Expression of LncRNA NEAT1 in osteosarcoma cells and study of the mechanism of cellular activity via miR-185-5p

doi:10.3877/cma.j.issn.2096-0263.2023.04.006

Abstract

Abstract:

Objective

To investigate the regulatory mechanism of LncRNA NEAT1 on osteosarcoma cells.

Methods

qRT-PCR was used to detect the differential expression of LncRNA NEAT1 in normal human osteoblasts and osteosarcoma cells. siRNA-NEAT1#1, siRNA-NEAT1#2 and siRNA-NEAT1#3 were constructed and transfected into osteosarcoma cells and their transfection efficiency was verified. Cell viability, apoptosis, migration and invasion ability were detected by CCK-8 assay, flow cytometry and transwell assay, respectively. NEAT1 localization was verified using nucleoplasmic isolation experiments. Predicted target genes of LncRNA NEAT1 was detected by bioinformatics prediction website, followed by dual luciferase assay to verify. Co-transfection of miR-185-5p inhibitor and siRNA-NEAT1 into osteosarcoma cells was applied to verify that LncRNA NEAT1 regulated the biological behavior of osteosarcoma cells through sponge binding of the target gene miR-185-5p.

Results

LncRNA NEAT1 was highly expressed in osteosarcoma cells. Knockdown of LncRNA NEAT1 expression in osteosarcoma cells inhibited cell viability, migration, invasion and promoted apoptosis. LncRNA NEAT1 sponged binding to miR-185-5p, knockdown of LncRNA NEAT1 expression with miR-185-5p in osteosarcoma cells partially reversed the above cellular behaviors.

Conclusion

Knockdown of miR-185-5p expression level could partially reverse the inhibitory effect of siRNA-NEAT1 on osteosarcoma cell viability.

Key words: Osteosarcoma, LncRNA NEAT1, MiR-185-5p, Cell viability

Mingxian Qiu, Xiao Kang, Lei Wang. Expression of LncRNA NEAT1 in osteosarcoma cells and study of the mechanism of cellular activity via miR-185-5p[J]. Chinese Journal of Geriatric Orthopaedics and Rehabilitation(Electronic Edition), 2023, 09(04): 233-239.

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Gene	Forward primer（5’-3’）	Reverse primer（5’-3’）
NEAT1	GCGAGGTGCCTTTACTACAT		TGGAACCCAGAAGACAGA
miR-185-5p	CGCTGGAGAGAAAGGCAGT	GTGCAGGGTCCGAGGT
GAPDH	ACGGCAAGTTCAACGGCACAG	GACGCCAGTAGACTCCACGACA
U6	CTCGCTTCGGCAGCACA	AACGCTTCACGAATTTGCGT

Gene	Forward primer（5’-3’）	Reverse primer（5’-3’）
NEAT1	GCGAGGTGCCTTTACTACAT		TGGAACCCAGAAGACAGA
miR-185-5p	CGCTGGAGAGAAAGGCAGT	GTGCAGGGTCCGAGGT
GAPDH	ACGGCAAGTTCAACGGCACAG	GACGCCAGTAGACTCCACGACA
U6	CTCGCTTCGGCAGCACA	AACGCTTCACGAATTTGCGT

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