ZEB1 promotes the proliferation， migration and invasion of prostate cancer cells by regulating Wnt/β-catenin signaling pathway

doi:10.3877/cma.j.issn.2095-1221.2025.03.004

Abstract

Abstract:

Objective

To explore whether zinc finger E-box binding homeobox 1 （ZEB1）promotes the proliferation， migration and invasion of prostate cancer cells by regulating the Wnt/β-catenin signaling pathway.

Methods

siZEB1 and Wnt3a overexpression plasmids were transfected into PC-3 and LNCaP prostate cancer cells， and named control group， the siZEB1 group， and the siZEB1+OE-Wnt3a group， respectively. Real-time fluorescence quantitative polymerase chain reaction （RT-qPCR） was used to detect the mRNA expression levels of ZEB1， Wnt3a， β-catenin，c-Myc， Cyclin D1， E-cadherin， N-cadherin in human normal prostate epithelial cell RWPE- 1 and PC-3， LNCaP prostate cancer cells. Western blot was used to detect the protein contents of Wnt3a，β-catenin， c-Myc， Cyclin D1， E-cadherin and N-cadherin in PC-3 as well as LNCaP prostate cancer cells. Cell counting kit-8 （CCK-8） assay and plate clone assay were used to detect the proliferation changes of prostate cancer cells after different treatments. Scratch assay and Transwell assay were used to detect the invasion and migration ability of prostate cancer cells after different treatments.Independent sample t-test was used for comparison between two groups， one- way ANOVA was used for comparison among multiple groups， and LSD-t test was used for pairwise comparison between groups.

Results

Compared with RWPE1 cells， the expression level of ZEB1 mRNA in prostate cancer cells PC-3 and LNCaP were upregulated [（1.99 ± 0.11），（1.96 ± 0.13） vs （1.00 ± 0.05）］（both P ＜ 0.05）. Compared with the control group， the viability， the colony numbers [PC-3 cells：（21.33 ± 2.05） vs （48.67 ± 3.68） cells， LNCaP cells：（21.67 ± 2.87） vs （47.33 ± 2.05） cells］， the migration rate [PC-3 cells：（30.67 ± 2.49）% vs （48.33 ± 2.49）%， LNCaP cells：（28.33 ± 3.40） %vs （43.00 ± 2.94）%］， and the number of invaded cells [PC-3 cells：（43.00 ± 3.56） vs （64.00 ± 3.74）cells， and LNCaP cells：（42.67 ± 4.50） vs （61.33 ± 2.62）cells］ were decreased in cells transfected with siZEB1 （all P ＜ 0.05）， while the apoptosis rate [PC- 3 cells：（9.63 ± 0.66）% vs （1.25 ± 0.34）%，LNCaP cells：（9.34 ± 0.72）% vs （1.54 ± 0.42）%］ were increased （all P ＜ 0.001）. Compared with the control group， the expression levels of protein and mRNA of Wnt3a， c-Myc， Cyclin D1， N-cadherin andthe expression of β-catenin protein was down-regulated， while the expression levels of of E-cadherin protein and mRNA in PC-3 and LNCaP cells after transfected with siZEB1 （all P ＜ 0.05），which could be partially restored after co-transfection of siZEB1 and OE- Wnt3a. After co-transfection of siZEB1 and OE-Wnt3a， the viability， the colony number [PC-3 cells：（35.67 ± 2.05） vs （18.33 ±2.87） cells； LNCaP cells：（31.33 ± 2.52） vs （15.00 ± 3.61） cells］， the migration rate [PC- 3 cells：（50.67 ± 3.30）% vs （43.67 ± 3.68）%； LNCaP cells：（53.67 ± 3.21）% vs （41.33 ± 5.50）%］（all P ＜ 0.05）， the invaded cell number [PC-3 cells：（35.67 ± 1.24） vs （24.00 ± 2.45） cells； LNCaP cells：（34.33 ± 4.04） vs （22.00 ± 3.00） cells］（all P ＜ 0.05） were increased， and the apoptosis rate was decreased [PC-3 cells：（6.74 ± 0.73）% vs （9.75 ± 1.08）% ； LNCaP cells：（8.30 ± 0.47）%vs （11.50 ± 0.56）%］（all P ＜ 0.05） in PC-3 and LNCaP cells compared with siZEB1 group.

Conclusion

ZEB1 plays a significant role in the proliferation， migration， invasion， and apoptosis of prostate cancer cells by regulating the Wnt/β-catenin signaling pathway. Knockdown of ZEB1 can significantly inhibit the malignant phenotype of prostate cancer cells， while overexpression of Wnt3a can partially reverse this inhibitory effect.

Key words: Prostate cancer, ZEB1, Wnt/β-catenin signaling pathway, Proliferation, Migration

Kai Liu, Xiangge Li, Cheng Wang, Run Tang. ZEB1 promotes the proliferation， migration and invasion of prostate cancer cells by regulating Wnt/β-catenin signaling pathway[J]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2025, 15(03): 157-166.

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Figures/Tables 14

References 20

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基因	序列（ 5' → 3'）	预期扩增长度（ bp）
ZEB1	上游GATGATGAATGCGAGTCAGATGC	86
	下游ACAGCAGTGTCTTGTTGTTGT
β-catenin	上游CTGAGGAGCAGCTTCAGTCC	161
	下游CCATCAAATCAGCTTGAGTAGCC
Wnt3a	上游TAGGAAGAGAGGTCCAGCCC	186
	下游CTCCAGGAAAGCGGACCATT
c-MYC	上游GGCTCCTGGCAAAAGGTCA	119
	下游CTGCGTAGTTGTGCTGATGT
Cyclin D1	上游GCTGCGAAGTGGAAACCATC	135
	下游CCTCCTTCTGCACACATTTGAA
N-cadherin	上游GGGAAATGGAAACTTGATGGC	164
	下游AAATCTGCAGGCTCACTGCT
E-cadherin	上游CGAGAGCTACACGTTCACGG	119
	下游GGGTGTCGAGGGAAAAATAGG
GAPDH	上游ACAACTTTGGTATCGTGGAAGG	101
	下游GCCATCACGCCACAGTTTC

基因	序列（ 5' → 3'）	预期扩增长度（ bp）
ZEB1	上游GATGATGAATGCGAGTCAGATGC	86
	下游ACAGCAGTGTCTTGTTGTTGT
β-catenin	上游CTGAGGAGCAGCTTCAGTCC	161
	下游CCATCAAATCAGCTTGAGTAGCC
Wnt3a	上游TAGGAAGAGAGGTCCAGCCC	186
	下游CTCCAGGAAAGCGGACCATT
c-MYC	上游GGCTCCTGGCAAAAGGTCA	119
	下游CTGCGTAGTTGTGCTGATGT
Cyclin D1	上游GCTGCGAAGTGGAAACCATC	135
	下游CCTCCTTCTGCACACATTTGAA
N-cadherin	上游GGGAAATGGAAACTTGATGGC	164
	下游AAATCTGCAGGCTCACTGCT
E-cadherin	上游CGAGAGCTACACGTTCACGG	119
	下游GGGTGTCGAGGGAAAAATAGG
GAPDH	上游ACAACTTTGGTATCGTGGAAGG	101
	下游GCCATCACGCCACAGTTTC

分组	实验次数	PC-3				LNCap
分组	实验次数	0 h	24 h	48 h	72 h	0 h	24 h	48 h	72 h
对照	3	0.12 ± 0.02	0.85 ± 0.04	1.01 ± 0.06	1.12 ± 0.06	0.10 ± 0.01	0.85 ± 0.03	1.03 ± 0.06	1.07 ± 0.05
siZEB1	3	0.10 ± 0.01	0.68 ± 0.02	0.79 ± 0.04	0.90 ± 0.05	0.11 ± 0.01	0.70 ± 0.03	0.79 ± 0.06	0.86 ± 0.06
t值		1.189	6.360	5.023	5.009	0.673	5.420	5.041	4.610
P值		0.300	0.003	0.007	0.007	0.538	0.006	0.007	0.010
F 组间/F时间/F 交互			5358.384/649.613/9.163				3205.177/724.065/13.224
P 组间/P 时间/P 交互			＜ 0.001/＜ 0.001/0.002				＜ 0.001/＜ 0.001/＜0.001

分组	实验次数	PC-3				LNCap
分组	实验次数	0 h	24 h	48 h	72 h	0 h	24 h	48 h	72 h
对照	3	0.12 ± 0.02	0.85 ± 0.04	1.01 ± 0.06	1.12 ± 0.06	0.10 ± 0.01	0.85 ± 0.03	1.03 ± 0.06	1.07 ± 0.05
siZEB1	3	0.10 ± 0.01	0.68 ± 0.02	0.79 ± 0.04	0.90 ± 0.05	0.11 ± 0.01	0.70 ± 0.03	0.79 ± 0.06	0.86 ± 0.06
t值		1.189	6.360	5.023	5.009	0.673	5.420	5.041	4.610
P值		0.300	0.003	0.007	0.007	0.538	0.006	0.007	0.010
F 组间/F时间/F 交互			5358.384/649.613/9.163				3205.177/724.065/13.224
P 组间/P 时间/P 交互			＜ 0.001/＜ 0.001/0.002				＜ 0.001/＜ 0.001/＜0.001

分组	实验次数	PC-3 细胞
分组	实验次数	Wnt3a	β-catenin	c-Myc	Cyclin D1	E-cadherin	N-cadherin
对照	3	1.00 ± 0.05	1.00 ± 0.01	1.00 ± 0.02	1.00 ± 0.01	1.00 ± 0.04	1.00 ± 0.01
siZEB1	3	0.32 ± 0.03^a	0.31 ± 0.02^a	0.25 ± 0.01^a	0.29 ± 0.01a	2.73 ± 0.11^a	0.24 ± 0.04^a
siZEB1+OE-Wnt3a	3	0.74 ± 0.05^ab	0.82 ± 0.02^ab	0.76 ± 0.03^ab	0.78 ± 0.02^ab	1.89 ± 0.07^ab	0.68 ± 0.01^ab
F值		20.162	59.315	40.244	149.33	24.592	34.846
P值		＜ 0.001	＜ 0.001	＜ 0.001	＜ 0.001	＜ 0.001	＜ 0.001
分组	实验次数	LNCaP 细胞
分组	实验次数	Wnt3a	β-catenin	c-Myc	Cyclin D1	E-cadherin	N-cadherin
对照	3	1.00 ± 0.01	1.00 ± 0.02	1.00 ± 0.03	1.00 ± 0.02	1.00 ± 0.02	1.00 ± 0.01
siZEB1	3	0.27 ± 0.02^a	0.27 ± 0.02^a	0.35 ± 0.02^a	0.24 ± 0.02^a	2.43 ± 0.07^a	0.27 ± 0.01^a
siZEB1+OE-Wnt3a	3	0.67 ± 0.02^ab	0.82 ± 0.03^ab	0.73 ± 0.03^ab	0.73 ± 0.04^ab	1.72 ± 0.02^ab	0.70 ± 0.01^ab
F值		37.032	53.335	35.217	32.116	36.475	43.792
P值		＜ 0.001	＜ 0.001	＜ 0.001	＜ 0.001	＜ 0.001	＜ 0.001

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