gene long non-coding RNA KIAA0125 was screened. The expression of long non-coding RNA KIAA0125 in peripheral blood of patients with acute myeloid leukemia was detected by qRT-PCR. The relationship between long non-coding RNA KIAA0125 mRNA expression and prognosis in bone marrow cells of 173 acute myeloid leukemia patients and 70 healthy people was statistically analyzed by GEPIA database. Subsequently, recombinant lentivirus technology and CRISPR/Cas9-SAM technology were used to construct U937 cell lines with knockdown/overexpression of long non-coding RNA KIAA0125. qRT-PCR was used to detect the knockdown/overexpression efficiency of long non-coding RNA KIAA0125. Next, CCK-8 assay, flow cytometry, and western blot assay were used to detect the effects of knockdown/overexpression of long non-coding RNA KIAA0125 on the proliferation and apoptosis of U937 cells. Finally, western blot assay was used to detect the effect of knockdown/overexpressed long non-coding RNA KIAA0125 on Wnt/β-catenin signaling pathway-related proteins.
RESULTS AND CONCLUSION: (1) The results of qRT-PCR showed that long non-coding RNA KIAA0125 was highly expressed in peripheral blood of acute myeloid leukemia patients. The results of GEPIA database showed that long non-coding RNA KIAA0125 was highly expressed in bone marrow cells of acute myeloid leukemia patients, and the high expression group had worse overall survival. (2) The knockdown efficiency of long non-coding RNA KIAA0125 in knockdown group was 70%, and the U937 cells that stably down-regulated long non-coding RNA KIAA0125 expression were successfully constructed. The expression of long non-coding RNA KIAA0125 in overexpression group was four times that of vector group, and stable U937 cells were successfully constructed. (3) Knockdown of long non-coding RNA KIAA0125 inhibited the proliferation of U937 cells and promoted their apoptosis. Overexpression of long non-coding RNA KIAA0125 promoted the proliferation of U937 cells but had no significant effect on the apoptosis of U937 cells. (4) Knockdown of long non-coding RNA KIAA0125 inhibited the activity of Wnt/β-catenin signaling pathway, while overexpression of long non-coding RNA KIAA0125 activated Wnt/β-catenin signaling pathway. These results confirm that long non-coding RNA KIAA0125 is highly expressed in acute myeloid leukemia peripheral blood. Long non-coding RNA KIAA0125 may affect the proliferation and apoptosis of U937 cells by regulating the Wnt/β-catenin signaling pathway, and may be a potential prognostic marker for acute myeloid leukemia.

Key words: acute myeloid leukemia, lncKIAA0125, Wnt/β-catenin, U937 cell, proliferation

CLC Number:

Hu Huali, Deng Fahua, Liu Yuancheng, Wang Siqi, Zhang Jingxin, Lu Tingting, Huang Hai, Wei Sixi. Effects of long non-coding RNA KIAA0125 on proliferation and apoptosis of acute myeloid leukemia U937 cells[J]. Chinese Journal of Tissue Engineering Research, 2025, 29(19): 3983-3991.

Figures/Tables 4

2.4 过表达lncKIAA0125对U937细胞增殖及凋亡的影响利用CRISPR/Cas9-SAM系统构建稳定过表达lncKIAA0125的U937细胞系，qRT-PCR验证lncKIAA0125过表达效率，sgRNA-1的过表达效率最高，因此选择sgRNA-1(即OE-lncKIAA0125组)进行后续研究(图3A)。CCK-8结果显示，与Vector组相比，OE-lncKIAA0125组的U937细胞增殖能力增加(图3B)，流式细胞术结果显示Vector组与OE-lncKIAA0125组的U937细胞凋亡率无显著差异(图3C，D)，Western blot结果显示，与Vector组相比，OE-lncKIAA0125组增殖相关蛋白PCNA表达增加，但抗凋亡蛋白Bcl-2及促凋亡蛋白Bax表达无显著差异(图3E，F)。 2.5 敲低lncKIAA0125对Wnt/β-catenin通路的影响研究表明，lncKIAA0125可通过Wnt/β-catenin通路调节结直肠癌的生长和转移[21]，说明lncKIAA0125可能通过影响Wnt/β-catenin信号通路来发挥作用。因此，该研究拟进一步探讨在急性髓系白血病细胞系——U937细胞中敲低lncKIAA0125对Wnt/β-catenin通路的影响。Western blot结果显示，与sh-NC组相比，sh-lncKIAA0125组的β-catenin、TCF4、c-Myc、Cyclin D1蛋白表达水平降低(图4A，B)。qRT-PCR结果显示，与sh-NC组相比，sh-lncKIAA0125组Wnt/β-catenin通路的下游基因c-Myc、Cyclin D1 mRNA表达水平降低(图4C)。以上结果提示，在敲低lncKIAA0125后，抑制了Wnt/β-catenin信号通路，导致通路中的关键蛋白β-catenin降低，与β-catenin相结合的蛋白TCF4水平降低，从而抑制下游基因c-Myc、Cyclin D1的转录，进一步抑制U937细胞的增殖并促进细胞凋亡。 2.6 过表达lncKIAA0125对Wnt/β-catenin通路的影响 Western blot结果显示，与Vector组相比，OE-lncKIAA0125组的β-catenin、TCF4、c-Myc、Cyclin D1蛋白表达水平增加(图5A，B)。qRT-PCR结果显示，与Vector组相比，OE-lncKIAA0125组Wnt/β-catenin通路的下游基因c-Myc、Cyclin D1 mRNA表达水平增加(图5C)。以上结果提示，在过表达lncKIAA0125后，激活了Wnt/β-catenin信号通路，使通路中的关键蛋白β-catenin增加，与β-catenin相结合的蛋白TCF4水平增加，从而促进下游基因c-Myc、Cyclin D1的转录，进一步促进了U937细胞的增殖。"

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