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. 2020 Jan 3;51:102603. doi: 10.1016/j.ebiom.2019.102603

Fig. 2.

Fig 2

The effect of TNFAIP1 on HCC cell proliferation, apoptosis and tumor growth. a. RT-qPCR analysis of the mRNA level of TNFAIP1 in MHCC97H cells infected with TNFAIP1 or the control lentivirus (left) (***P<0.001, Student's t-test) and in SMMC7721 cells infected with shTNFAIP1 or shControl lentivirus (right) (*P < 0.05, **P < 0.01, one-way ANOVA). b. Western blot analysis of TNFAIP1 protein expression in MHCC97H infected with TNFAIP1 or the control lentivirus (upper) and in SMMC7721 cells infected with shTNFAIP1 or shControl lentivirus (lower). c. CCK8 assay was used to determine cell proliferation in MHCC97H cells infected with TNFAIP1 or the control lentivirus (left) (**p<0.01, Student's t-test) and in SMMC7721 cells infected with shTNFAIP1 or shControl lentivirus (right) (**p < 0.01, one-way ANOVA) at 24, 48, 72 and 96 h. d. Representative photographs of the tumors at 6 weeks after injection with MHCC97H-TNFAIP1 or Control stable cells (n = 5/group). e. Tumor weight and tumor volume of nude mice subcutaneous injected with MHCC97H-TNFAIP1 or MHCC97H—Control stable cells were measured (n = 5/group) (**P<0.01, Student's t-test). f. Representative photographs of the tumors at 6 weeks after injection with SMMC7721-shTNFAIP1 or shControl stable cells (n = 5/group). g. Tumor weight and tumor volume of nude mice subcutaneous injected with SMMC7721-shTNFAIP1 or SMMC7721-shControl stable cells were measured (n = 5/group) (*P<0.05, Student's t-test). h. Immunostaining of xenograft tumors from MHCC97H-TNFAIP1 and MHCC97H—Control groups (upper) or from SMMC7721-shTNFAIP1 and SMMC7721-shControl groups (lower) were conducted using a specific anti-Ki67 antibody, Scale bars, 25 μm. i. The Ki67-positive cells were quantitatively analyzed in MHCC97H-TNFAIP1 and MHCC97H—Control groups (left) (***P<0.001, Student's t-test) or in SMMC7721-shTNFAIP1 and SMMC7721-shControl groups (Right), (**P<0.01, Student's t-test) (n = 5). j. TUNEL assay to determine cell apoptosis in MHCC97H cells infected with TNFAIP1 or the control lentivirus (upper) and in SMMC7721 cells infected with shTNFAIP1 or shControl lentivirus (lower). Scale bar, 25 μm. k. Quantitative analysis of TUNEL staining in MHCC97H cells infected with TNFAIP1 or the control lentivirus (**P<0.01, Student's t-test) and in SMMC7721 cells infected with shTNFAIP1 or shControl lentivirus (**P<0.01, Student's t-test). l. RT-qPCR analysis of the mRNA level of TNFAIP1, RhoB, Bcl-XL and Bcl-2 in MHCC97H cells infected with TNFAIP1 or the control lentivirus and in SMMC7721 cells infected with shTNFAIP1 or shControl lentivirus (*P<0.05, **P<0.01 and ***P<0.001, Student's t-test). m. Western blot analysis of the expression of Cleaved-caspase3, Bcl2, Bcl-XL, Bax, RhoB and TNFAIP1 in MHCC97H infected with TNFAIP1 or the control lentivirus and in SMMC7721 infected with shTNFAIP1 or shControl lentivirus. Data are presented as means ± SEM from triplicate independent experiments. P-values were determined by two-tailed Student's t-test or one-way ANOVA (*P < 0.05, **P < 0.01, ***P < 0.001).