Objective To explore the expression of amino acid transporters solute carrier family-1 member-5 (SLC1A5) in esophageal squamous cell carcinoma (ESCC) and its correlation with clinicopathological features, and investigate the potential molecular mechanisms of SLC1A5 in ESCC metastasis.
Methods The expression of SLC1A5 gene in ESCC and normal tissues was analyzed by TNM plot online database. The SLC1A5 mRNA expression in ESCC and paracancer tissues was examined by quantitative real time polymerase chain reaction (qRT-PCR); the protein expression of SLC1A5 was detected by immunohistochemistry (IHC) and the correlations between SLC1A5 and clinic pathological characteristics in patients were analyzed; SLC1A5 overexpression (SLC1A5-OE group), transmembrance-4 L-six family member-1 overexpression (TM4SF1- OE group), SLC1A5 and TM4SF1 co-overexpression Eca109 cells were constructed respectively. Cell proliferation test and Transwell test were used to detect the proliferation, migration and invasion ability of Eca109 cells. The interaction between SLC1A5 and TM4SF1 was confirmed by immunoprecipitation (IP) assays; the expression level of mammalian target of rapamycin (mTOR) signaling pathway-related protein was detected by Western blot (WB).
Results TNM plot database analysis showed that the expression level of SLC1A5 gene in the ESCC tissues was significantly higher than that in the normal tissues (P < 0.05). The expression of SLC1A5 mRNA in ESCC tissues was significantly higher than that in paired para-cancerous tissues (P < 0.01). IHC results showed that SLC1A5 protein was highly expressed in cancer tissues, and was significantly correlated with clinical stage (P=0.036) and lymph node metastasis (P=0.029). The results of cell proliferation experiment showed that there was no significant difference in proliferation ability between the SLC1A5-OE group and control group (Con group) (P>0.05). The results of Transwell experiment showed that compared with the Con group, the cell migration and invasion ability of the SLC1A5-OE group were significantly enhanced (P < 0.01). The IP results showed that SLC1A5 interacted with TM4SF1. WB results showed that the expression levels of p-mTOR and p-S6 proteins were the highest in the cells co-overexpressing SLC1A5 and TM4SF1, relative to the SLC1A5-OE group or TM4SF1-OE group. Transwell experiment confirmed that ESCC cells overexpressing SLC1A5 and TM4SF1 had the strongest migration ability.
Conclusion SLC1A5 is highly expressed in ESCC, and is significantly correlated with clinical stage and lymph node metastasis of patients. SLC1A5 may activate mTOR signaling pathway through interaction with TM4SF1, thus promoting ESCC cell migration.
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