Expression of SAPCD2 in esophageal squamous cell carcinoma tissues and its effects on the biological function of esophageal squamous cell carcinoma cells in vitro and the possible mechanisms
10.3760/cma.j.cn115355-20240809-00385
- VernacularTitle:SAPCD2在食管鳞状细胞癌组织中的表达及其体外对食管鳞状细胞癌细胞生物学功能的影响和可能机制的研究
- Author:
Qixiong LONG
1
;
Jingchun LI
;
Junxia FENG
;
Wenxi HE
;
Wei SUN
Author Information
1. 广州市花都区人民医院消化内科，广州 510800
- Publication Type:Journal Article
- Keywords:
Esophageal neoplasms;
Carcinoma, squamous cell;
Cell proliferation;
Neoplasm invasiveness;
SAPCD2
- From:
Cancer Research and Clinic
2025;37(5):334-342
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the expression of SAPCD2 in esophageal squamous cell carcinoma (ESCC) tissues and its effects on the biological function of ESCC cells in vitro, as well as the possible molecular mechanisms.Methods:By using the Gene Expression Profile Interaction Analysis (GEPIA) platform, the transcriptional level expressions of SAPCD2 in 182 ESCC samples and 286 normal esophageal tissue samples from the Cancer Genome Atlas (TCGA) database and Genotype-Tissue Expression (GTEx) database were analyzed. Immunohistochemical (IHC) staining was performed on clinical tissue chips of ESCC patients, and staining scores were evaluated. The expression differences of SAPCD2 protein between 61 cancer tissues and the paired adjacent tissues with the complete clinical data, as well as the distribution of patients with SAPCD2 high expression among patients stratifies by different clinicopathological features were compared. ESCC cell line KYSE-150 was transfected with plasmids carrying SAPCD2 sequence and short hairpin RNA sequence targeting SAPCD2, respectively, which was treated as the SAPCD2 overexpression group and SAPCD2 knockdown group; and the cells transfected with empty plasmids and plasmids carrying negative RNA sequence were treated as the overexpression control group and the knockdown control group. CCK-8 method (expressed with the absorbance value) and plate clone formation assay were used to detect the cell proliferation ability. Cell migration was detected by using cell scratch assay and Transwell cell migration assay were used to detect the cell migration ability. The reverse-real time fluorescence quantitative polymerase chain reaction (qPCR) was used to detect the mRNA levels of SAPCD2, matrix metalloproteinase-9 (MMP9), proliferating cell nuclear antigen (PCNA), and Vimentin in cells of all groups. Western blotting was used to detect the expression levels of SAPCD2 protein and proteins related to cell proliferation, invasion, metastasis, and AKT signaling pathway.Results:GEPIA platform analysis showed that the transcriptional expression level of SAPCD2 in ESCC tissues was higher than that in adjacent tissues, and the difference was statistically significant ( P < 0.01). IHC staining showed that the staining score of SAPCD2 protein in cancer tissues was higher than that in adjacent tissues [(8.2±2.8) points vs. (2.2±1.7) points], and the proportion of patients with positive SAPCD2 protein (staining score > 0 point) in cancer tissues was higher than that in adjacent tissues [95.1% (58/61) vs. 57.4% (35/61)], and the differences were statistically significant (all P < 0.001), while there were no statistically significant differences in the distribution of the high expression of SAPCD2 protein (staining score > 3 points) patients stratified by gender, age, tumor size, pathological grade, and T stage, N stage and M stage (all P > 0.05). CCK-8 assay showed that the absorbance value of KYSE-150 cells in the SAPCD2 overexpression group after 96 h of culture was higher than that in the overexpression control group, while the absorbance values of the SAPCD2 knockdown group after 72 h and 96 h of culture were lower than those in the knockdown control group, and the differences were statistically significant (all P < 0.05). The plate clone formation assay showed that the number of colonies of KYSE-150 cells cultured for 14 d in the SAPCD2 overexpression group was more than that in the overexpression control group [(800±30) vs. (458±47)], and that in the SAPCD2 knockdown group was less than that in the knockdown control group [(52±7) vs. (81±2)], and the differences were statistically significant (all P < 0.05). The cell scratch assay showed that after 24 h of culture, the scratch width of KYSE-150 cells in the SAPCD2 overexpression group was narrower than that in the overexpression control group [(51±9) μm vs. (89±7) μm], while that in the SAPCD2 knockdown group was wider than that in the knockdown control group [(120±22) μm vs. (37±10) μm], and the differences were statistically significant (all P < 0.01). Transwell cell migration assay showed that the migration number of KYSE-150 cells in the SAPCD2 overexpression group was more than that in the overexpression control group [(202±18) vs. (50±14)], and that in the SAPCD2 knockdown group was less than that in the knockdown control group [(227±27) vs. (483±16)], and the differences were all statistically significant (all P < 0.01). qPCR assay showed that the mRNA relative expression levels of MMP9, PCNA and Vimentin in KYSE-150 cells in the SAPCD2 overexpression group were all higher than those in the overexpression control group, while those in the SAPCD2 knockdown group were all lower than those in the knockdown control group, and the differences were statistically significant (all P < 0.05). Western blotting showed that the relative expression levels of PCNA and Vimentin proteins in KYSE-150 cells of the SAPCD2 overexpression group were higher than those of the overexpression control group, while the relative expression levels of epithelial cadherin (E-cad) and cleaved cysteine aspartate protease 3 (CASP3) proteins were lower than those of the overexpression control group; however, the expression levels in SAPCD2 knockdown group showed the opposite results, and the differences were statistically significant (all P < 0.05); the relative expression level of phosphorylated AKT (p-AKT) protein in the SAPCD2 overexpression group was higher than that in the overexpression control group, while that in the SAPCD2 knockdown group was lower than that in the knockdown control group, and the differences were statistically significant (all P < 0.01). Conclusions:SAPCD2 is highly expressed at both the transcriptional level and the protein level in ESCC tissues. SAPCD2 promotes the proliferation and migration of ESCC cells in vitro, which may be related to the AKT signaling pathway.
