Expression of SH2 domain containing 2A in pancreatic ductal adenocarcinoma and its effect on gemcitabine resistance
Objective To explore the effect of SH2 domain containing 2A(SH2D2A)on gemcit-abine resistance in pancreatic ductal adenocarcinoma(PDAC).Methods Exploring SH2D2A expression in PDAC using the Kaplan-Meier Plotter and GEPIA platforms,and the impact on overall survival and dis-ease-free survival of PDAC patients;Forty cases of PDAC tissues and paracancerous tissues surgically resec-ted at Zhengzhou University People's Hospital from June 2022 to June 2023 were selected,and Western blotting was performed to analyze SH2D2A protein expression levels in the tissues;quantitative real-time polymerase chain reaction(qPCR)and Western blotting were performed to analyze the expression levels of SH2D2A protein and mRNA in the humol/Lan normal pancreatic ductal epithelial cells HPDE and humol/Lan PDAC cell line SW1990,MIA PaCa-2,PANC-1,HPAFⅡ,and BxPC-3;short hair RNA construction of stable knockdown SH2D2A cell lines,divided into Sh and NC groups;qPCR and Western blotting ana-lyzed the expression level of SH2D2A in the two groups of cells;cell counting kit-8(CCK-8)analyzed the half maximal inhibitory concentration(IC50)of the two groups of cells;colony formation assay,CCK-8,flow cytometry and tumol/Lor spheroid formation assay analyzed the proliferation,apoptosis and tumol/Lor stem-ness of the GEM-treated cells;t-test was used for comparison between groups.Results Bio-confidence a-nalysis showed that OS and DFS were significantly shorter in patients with high SH2D2A expression(both P＜0.01).SH2D2A protein expression in pancreatic cancer tissues was higher than that in paracancerous tissues,[(0.88±0.27)vs.(0.36±0.07),t=6.416,P＜0.05].The expression levels of SH2D2A were higher than those of HPDE in SW1990,MIAPaCa-2,PANC-1,HPAF Ⅱ and BxPC-3[mRNA:(5.86±0.45,3.47±0.27,4.74±0.37,2.38±0.18,2.54±0.20)vs.(1.00±0.08),t=15.780,12.280,14.490,9.040,9.632,all P＜0.05);Protein:(2.24±0.11,1.67±0.07,1.81±0.18,1.31±0.08,1.53±0.17)vs.(1.00±0.01),t=18.110,16.400,7.320,6.406,5.220,all P＜0.05].The expression level of SH2D2A in Sh group was lower than NC group[mRNA:(0.15±0.03,0.14±0.03)vs.(1.01±0.07),t=17.970,18.570,both P＜0.05);Protein:(0.35±0.02,0.30± 0.02)vs.(1.00±0.08),t=11.690,14.300,both P＜0.05].The IC50 of Sh group was lower than that of NC group,[(18.27±0.04,16.54±0.10)vs.(25.81±0.32),t=45.280,54.850,both P＜0.05];CCK-8 showed that the absorbance of cells in Sh group on day 4 was lower than NC group[(1.20± 0.06,1.02±0.08)vs.(2.22±0.08),t=17.440,18.490,both P＜0.05];the number of cell colonies in Sh group was lower than in NC group[(367.70±31.50,331.00±58.66)vs.(738.00±101.50),t=7.211,9.676,both P＜0.05];the proportion of apoptosis in Sh group was higher than NC group,[(28.59±3.44)％,(22.68±1.78)％vs.(17.67±0.38)％,t=5.166,4.375,both P＜0.05];the size of cell formation in Sh group was significantly smaller than NC group[(8.79±1.43,7.63±1.23)vs.(45.31±11.85),t=4.772,5.191,both P＜0.05].Conclusion SH2D2A expression is upregulat-ed in human PDAC and is involved in the regulation of PDAC resistance to gemcitabine.
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