In order to study the effects and the possible mechanisms of Daxx overexpressed in HepG2 to hydrogen peroxide treatment, and to search new targets for cancer chemotherapy, HepG2cells were transfected using lipofectamine 2000, and selected by treatment with G418. Stable cell lines were confirmed by reverse transeriptase polymerase chain reaction (RT-PCR) targeting vector gene. Experiments include the following groups: (1) control group (non-transfected cells); (2) transfected with empty vector (HepG2/GFP cells); and (3) transfected with pEGFP-C1-Daxx (HepG2/GFP-Daxx cells). After incubation with hydrogen peroxide (H2O2) for 24 h, cellular viability was analyzed by MTT, and cellular apoptosis was measured by flow cytometric analysis. Gene expression at protein level was detected by Western blot. The RT-PCR results showed that Daxx RNA in cells transfected with pEGFP-C1-Daxx was increased significantly compared with that in the HepG2/GFP cells. Fluorescence microscopy revealed that Daxx protein was localized in the nuclei. Hydrogen peroxide was used to induce apoptosis of HepG2 cells and observed that the hydrogen peroxide decreased the viability of HepG2 cells in concentration-dependent pattern. The IC50 values in three groups (Normal cells, HepG2/GFP cells and HepG2/GFP-Daxx cells) were 0.72, 0.76, and 0.49 mmol/L respectively. The apoptotic ratio was significantly higher in HepG2/GFP-Daxx cells as compared to the other two groups. HepG2/GFP-Daxx cell incubated with hydrogen peroxide, showed a significant increase in the activation of caspase-3 and JNK as compare with the other groups. Over-expression of Daxx facilitated HepG2 cells apoptosis induced by hydrogen peroxide. Furthermore, there may be a synergetic relation with apoptosis and increase of JNK activity.

Objective To investigate subtle structural changes of white matter in patients with amnestic mild cognitive impair-ment (aMCI)using a tractography-based method.Methods Thirty patients with clinical diagnosis of aMCI and thirty-one cases of normal control(NC)and undergone diffusion tensor imaging.Cingulum (CG),corpus callosum (CC),uncinate fasciculus (UNC) and inferior occipitofrontal fasciculus (IOFF)were reconstructed,and fractional anisotrophy (FA)values were measured along these tracts using dTV II software.Differences of white matter tracts’FA values were explored between aMCI group and NC group.In addition,correlation analyses were also done between FA values and the MMSE(mini-mental state examination)scores in the aMCI patients.Results ① aMCI patients exhibited significant lower FA values in the bilateral CG,bilateral UNC and CC than NC group. Although there were no statistically significant differences,aMCI patients exhibited lower FA values than NC group in the left IOFF.② The FA values of right CG were positively correlated with MMSE scores.Conclusion Abnormal changes of FA values in CG,UNC and CC of aMCI patients suggest that subtle damages of white matter tracts related to memory exist in the prodromal phase of Alzheimer’s disease (AD).Fiber tractography has high sensitivity in detecting early damages of white matter.

Objective To observe the curative effect of Bushen-Tongdu decoction combined with percutaneous minimally invasive decompression for degenerative lumbar spinal stenosis. Methods A total of 80 patients with degenerative lumbar spinal stenosis were selected and divided into two groups of 40 cases by random number table. The control group was treated with percutaneous transforaminal minimally invasive decompression. The treatment group was given Bushen-Tongdu decoction on the basis of the control group. Both groups were treated for 14 days. The Japanese orthopedic Society back pain evaluation criteria for waist pain, leg pain and numbness, walking ability, sensory dysfunction, dyskinesia, straight leg raising test score were collected and compared. Results The total effective rate was 95.00％ (38/40) in the treatment group and 82.50％ (33/40) in the control group. The total effective rate of the two groups was statistically significant (Z=-2.357,P=0.018). After treatment, the levels of waist pain score, leg pain and numbness score, walking ability score, sensory dysfunction score, dyskinesia score and the straight leg raising test score in the treatment group were significantly higher than those in the control group(t=-2.180, -2.059, -1.985, -2.177, -2.045, -2.238, P<0.05).Conclusions The Bushen-Tongdu decoction combined with percutaneous minimally invasive decompression can reduce pain, improve walking ability, improve sensory and motor dysfunction, and improve clinical efficacy of the patients with degenerative lumbar spinal stenosis.

OBJECTIVE: To construct a recombinant lentiviral expression vector pCDH-Daxx-EGFP to investigate the effect of Daxx on the proliferation of vascular smooth muscle cells (VSMCs). METHODS: The recombinant lentiviral expression vector pCDHDaxx-EGFP was constructed using PCR-based accurate synthesis method. After identification by sequencing and enzyme digestion, the recombinant lentiviral vector was contransfected into 293T cells with lentivirus packaging vector. The recombinant lentivirus particles were collected and purified to infect VSMCs, whose expression of Daxx was detected with Western boltting. The cells infected with the empty vector pCDH-EGFP or pCDH-Daxx-EGFP were incubated in serum-free medium or in the presence of angiotensin Ⅱ (AngⅡ). The cell viability was determined with MTT assay, and the cell cycle changes were analyzed with flow cytometry. The cell migration ability was assessed using a scratch wound healing assay. The expression of p-Akt protein in the cells was detected using Western blotting. RESULTS: Double enzyme digestion and sequencing confirmed successful construction of the recombinant plasmid. Compared with the cells infected with the empty vector, the cells infected with pCDH-Daxx-EGFP exhibited significantly increased expressions of Daxx protein ( < 0.05). AngⅡ treatment of the cells infected with the pCDH-Daxx-EGFP, as compared with the cells infected with the empty vector, significantly lowered the cell viability, S phase cell ratio and cell migration ability ( < 0.05), and significantly decreased the expression level of p-Akt protein ( < 0.05). CONCLUSIONS We successfully constructed the recombinant lentiviral vector pCDH-Daxx-EGFP and overexpressed Daxx in primary cultured VSMCs using this vector. Daxx overexpression can inhibit AngⅡ-induced proliferation and migration in VSMCs probably by regulating p-Akt protein.