Study On The Influence Of SALL2 Expression On The Growth Of Cervical Cancer Cell

Objective: This study was undertaken to define the expression and function of Spalt-like Gene-2 (SALL2) in cervical cancer cells and cervical cancer stem cells.Methods:Firstly, the expression and function of SALL2 in cervical cancer cells were studied. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the expression of SALL2 in cervical cancer cell lines C33A, ME-180, HeLa, SiHa, MS751 and CaSKi. The expression of SALL2 in ME-180 and HeLa cells was detected by immunoblotting (Western blot) and cell immunofluorescence. Three small interfering RNAs (siRNA) against SALL2 genes (siRNA-1,2,3)were constructed. Liposomes were used to transfect siRNA into human cervical cancer cell lines ME-180 and HeLa, and the carrier group and the control group were established. After transfected the siRNAs, RT-PCR and Western blot were used to detect the expression of SALL2. CCK-8 and flow cytometry were used to determine the growth and apoptosis of ME-180 and HeLa with silencing of SALL2. Scratch assay was used to detect the migration of cervical cancer cell. Reverse transcription quantative polymerase chain reaction (RT-qPCR) were used to detect the expression of CDKN1A mRNA in the transfected cells.The expression and function of SALL2 in cervical cancer stem like cells were discussed.ME-180, C33A and HeLa were suspension cultured in vitro with serum-free medium to enrich the stem-like cells. RT-qPCR was used to detect the expression of stem cell markers OCT4, SOX2 and EMT marker TWIST1 in cervical stem-like cells. RT-qPCR was used to detect the expression of SALL2 in cervical stem-like cells. Colony-formation assay was used to test the ability of stem-like cells to form clones.Results:SALL2 is expressed in C33A, ME-180, HeLa and MS751 cells. The expression level of SALL2 in HeLa was higher than other cell lines. The expression of SALL2 in HeLa and ME-180 has no difference between the control and carrier groups (P>0.05). Compared with the control group, the expression of SALL2 in HeLa and ME-180 was down-regulated 48h after transfection with the siRNA-1 and 2 (P<0.05), but there is no difference in protein expression of siRNA-3 group.siRNA-1 exerted the highest efficiency of SALL2 silencing in the 3 siRNAs. Compared to the control group, the proliferation of HeLa cell in the siRNA-1 group was increased 48, 72h after transfection (P<0.05). Compared to the control group, the proliferation of ME-180 cell in the siRNA-1 group was increased 24, 48, 72h after transfection (P<0.05). In the siRNA-1 group,the ratio of HeLa and ME-180 cells in G0/G1 phase was decreased,and apoptosis was reduced 48h after transfection (P<0.05). Compared with the control group, CDKN1A mRNA expression level in ME-180 cells was significantly decreased in ME-180 cells after transfection (P<0.001).The expressions of stem cell markers OCT4, SOX2 and epithelial-mesenchymal transition markers TWIST1 were elevated in cervical cancer stem-like cells. Compared to the normal C33A and HeLa cells,the expression of OCT4 in the stem-like cells was significantly increased (P<0.001).The expression of SALL2 in cervical cancer stem-like cells was significantly higher than normal cervical cancer cells (P<0.001). The colony-forming ability of cervical cancer stem-like cells was significantly higher than normal cancer cells (P<0.001).Conclusion:1. SALL2 is expressed in cervical cancer cell lines ME-180, HeLa, C33A and MS751.2. SALL2 silencing promotes cell proliferation and inhibits apoptosis of HeLa and ME-180.The expression of SALL2 negatively regulates the growth of cervical cancer cells HeLa and ME-180.3. SALL2 and CDKN1A synergistically inhibit the growth of cervical cancer cells.4. The cervical cancer stem-like cells can be enriched by suspension culture method. The expression of SALL2 was increased in cervical cancer stem-like cells.