Effects of Mesothelin on proliferation, apoptosis, migration and invasion of breast cancer MDA-MB-231 cells

doi:10.11958/20230452

Abstract

Abstract:

Objective To investigate the effect of Mesothelin on proliferation, apoptosis, migration and invasion of human triple-negative breast cancer cells (TNBC). Methods Immunohistochemical staining was used to detect the expression of Mesothelin in TNBC tissue, paracancer tissue and breast cancer cell lines. TNBC cell line MD-MB-231 cells were transfected with the overexpressed Mesothelin lentivirus plasmid (the mesothelin group) and the control Scramble plasmid (the Scramble group). MD-MB-231 cells with no infected lentivirus were used as the control group. Flow cytometry was used to detect the efficiency of virus infection. The expression of Mesothelin was detected by qPCR, flow cytometry and Western blot assay. Cell proliferation was detected by CCK-8. Annexin V-APC/PI was used to detect apoptosis. The expression of anti-B lymphoblastoma-2 (Bcl-2), anti-Bcl-2 associated X protein (Bax), E-cadherin, N-cadherin and Vimentin were detected by Western blot assay. Cell migration ability was detected by scratch test. The invasive ability of cells was detected by Transwell assay. Results The expression of Mesothelin was significantly higher in TNBC tumor tissue than that in paracancer tissue (P<0.05), and Mesothelin was expressed in breast cancer cell line. Flow cytometry showed that the efficiency of lentivirus infection was more than 90%. Compared with the Scramble group, the expression level of Mesothelin mRNA was increased in the Mesothelin group, and the cell proliferation rate was increased, while the cell apoptosis rate was decreased. The expression of Bax and E-cadherin protein decreased, the expression of Bcl-2, N-cadherin and Vimentin protein increased, and the migration and invasion ability of cells were significantly enhanced (P<0.05). Conclusion Mesothelin can promote the proliferation, inhibit the apoptosis and increase the ability of migration and invasion of human triple-negative breast cancer MDA-MB-231 cells.

Key words: triple negative breastneoplasms, cell proliferation, apoptosis, cell movement, Mesothelin

CLC Number:

R737.9

Figures/Tables 13

References 21

[1]	DERAKHSHAN F, REIS-FILHO J S. Pathogenesis of triple-negative breast cancer[J]. Annu Rev Pathol, 2022, 17:181-204. doi:10.1146/annurev-pathol-042420-093238.
[2]	BIANCHINI G, DE ANGELIS C, LICATA L, et al. Treatment landscape of triple-negative breast cancer - expanded options,evolving needs[J]. Nat Rev Clin Oncol, 2022, 19(2):91-113. doi:10.1038/s41571-021-00565-2.
[3]	BORRI F, GRANAGLIA A. Pathology of triple negative breast cancer[J]. Semin Cancer Biol, 2021, 72:136-145. doi:10.1016/j.semcancer.2020.06.005.
[4]	LI Y, XIAO F, ZHANG A, et al. Oncolytic adenovirus targeting TGF-β enhances anti-tumor responses of mesothelin-targeted chimeric antigen receptor T cell therapy against breast cancer[J]. Cell Immunol, 2020, 348:104041. doi:10.1016/j.cellimm.2020.104041.
[5]	SHEN K, LIU K, WANG Y, et al. Polymorphisms of an oncogenic gene,mesothelin,predict the risk and prognosis of gastric cancer in a Chinese Han population[J]. Arch Toxicol, 2022, 96(7):2097-2111. doi:10.1007/s00204-022-03290-6.
[6]	TCHOU J, WANG L C, SELVEN B, et al. Mesothelin,a novel immunotherapy target for triple negative breast cancer[J]. Breast Cancer Res Treat, 2012, 133(2):799-804. doi:10.1007/s10549-012-2018-4.
[7]	XU X, ZHANG M, XU F, et al. Wnt signaling in breast cancer:biological mechanisms,challenges and opportunities[J]. Mol Cancer, 2020, 19(1):165. doi:10.1186/s12943-020-01276-5.
[8]	LI Y, ZHANG H, MERKHER Y, et al. Recent advances in therapeutic strategies for triple-negative breast cancer[J]. J Hematol Oncol, 2022, 15(1):121. doi:10.1186/s13045-022-01341-0.
[9]	HU Z I, GHAFOOR A, SENGUPTA M, et al. Malignant mesothelioma:advances in immune checkpoint inhibitor and mesothelin-targeted therapies[J]. Cancer, 2021, 127(7):1010-1020. doi:10.1002/cncr.33433.
[10]	HILLIARD T S, KOWALSKI B, IWAMOTO K, et al. Host mesothelin expression increases ovarian cancer metastasis in the peritoneal microenvironment[J]. Int J Mol Sci, 2021, 22(22):12443. doi:10.3390/ijms222212443.
[11]	STRASSER A, VAUX D L. Cell death in the origin and treatment of cancer[J]. Mol Cell, 2020, 78(6):1045-1054. doi:10.1016/j.molcel.2020.05.014.
[12]	GANESH K, MASSAGUÉ J. Targeting metastatic cancer[J]. Nat Med, 2021, 27(1):34-44. doi:10.1038/s41591-020-01195-4.
[13]	UEHARA N, MATSUOKA Y, TSUBURA A. Mesothelin promotes anchorage-independent growth and prevents anoikis via extracellular signal-regulated kinase signaling pathway in human breast cancer cells[J]. Mol Cancer Res, 2008, 6(2):186-193. doi:10.1158/1541-7786.MCR-07-0254.
[14]	YUAN Z, JIANG H, ZHU X, et al. Ginsenoside Rg3 promotes cytotoxicity of paclitaxel through inhibiting NF-κB signaling and regulating Bax/Bcl-2 expression on triple-negative breast cancer[J]. Biomed Pharmacother, 2017, 89:227-232. doi:10.1016/j.biopha.2017.02.038.
[15]	HUO Q, XU C, SHAO Y, et al. Free CA125 promotes ovarian cancer cell migration and tumor metastasis by binding mesothelin to reduce DKK1 expression and activate the SGK3/FOXO3 pathway[J]. Int J Biol Sci, 2021, 17(2):574-588. doi:10.7150/ijbs.52097.
[16]	WANG S, TONG X, LI C, et al. Quaking 5 suppresses TGF-β-induced EMT and cell invasion in lung adenocarcinoma[J]. EMBO Rep, 2021, 22(6):e52079. doi:10.15252/embr.202052079.
[17]	MOHAMMADI GHAHHARI N, SZNURKOWSKA M K, HULO N, et al. Cooperative interaction between ERα and the EMT-inducer ZEB1 reprograms breast cancer cells for bone metastasis[J]. Nat Commun, 2022, 13(1):2104. doi:10.1038/s41467-022-29723-5.
[18]	JINESH G G, BROHL A S. Classical epithelial-mesenchymal transition(EMT) and alternative cell death process-driven blebbishield metastatic-witch (BMW) pathways to cancer metastasis[J]. Signal Transduct Target Ther, 2022, 7(1):296. doi:10.1038/s41392-022-01132-6.
[19]	FANG C, KANG Y. E-Cadherin:Context-dependent functions of a quintessential epithelial marker in metastasis[J]. Cancer Res, 2021, 81(23):5800-5802. doi:10.1158/0008-5472.CAN-21-3302.
[20]	BAEK J, KUMAR S, SCHAFFER D V, et al. N-Cadherin adhesive ligation regulates mechanosensitive neural stem cell lineage commitment in 3D matrices[J]. Biomater Sci, 2022, 10(23):6768-6777. doi:10.1039/d2bm01349e.
[21]	RIDGE K M, ERIKSSON J E, PEKNY M, et al. Roles of vimentin in health and disease[J]. Genes Dev, 2022, 36(7/8):391-407. doi:10.1101/gad.349358.122.

组别	Mesothelin阳性细胞比例/%	Mesothelin mRNA	Mesothelin 蛋白
Control组	8.67±0.97	1.03±0.10	0.17±0.02
Scramble组	9.74±1.69	2.09±0.33	0.17±0.03
Mesothelin组	91.60±1.99^ab	35.02±3.98^ab	0.88±0.04^ab
F	2 627.000^＊＊	210.600^＊＊	517.000^＊＊

组别	Mesothelin阳性细胞比例/%	Mesothelin mRNA	Mesothelin 蛋白
Control组	8.67±0.97	1.03±0.10	0.17±0.02
Scramble组	9.74±1.69	2.09±0.33	0.17±0.03
Mesothelin组	91.60±1.99^ab	35.02±3.98^ab	0.88±0.04^ab
F	2 627.000^＊＊	210.600^＊＊	517.000^＊＊

组别		0 h		24 h
Control组		103.96±5.09		205.41±7.25
Scramble组		101.30±2.03		200.78±11.19
Mesothelin组		102.41±5.93		217.59±7.27
F		0.247		2.942
组别	48 h		72 h		96 h
Control组	255.92±8.16		331.49±20.42		522.72±15.75
Scramble组	235.03±18.07		290.10±17.78		475.78±52.99
Mesothelin组	263.15±17.13		529.60±32.30^ab		756.71±49.05^ab
F	2.797		83.020^＊＊		37.320^＊＊

组别		0 h		24 h
Control组		103.96±5.09		205.41±7.25
Scramble组		101.30±2.03		200.78±11.19
Mesothelin组		102.41±5.93		217.59±7.27
F		0.247		2.942
组别	48 h		72 h		96 h
Control组	255.92±8.16		331.49±20.42		522.72±15.75
Scramble组	235.03±18.07		290.10±17.78		475.78±52.99
Mesothelin组	263.15±17.13		529.60±32.30^ab		756.71±49.05^ab
F	2.797		83.020^＊＊		37.320^＊＊

组别	24 h凋亡率/%		48 h凋亡率/%		12 h愈合率/%
Control组	7.18±0.96		12.72±0.79		12.68±1.39
Scramble组	8.51±0.55		13.77±1.04		12.73±3.50
Mesothelin组	3.44±0.36^ab		8.43±0.84^ab		27.40±0.42^ab
F	46.380^＊＊		29.870^＊＊		45.050^＊＊
组别		24 h愈合率/%		24 h侵袭细胞数/（个/视野）
Control组		24.86±4.59		13.67±0.58
Scramble组		25.94±8.15		17.33±2.31
Mesothelin组		60.03±6.55^ab		69.67±8.50^ab
F		27.590^＊＊		113.200^＊＊