Expression Of SALL4 In Laryngeal Cancer Tissues And Its Effects On The Migration And Invasion Of Laryngeal Cancer Cells

Objective The purpose of this study was to investigate the expression of human spalt like transcription factor 4(SALL4)in laryngeal squamous cell carcinoma(LSCC)tissues and its effect on LSCC cell proliferative activity,invasion,migration ability and epithelial to mesenchymal transition(EMT).Methods 1.Tissue specimens of 48 laryngeal cancer patients(n=48)and their corresponding paraneoplastic normal tissue specimens(n=48)treated in our hospital from January 2020 to December 2022 were collected and divided into tumor(T)group and normal(N)group.The expression of SALL4 gene and protein in laryngeal cancer tissue specimens and paraneoplastic tissue specimens were detected by q RT-PCR and Western blot method.2.Human bronchial epithelial-like cell line(16HBE)and five laryngeal cancer cell lines(AMC-HN-8,TU686,TU177,M4 E,LSC-1)were cultured.QRT-PCR and Western blot assays were performed to detect the expression levels of SALL4 mRNA and protein in 16 HBE and each laryngeal cancer cell line,and two cell lines that met the experimental objectives were selected for subsequent studies.The specific experimental groups were as follows: laryngeal cancer cell lines without any treatment were used as blank control group(Control group),laryngeal cancer cell lines transfected with sh-NC sequences only were used as negative control group(sh-NC group),and laryngeal cancer cell lines transfected with sh RNA-SALL4 sequences only were used as experimental group(sh-SALL4 group).The effect of SALL4 knockdown in AMC-HN-8 and TU686 cells was verified using q RT-PCR and Western blot.MTT assay was performed to detect the effect of SALL4 knockdown on the proliferation activity of laryngeal cancer cell lines;cell scratch assay was performed to detect the migration ability of AMC-HN-8 and TU686 cells after SALL4 knockdown;Transwell assay was performed to detect the invasion ability of AMCHN-8 and TU686 cells after SALL4 knockdown;Western blot assay was performed to detect the expression levels of EMT-related proteins(including E-cadherin and vimentin)in LSCC cells after SALL4 knockdown.Results 1.SALL4 mRNA and protein expression was significantly increased in LSCC tissues compared with normal tissues adjacent to cancer(P < 0.001);SALL4 mRNA expression was higher in cancer tissues of LSCC patients with clinical stage III/IV compared with LSCC patients with clinical stage I/II(P < 0.001).2.SALL4 mRNA and protein expression levels were significantly elevated in all five laryngeal cancer cell lines compared with 16HBE(P < 0.05).SALL4 mRNA and protein expression were most significantly elevated in AMC-HN-8 and TU686 cells compared with TU177,M4 E and LSC-1 cells(P < 0.001),therefore we selected AMC-HN-8 and TU686 cells as the subjects for the follow-up study.3.Compared with the Control group and sh-NC group,the expression of SALL4 in laryngeal cancer cell lines in the experimental group was significantly lower after transfection with sh RNA-SALL4(P < 0.01),indicating that the SALL4 knockdown model was successfully constructed.4.The MTT results suggested that the cell proliferation activity of laryngeal cancer cell lines in the sh-SALL4 group was significantly lower than that in the Control and sh-NC groups(P < 0.01).5.The results of the cell scratch assay showed that the scratch healing width of both laryngeal cancer cell lines in the sh-SALL4 group was significantly shorter and the migration rate was significantly lower(P < 0.01).6.The results of Transwell assay suggested that the number of cell-penetrating cells was significantly lower in the sh-SALL4 group(P < 0.01).7.Western blot results showed increased expression of EMT-related protein E-cadherin and decreased expression of vimentin in AMC-HN-8 and TU686 cells after knockdown of SALL4,and the difference was statistically significant(P < 0.05).Conclusions 1.The expression levels of SALL4 gene and its protein were elevated in LSCC tissues.2.The expression of SALL4 gene was higher in clinical stage III/IV than in clinical stage I/II.3.Knockdown of SALL4 inhibited LSCC cell proliferation,migration,invasion and EMT.2.In vitro experiments have shown that knockdown of SALL4 inhibits proliferation,migration,invasion and EMT of LSCC cells,and treatment targeting SALL4 is expected to be a new strategy against malignant progression of LSCC.