Combined effects of hydrostatic pressure and estrogen on osteogenic and chondrogenic differentiation of bone marrow mesenchymal stem cells / 医用生物力学

Objective To investigate the effect of hydrostatic pressure and estrogen on the proliferation, F-actin cytoskeleton, osteogenic and chondrogenic differentiation of rat bone marrow mesenchymal stem cells (BMSCs), and to test whether combined stimulation can exert the fortified stimulating effort on BMSCs． Methods BMSCs were separated by using the whole bone marrow culture method and purified by differential adherence method. BMSCs surface markers were detected by flow cytometer. BMSCs were randomly assigned to six groups：blank control group (Group C), 1 nmol/L 17β-Estradiol treatment group (Group E), 1 nmol/L tamoxifen treatment group (Group T), 90 kPa pressure treatment group for 1 h (Group P); 17β-Estradiol pretreatment for 12 h and 90 kPa pressure group for 1 h (Group P+E); and tamoxifen pretreatmet for 12 h and 90 kPa pressure group for 1 h (Group P+T). Cell cycle was measured by flow cytometry. Fluorescent staining under laser scanning confocal microscope observation was observed for F-actin cytoskeleton expression and re-assembly. After osteogenic differentiation for 7 d and 14 d, calcified nodules were detected with alizarin red staining. Further, the osteogenic markers including Col I, ON, OPN and BSP were analyzed by real-time PCR. Following chondrogenesis of BMSCs for 14 d and 28 d, proteoglycan contents were detected with toluidine blue staining, and chondrogenic markers including Sox9, Aggrecan and ColⅡwere evaluated by real time PCR. ANOVAs followed by the Dunnett t tests were adopted for comparisons among subgroups. All the experimental data were analyzed by SPSS 16.0 software. Results Both hydrostatic pressure (90 kPa, 1 h) and 1 nmol/L17β-estradiol could increase the proliferation of BMSCs and F-actin activation, but no bio-cooperation effects appeared. Calcified nodules were observed after 14 d osteogenic induction. Real-time PCR showed the estrogen enhanced osteogenetic gene (Col I, ON, OPN and BSP) expression in 7 d and 14 d. Combined effects of pressure and estrogen showed synergistic improving effects on early osteogenetic differentiation, but oppositional effects on advanced osteogenetic differentiation. Toluidine blue staining was positive after 28 d chondrogenic induction. With the hydrostatic pressure loading regime, the mRNA expression of chondrogenic genes (Sox9, Aggrecan and ColⅡ) was increased significantly, but with oppositional effects from estrogen on advanced chondrogenic differentiation. Conclusions The superposition effects of mechanical stimulation and estrogen acting only enhanced the differentiation of BMSCs in the early osteogenetic differentiation, but no effect was found in the proliferation and F-actin activation. Hydrostatic pressure and estrogen show antagonistic action in advanced osteogenetic differentiation and chondrogenic differentiation. Estrogen promotes osteogenetic differentiation, while hydrostatic pressure can enhance chondrogenic differentiation of BMSCs.

The mechanism of HBV infection of human trophoblast cell / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To observe the changes of human trophoblast cells after infected with hepatitis B virus.</p><p><b>METHODS</b>HBV positive serum was used to infect human trophoblast cells in vitro. HBsAg in cell culture medium were detected by ELISA method and HBV DNA in cell culture medium and cells were detected by PCR method. HBV fluorescence polymerase chain reaction diagnose kit were used to detect the HBV DNA concentration. Ultra structure of trophoblast cells were observed with transmission electron microscopy (TEM).</p><p><b>RESULTS</b>HBsAg could be detected in infection group by ELISA. Infection group cell culture medium and infection group cells were HBV DNA positive. HBV DNA concentrations in HBV infection cell culture medium in 0, 12, 36, 60, 84 h after extensively PBS washed were < 10(3), 3 x 10(4), 6 x 10(5), 5 x 10(5), 3 x 10(5) copies/mL. HBV infected trophoblast cells were found many forms of endosomes, some of which contents virus like particle.</p><p><b>CONCLUSION</b>HBV might take advantage of clathrin-mediated endocytosis to enter trophoblast cell, which might lead to cell infection or across the cell bar by transcytosis.</p>

In vitro infection of human hepatoma (Hep G2) cell line by hepatitis B virus positive serum / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To establish a culture system of HBV positive serum infected Hep G2 cells in vitro.</p><p><b>METHODS</b>Hep G2 cells were seeded into six-well cluster dishes, at 1 x 10(-6) cells per well and incubated with 3 ml 10% fetal calf serum/ Dulbecco's modified Eagle's medium (10% FCS/DMEM) at 37 degrees in 5% CO2 air. At 24 h after plating, infection group Hep G2 cells were cultured with 0.5 ml HBV positive serum, in control group HBV negative serum was used, 24 h later the inoculums was removed. The cells were then extensively washed with 0.01 mol/L phosphate-buffered saline (PBS). After washing with PBS, 4 ml 2% FCS/DMEM were added to each well and the medium was collected every 12 h. ELISA method was used to detect HBsAg in culture medium. HBV DNA in cells and culture medium was detected by PCR.</p><p><b>RESULTS</b>In infection group, HBsAg could be detected from cell culture medium from 12 h (after PBS washed) to 84 h. HBV DNA could be detected by PCR in culture medium and cells.</p><p><b>CONCLUSION</b>Infection of Hep G2 cells by HBV positive serum is feasible.</p>

Meta-analysis on the relationship between tobacco smoking, alcohol drinking and p53 alteration in cases with esophageal carcinoma / 中华流行病学杂志

<p><b>OBJECTIVE</b>To investigate the relationship between tobacco smoking, drinking and p53 alteration in esophageal carcinoma.</p><p><b>METHODS</b>Literature on the relationship between p53 alteration in esophageal carcinoma and tobacco smoking, drinking through Meta-analysis were reviewed.</p><p><b>RESULTS</b>In 14 selected papers related to tobacco smoking, pooled odds ratio (OR) of tobacco smoking with P53 overexpression and p53 alteration were 1.99 (95% CI: 1.30- 3.06) and 1.64 (95% CI: 1.13 - 2.37), respectively (P < 0.05). Pooled OR of tobacco smoking with p53 mutation was 1.11 (95% CI: 0.47 - 2.76) (P > 0.05). In 11 selected papers on alcohol drinking, pooled OR of drinking with P53 overexpression, p53 mutation and p53 alteration were 1.30 (95% CI: 0.83 - 2.04), 1.13 (95% CI: 0.67 - 1.90) and 1.22 (95% CI: 0.87 - 1.72) respectively (P > 0.05).</p><p><b>CONCLUSION</b>There were significant relations between tobacco smoking and p53 alteration while there were no significant relations between alcohol drinking and p53 alteration.</p>