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Objective To construct pcDNA3.1 RASSF1 eukaryotic vector and observe the influence of RASSF1 on the apoptosis of hepatocarcinoma cell line HepG2. Methods RASSF1 gene was amplifled from human RASSF1 cDNA by polymerase chain reaction (PCR) and cloned into pcDNA3.1. The recombinant plasmid pcDNA3. 1 RASSF1 was transfected into hepatocarcinoma HepG2 cell line. The expression of RASSF1 was examined by Western blot. The influence of RASSF1 on the cell apoptosis was measured by Annexin V/PI assay. Results DNA enzyme digestion and sequencing results showed that recombinant plasmid pcDNA3. 1-RASSF1 was successfully constructed. RASSF1 protein was overexpressed in HepG2 cell line transfected with pcDNA3. 1-RASSF1 plasmid. The apoptosis rate of blank, pcDNA3. 1 and pcDNA3. 1-RASSF1 group was (5.8 ±0.42)%, (7.48 ±0.68)% and (35. 1 ±3. 15)%, respectively.Conclusion The pcDNA3. 1- RASSF1 eukaryotic vector was successfully constructed, RASSF1 protein overexpression could induce apoptosis in HepG2 cell line.
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Objective To investigate the change of gene rstn expression in brain tissues following traumatic brain injury (TBI). Methods A total 90 SD rats were involved in the study and divided into normal control group (5 rats), sham operation group (10 rats), mild, moderate and severe trauma groups (25 rats per group). Rat model was made with sap pressure clash method and RT-PCR assay was employed to detect expression change of gene rstn at 3, 6, 24, 72 h and 1, 2, 4 weeks after TBI. The change of peripheral blood glucose concentration was measured in moderate trauma group to observe its relationship with gene rstn expression in brain tissue. Results Postoperative expression of gene rstn was increased in severe trauma group at 24 hours, in moderate trauma group at 72 hours and in mild trauma group at four weeks (P＜0.05). The expression of gene rstn was increased in hippocampus, thalamus and cortex of all trauma groups at four weeks postoperatively, when the hippocampus showed the highest expression while the thalamus showed the least (P＜ 0.05). Moreover, the expression of gene rstn showed high level in injury side but low level in contralateral side in different districts (P ＜ 0.05). The expression of gene rstn was increased the most obviously in severe trauma group (P ＜0.05). Peripheral blood glucose concentration showed a linearity positive correlation with gene rstn expression in brain tissue (R=5.32,P＜0.05). Conclusions Expression of gene rstn shows obvious increase after TBI, and the time course correlates with the injury severity. The gene rstn expresses the most in the ipsilateral hippocampus. There shows a certain correlation between gene rstn expression and peripheral blood glucose concentration in brain tissues after TBI.