RESUMO
Objective:To investigate the effects of miRNA-26a (miR-26a) on the target gene HMGA2 on the proliferation and migration of hepatoma cells and the underlying mechanism. Methods:Liver cancer tissue samples ( n = 30) and adjacent normal tissue samples ( n = 30) pathologically confirmed by Wenzhou Hospital of Traditional Chinese Medicine between September 2018 and September 2019 were collected. MiR-26a mimics, control mimics (miR-Control), high-mobility group A2 protein (HMGA2) siRNA or negative control siRNA (Control) were transfected into human hepatoma cell lines HepG2 or Huh-7 cells. The expression of miR-26a in hepatocellular carcinoma tissue was detected by reverse transcription quantitative polymerase chain reaction (RT-qPCR). MTT assay and scratch test were performed to determine the ability of cell proliferation and migration. RT-qPCR and western blotting were performed to detect miR-26a and HMGA2 mRNA expression. The relationship between miR-26a and HMGA2 mRNA was analyzed using Bioinformatics and luciferase reporter gene assay. Results:RT-qPCR results showed that the expression level of miR-26a in hepatocellular carcinoma tissue was 0.11 ± 0.02, which was significantly lower than that in normal tissues (0.25 ± 0.03, t = 21.268, P < 0.05). The expression level of miR-26a in stage III + IV was 0.05 ± 0.01, which was significantly lower than that in stage I + II (0.09 ± 0.01, t = 15.491, P < 0.05). Cell experiment showed that in the miR-26a group, the proliferation ability of Huh-7 cells was (3.10 ± 0.30) and (4.10 ± 0.40), and the proliferation ability of HepG2 cells was (3.08 ± 0.31) and (4.11 ± 0.40), which was significantly lower than that in the control group [(3.90 ± 0.40), (5.50 ± 0.60), (3.92 ± 0.41), (5.49 ± 0.58), t = 8.764, 10.634, 11.148, 10.728, all P < 0.05]. In the miR-26a group, the migration ability was (0.50 ± 0.06), (0.65 ± 0.07), which was significantly lower than that in the control group [(1.00 ± 0.10), (0.96 ± 0.10), t = 23.483, 13.910, both P < 0.05]. Bioinformatics and in vitro experiments showed that HMGA2 was a direct target of miR-26a. Restoring the expression of HMGA2 in miR-26a mimics-transfected cells, compared with that in the miR-26a group [(0.24 ± 0.02), (0.31 ± 0.03);(0.45 ± 0.05)], could significantly reverse the inhibitory effect of miR-26a on tumor cell proliferation and migration [(0.31 ± 0.03), (0.40 ± 0.04);(0.93 ± 0.08), t = 10.634, 9.859, 27.868, all P < 0.05). Conclusion:miR-26a inhibits the proliferation and migration of hepatoma cells by directly targeting HMGA2. The abnormal decrease of miR-26a and the increase of HMGA2 may be the important factors that participate in the occurrence and development of liver cancer.
RESUMO
Background:Acute-on-chronic liver failure( ACLF)is a commonly seen liver failure in China,and lacking an animal model that can effectively simulate the dynamic change of immune status of ACLF. Aims:To establish an animal model that can simulate dynamic change of immune status of ACLF by repeated administration of concanavalin A(ConA). Methods:Mice were randomly divided into normal control group and ConA repeated administration group. Mice in ConA repeated administration group were injected with ConA 15 mg/ kg through retrobulbar angular vein every 48 hours for 5 times,and mice in control group were injected with same volume of 0. 9% NaCl solution. Serum levels of IL-6,IL-10,IL- 12,TNF-α,IFN-γ,MCP-1 in peripheral blood were assessed by CBA assay,and the ratio of IL-10/ TNF-α was calculated. The expression of HLA-DR,number and proportion of CD4+ T cells and the expression of PD-1 of monocytes in peripheral blood were detected by flow cytometry. Results:Peripheral blood cytokines changed from predominated proinflammatory cytokines into predominated anti-inflammatory cytokines with the increasing in time of administration in ConA repeated administration group. Compared with control group,HLA-DR expression of monocytes in peripheral blood was significantly decreased(P <0. 05),number and proportion of CD4+ T cells were significantly decreased(P <0. 05), and PD-1 expression was significantly increased( P < 0. 05)in ConA repeated administration group. Conclusions:An animal model of ACLF immune status from systemic inflammatory response syndrome( SIRS) to compensatory antiinflammatory response syndrome(CARS)induced by repeated ConA stimulation is successfully established.