Expression characterization OPA1 expression in breast cancer tissues and its biological function in ER-positive breast cancer cells

doi:10.3877/cma.j.issn.2095-1221.2024.05.003

Abstract

Abstract:

Objective

To analyze the expression of Optic Atrophy 1 （OPA1） in breast cancer and its effect on the biological function of estrogen receptor （ER）-positive breast cancer cells， seeking potential new targets for breast cancer therapy.

Methods

Expression differences of OPA1 in breast cancer and normal/paracancerous tissues were analyzed using publicly available database information （HPA and GSE115144） and tissue microarrays from ER-positive breast cancer patients， respectively. The correlation between OPA1 expression and survival in breast cancer patients was analyzed by the GEPIA and TCGA databases. In cellexperiments， MCF7 and T47D cells were transfected with OPA1 plasmid and small interfering RNA （siRNA）， respectively， and the relative expression levels of mRNA and protein of OPA1 were detected by RT-qPCR and Western blot. Cell proliferation and clone formation ability were detected by CCK-8 assay and plate clone formation assay， respectively. Reactive oxygen species （ROS） level， mitochondrial membrane potential （JC- 1 method） and apoptosis rate were detected by flow cytometry. Two independent samples t-test was used for comparison between groups. Differences among groups were compared by one-way ANOVE analysis， and Dunnett-t test was used for pair-to-group comparisons.

Results

OPA1 was highly expressed in breast cancer tissues compared with paracancerous tissues （P＜ 0.01）， and high OPA1 expression was associated with poor clinical prognosis in breast cancer patients （P＜ 0.05）. In cell experiments， overexpression of OPA1 promoted cell proliferation and clone formation and decreased the ROS levels ［（37.37 ± 1.48）% vs （62.51 ± 2.66）%， P＜ 0.001］， along with an increase in mitochondrial membrane potential ［decrease in the proportion of JC-1 monomers，（7.32 ± 1.84）% vs（17.6 ± 3.35）%， P＜ 0.01］； while inhibition of OPA1 was able to inhibit cell proliferation and clone formation， increase ROS levels ［（77.81 ± 2.37）% vs （58.37 ± 1.36）%， P＜ 0.001］， decreasing mitochondrial membrane potential ［elevated proportion of JC-1 monomer，（28.13 ± 3.47）% vs（15.96 ± 1.14）%， P＜ 0.01］， and promote apoptosis ［（20.10 ± 1.20）% vs （3.85 ± 0.76）%， P＜0.001］.

Conclusion

OPA1 is highly expressed in breast cancer and is significantly associated with poor clinical prognosis. Inhibition of OPA1 expression inhibits the cell proliferation and clone formation of ER-positive breast cancer， while decreasing mitochondrial membrane potential， affecting mitochondrial function，inducing ROS generation andultimately promote cell apoptosis.

Key words: Breast neoplasms, OPA1, Cell Proliferation, Apoptosis, Mitochondria

Chengxin Huang, Li Chen, Yichu Liu, Shuiliang Wang, Xiaofeng Lai. Expression characterization OPA1 expression in breast cancer tissues and its biological function in ER-positive breast cancer cells[J]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2024, 14(05): 275-284.

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Figures/Tables 12

References 26

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基因	序列（5'→3'）	预期扩增长度（bp）
OPA1	上游TGAAAGCATCAAGTTTTTCTTG	105
	下游TGCTGAAGATGGTGAGAAGAAG
β-actin	上游CGCGAGAAGATGACCCAGAT	98
	下游GTACGGCCAGAGGCGTACAG

基因	序列（5'→3'）	预期扩增长度（bp）
OPA1	上游TGAAAGCATCAAGTTTTTCTTG	105
	下游TGCTGAAGATGGTGAGAAGAAG
β-actin	上游CGCGAGAAGATGACCCAGAT	98
	下游GTACGGCCAGAGGCGTACAG

序号	Sense（5'→3'）	Antisense（5'→3'）
siOPA1-1	UUUAUAUUCACUAAGGUCCGG（dT）（dT）	CCGGACCUUAGUGAAUAUAAA（dT）（dT）
siOPA1-2	GCCUGACAUUGUGUGGGAAAU（dT）（dT）	AUUUCCCACACAAUGUCAGGC（dT）（dT）
siOPA1-3	GGAUGGUGCUUGUUGACUUAC（dT）（dT）	GUAAGUCAACAAGCACCAUCC（dT）（dT）
siNC	UUCUCCGAACGUGUCACGU（dT）（dT）	ACGUGACACGUUCGGAGAA（dT）（dT）

序号	Sense（5'→3'）	Antisense（5'→3'）
siOPA1-1	UUUAUAUUCACUAAGGUCCGG（dT）（dT）	CCGGACCUUAGUGAAUAUAAA（dT）（dT）
siOPA1-2	GCCUGACAUUGUGUGGGAAAU（dT）（dT）	AUUUCCCACACAAUGUCAGGC（dT）（dT）
siOPA1-3	GGAUGGUGCUUGUUGACUUAC（dT）（dT）	GUAAGUCAACAAGCACCAUCC（dT）（dT）
siNC	UUCUCCGAACGUGUCACGU（dT）（dT）	ACGUGACACGUUCGGAGAA（dT）（dT）

编号	类型	部位	年龄	肿块大小	TNM分期	分期	ER	PR	CerbB-2	Ki-67	淋巴结
1#	癌	右侧乳腺	35	2	T1cN0M0	Ⅰ	90 % +	70 % +	-	20 % +	0/15
2#	癌	右侧乳腺	56	2.5×2.0×2.0	T2N2M0	Ⅲ	80 % +	10 % +	+	30 % +	6/16
3#	癌	右侧乳腺	70	1.5	T4aNxM1	Ⅳ	95 % +	50 % +	无	20 % +	无

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