Aim To investigate the effects of ampelopsin on induction of apoptosis in human hepatocellular carcinoma Bel-7402 cells.Methods Bel-7402 cells were treated with ampelopsin with different concentrations for 24,48 and 72 h.The cell proliferation was detected by MTT assay.The morphological change of cells was observed through microscope observation by fluorescence staining.DNA fragmentation was visualized by agarose gel electrophoresis.The apoptosis rate was analyzed by flow cytometry.The expressions of caspase-3,Bcl-2 and Bax protein were detected by Western blot.Results Ampelopsin inhibited the proliferation of Bel-7402 cell line in a dose-and time-dependent manner.The IC50 values were 89.6?16.1,36.2?6.5 and 15.3?3.0 mg?L-1 at 24,48 and 72 h,respectively.The fluorescence microscope showed clearly cell apoptosis with apoptotic body.Agarose gel electrophoresis result showed that Bel-7402 treated with ampelopsin produced a DNA ladder band.The sub-G1 peak was detected and resulted in dose-and time-dependent increasing of the population of sub-G1 DNA content by flow cytometry.The expression of Bcl-2 protein was down-regulated,while the expression of Bax protein was up-regulated.The pro-caspase-3 protein was down-expressed and activated.Conclusions Ampelopsin could inhibit Bel-7402 proliferation through inducing cell apoptosis.The mechanism might be that ampelopsin could directly or indirectly enhance the level of anti-apoptosis protein Bcl-2 and decrease the level of apoptosis protein Bax.The pathway of pro-caspase-3 activated was initiated and effector caspase-3 was sequentially activated.

Objective To qualitatively analyze the expressions of key marker genes involved in osteogenic differentiation of bone marrow stromal cells (BMSCs) of mice at defined stages in vitro using real-time RT-PCR. Methods The BMSCs of mice were cultured in vitro and induced to osteoblasts for 7, 14, and 21 days. The total RNA was extracted from the cultured cells and the gene expression levels of collagenⅠ, bone alkaline phosphatase (APL), osteopontin (OPN) and bone sialoprotein (BSP) in the osteoblasts at the defined stages were determined by real-time RT-PCR. Results The ColⅠmRNA levels in 14 and 21 days were (0. 75?0. 04) and (0. 34?0. 03) times respectively of that in 7 days. ALP, OPN and BSP mRNA levels in 21 days were (19. 70?2. 36), (150. 12?9. 31) and (7. 73?0. 58) times of those in 7 days respectively. Conclusions ColⅠexpression levels tend to gradually decline while ALP, OPN and BSP levels gradually increase along with the culture time. Real-time RT-PCR is a reliable method for investigation of gene expressions in BMSCs cultured in vitro.

Objective To analyze the expression and regulation of HES1 in sperms with low motility.Methods Thirty semen samples from asthenospermia patients and 20 semen samples from healthy and fertile adults were collected,total RNAs were extracted to produce cDNAs probes.Hybridization with Phalanx OneArray~(TM) containing 30 968 probes was carried out after the labeled cDNAs were purified by PCR product purification kit.Realtime RT-PCR was used to analyze the expression of hsa-miR-487a and hsa-miR-193b;the expression of the target genes of hsa-miR-487a and hsa-miR-193b were searched from gene-expression profiles in asthenospermia patients' sperms.Results The expression level of HES1 in low motility sperms was up-regulated.The expression level of hsamiR-193b in low motility sperms was 2.19 times higher than that in high motility sperms,hsa-miR-487a was 0.43% of that in high motility sperms.Conclusion The expression level of HES1 in low motility sperms was up-regulated.Hsa-miR-487a and hsa-miR-193b may affect the expression of HES1 and so regulate sperm motility.

OBJECTIVETo study the anti-halitosis effect of sugar-free chewing gum through their influence on odor induced by cysteine.

METHODSTen volunteers were randomly divided into the treatment group and the untreated group; each group consisted of five volunteers. All volunteers consented to participate in a test in which breath odor was induced by cysteine. After the test, the treatment group chewed sugar-free chewing gum for 1 min, whereas the untreated group did not undergo any treatment. The effectiveness was determined by the percent reduction of H2S, CH3SH, and (CH3)2S response after the volunteers chewed gum for 1, 10, and 20 min.

RESULTSAt 1, 10, and 20 min, H2S of the treatment group was reduced by 82.68%, 92.27%, 97.47%, respectively, CH3SH was reduced by 65.49%, 73.79%, and 82.89%, respectively, and (CH3)2S was reduced by 60.45%, 73.82%, and 59.72%, respectively. The differences between the two groups at different times were significant (P < 0.05).

CONCLUSIONChewing gum can effectively inhibit cysteine-induced odor.

[Objective]To assess the clinical effect and radiological results of three methods of anterior cervical decompression and reconstruction for the treatment of multilevel cervical myelopathy.[Method]Fifty-two patients with continuous multilevel cervical myelopathy treated by the same team were divided into three groups.Group Ⅰ was performed the twosegment-corpectomy and reconstructed with a long titanium mesh and plate.Group Ⅱ was performed the three-level-discectomy and fused with three cages.Group Ⅲ was performed the combination with corpectomy and discectomy.The parameters included intervertebral height,JOA score.The data was compared statistically with Student s T-test or ANOVA.[Result]JOA score in all groups was improved post operation,but there was no significant difference among the three groups.The post-operation intervertebral height lose of group Ⅰ was more significant than other two groups,three cases of titanium meshs subsidence happened in group Ⅰ and one same case happened in group Ⅲ.[Conclusion]All the three surgical methods of anterior decompression and fusion can procure wonderful clinical effect,but group Ⅱ and Ⅲ were better than group Ⅰ in the stability and the maintenance of intervertebral height post operation.

External applies and washing of traditional Chinese medicine has sound curative effect in treating muscular and bone pain.The author reported some his experiences on the treatment of muscular and bone pain with hot applies and tendon-relaxing and collaterals-activating lotion.

Objective To explore the safety of the human bone marrow‐derived mesenchymal stem cells (hBMSCs) after silencing of human leukocyte antigen A2 expression .Methods We divided the cells into three groups:normal cultured cells of the 8th passage served as control group , and hBMSCs after HLA A2 silencing expression of the 5th and 15th passage as experimental groups 1 and 2 ,respectively .The hBMSCs were recultured by sterile methods .The growth curve ,telomerase activation ,and expressions of P27 ,cyclin D2 and cyclin‐dependent kinase 4 (CDK4) were utilized to explore the safety of the hBMSCs induced by LV‐siRNA‐HLA A2 .The BMSCs were transplanted to the subcutaneous layer of nude mice .Tissue types were detected 24 weeks after transplantation . Results The cell curves had no obvious left or right shift in all the groups . The telomerease activation in experimental groups 1 and 2 did not significantly differ from those in control group . The expressions of anti‐oncogene P27 ,cyclin D2 and CDK4 had no obvious difference between the two experimental groups and control group , either . There was only ectopic osteogenesis 24 weeks after the BMSCs (HLA A2 gene silenced ) were transplanted to the subcutaneous layer of the nude mice .Conclusion There was no obvious evidence to support that hBMSCs had undergone change in safety after the silencing of HLA A 2 expression .

BACKGROUND:Human leukocyte antigen (HLA), the major histocompatibility complex of human, plays an important function in the transplant rejection. Decreasing the expression of HLA wil prolong the survival time of transplants.
OBJECTIVE:To design smal interfering RNA (siRNA) of HLA-A2 and to detect the effect of siRNA-HLA-A2 on the expression of HLA-A2.
METHODS:Four kinds of siRNA-HLA-A2 domains were designed, and recombinant lentivirus expression vector were formed. The 293T cells, highly expressing HLA-A2, were infected in vitro. Then the knockout efficacy of four domains was detected to select the highly efficient siRNA-HLA-A2 target sequences. The human embryo lung fibroblasts were cultured in vitro and infected with the lentivirus carrying the target sequence. The infecting efficiency of LV-siRNA-HLA-A2 was observed under the fluorescence microscope and the silence function of this siRNA in human embryo lung fibroblasts was detected by western blot analysis.
RESULTS AND CONCLUSION:According to the mRNA sequence of HLA-A2 in Genbank, three siRNAs were designed and synthesized. In vitro, the over expression of HLA-A2 in 293K cells was successful y silenced. The HLA-A2 expression in human embryo lung fibroblasts was also efficiently silenced after the human embryo lung fibroblasts were infected by the highly efficient siRNA of HLA-A2. The efficacy was up to 80%.

Objective To elucidate the etiology feature of viral infection in hospitalized children with acute lower respiratory infection. Methods A total of 5 480 children with acute lower respiratory tract infection, hospitalized from September 2007 to September 2009, were studied. Nasopharyngeal aspirates were screened for 8 types of viruses by direct immunofluorescence (DIF) assay. Results At least one type of viral pathogen was detected in 2 710 out of 5 480 patients and the overall positive rate was 49.5%. The most common virus was RSV (51.1%), followed by hMPV (18.9%), PIVⅢ (12.5%), ADV (7.1%), IFA (4.7%), IFB (2.9%), PIV Ⅰ (1.5%) and PIV Ⅱ (1.2%). The positive rate was highest in children under 6 months (43.5%). The seasonal change of RSV, hMPV was more obvious. The peak of RSV, hMPV appeared in the winter and the spring. The prevalence of viral infection in children with pneumonia, bronchitis, asthmatic bronchitis, non asthmatic bronchitis and asthma were 47.4%、63.6%、 50.5%、 30.1% and 43.5% respectively. Conclusions Viruses are the main cause of lower respiratory tract infections in children, especially in infants and young children. RSV and hMPV were the most common viruses in these years.

10.3969/j.issn.2095-4344.2013.25.010