Aim To construct the eukaryotic expression vectors of short hairpin RNA targeting survivin and observe its effect on biologic behavior of A549 cells and sensitivity of A549 cells to paclitaxel.Methods The DNA fragment targeting human survivin was inserted into the plasmid,and the recombinant plasmid was constructed.The recombinant plasmids cells were transfected into A549 cells by FuGENE transfection reagent.The expression levels of survivin gene were detected with RT-PCR and Western blot before and after transfection,respectively.Cell apoptosis was detected by TUNEL method.The sensitivity of A549 cells to paclitaxel was detected by MTT after transfection.Results The recombinant plasmid was successfully constructed.RNAi group cells showed lower expression of survivin than control group.The apoptosis rate of A549 cells increased after transfection.The IC50 of paclitaxel inhibiting A549 cells was 11.9 fold before transfection compared with those after transfection.There was significant difference between the two groups(P

ObjectiveTo detect the mutations of GJB3 and GJB4 genes in two sporadic cases of erythrokeratodermia variabilis(EKV).MethodsGenomic DNA was extracted from two sporadic patients with EKV,their family members,and 100 normal human controls.All the exons and adjacent splice sites of GJB3 and GJB4 genes were amplified by PCR.Mutation scanning was carried out via direct bidirectional DNA sequencing.ResultsA G134C mutation was found at the GJB3 gene in patient 1,which caused a substitution of glycine by alanine at codon 45 (G45A).No mutation was found in the GJB4 gene in case 1 or GJB3 and GJB4 genes in case 2.ConclusionA missence mutation G45A in GJB3 gene is found in a patient with EKV.

Background Laser peripheral iridoplasty (LPI) is widely used in the treatment of glaucoma by flattening the iris and widening angle of anterior chamber (AA).However,no evidence suggests the optimal site of LPI in iris.Objective This study was to compare the therapeutic effects of LPI at different sites of iris for glaucoma.Methods Glaucoma models were established in the right eyes of 40 healthy adult male pigment rabbits by intrachamber injection of 0.1 ml compound carbomer solution with 0.3％ carbomer and 0.025％ dexamethasone.The models were randomly divided into model control group,corneoscleral limbus group,one spot from corneoscleral limbus group and two spots from corneoscleral limbus group.LPI was performed at corresponding site of iris by 532 nm argon laser with the spot diameter 500 μm,energy 300 mW,exposure time 0.3 seconds and laser number 24 spots,and the rabbits in the model control group did not receive LPI.Intraocular pressure (IOP),coefficient of outflow facility (C value) were measured and calculated with Schi(o)tz tonometer before LPI and 2,4,7,14 and 30 days after LPI,and anterior chamber depth (ACD),AA,anterior chamber angle opening distance within 500 μm radius from scleral spur (AOD500) were measured with ultrasound biomicroscope (UBM).The eyeballs were extracted 30 days after LPI,and the chamber angle were observed under the optical microscope after hematoxylin and eosin staining.The use and care of the animals complied with the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health.Results UBM showed that compared with the model control group,the anterior chamber angle was evidently widened in all the LPI groups,with the best effectiveness in the one spot from corneoscleral limbus group and the worst one in the two spots from corneoscleral limbus group.Compared with the model control group,the IOP was evidently reduced,and C values,AA and AOD500 were significantly increased in the corneoscleral limbus group,one spot from corneoscleral limbus group and two spots from corneoscleral limbus group after LPI,showing significant differences among the four groups (IOP:Fgroup =16.848,P ＜ 0.01;C value:Fgroup =9.629,P ＜ 0.01;AA:Fgroup =62.336,P＜0.01;AOD500:Fgroup =77.779,P ＜ 0.01).IOP was reduced and C value,AA and AOD500 were increased in 2,4,7,14 and 30 days after LPI as compared with before LPI,with significant differences over time (IOP:Ftime =3.041,P =0.011;C value:Ftime =4.311,P＜0.01;AA:Ftime =14.627,P＜0.01;AOD500:Ftime =20.378,P＜0.01).Compared with the model control group,the ACD was significantly increased in the corneoscleral limbus group and one spot from corneoscleral limbus group,and that in the two spots from corneoscleral limbus group was significantly reduced,and the ACD was insignificantly increased over time after LPI (Fgroup =18.017,P＜0.01;Ftime =0.022,P =1.000).Hematoxylin and eosin staining showed that the trabecular meshwork and adhesion of tissure were reopened and the anterior chamber angle was widened after LPI.Conclusions LPI can widen anterior chamber angle and lower the IOP.The best therapeutic outcome for glaucoma is displayed when LPI is performed at the iris site corresponding to one spot from the corneoscleral limbus.

Objective To investigate the effect of sevoflurane postconditioning on the activities of Na+ -K+ -ATPase and Ca2+ -Mg2+ -ATPase during myocardial ischemia-reperfusion (I/R) in rats and the possible mechanism. Methods Forty-five healthy male Wistar rats weighing 250-280 g were randomly divided into 3 groups ( n = 15 each): sham operation group (group S), I/R group and sevoflurane postconditioning group (group Spo). Myocardial I/R was induced by occlusion of anterior descending branch of left coronary artery for 30 min followed by 120 min of reperfusion. In group S the anterior descending branch was only exposed but not ligated. Group Spo received 5 min inhlation of 2.5% sevoflurane 1 min before reperfusion. The myocardial tissues were taken at 2 h of reperfusion for determination of infarct size and activities of Na+ -K+ -ATPase and Ca2 * -Mg2 * -ATPase. Results The infarct size was significantly larger and the activities of Na+ -K+ -ATPase and Ca2+ -Mg2+ -ATPase were signifi cantly lower in group I/R than in group S ( P ＜ 0.05). The infarct size was significantly smaller and the activities of Na+ -K+ -ATPase and Ca2+ -Mg2+ -ATPase were significantly higher in group Spo than in group I/R (P ＜ 0.05 ). Conclusion Sevoflurane postconditioning can reduce myocardial I/R injury through increasing the activities of Na+ -K+ -ATPase and Ca2+ -Mg2+ -ATPase.

Objective To investigate the effects of sevoflurane postconditioning on cardiomyocyte apoptosis during myocardial ischemia-repeffusion (I/R) in rats.Methods Forty-five healthy male Wistar rats weighing 250-280 g were randomly divided into 3 groups ( n =15 each):group sham operation ( group S) ; group I/R and group sevoflurane postconditioning (group Spo).The animals were anesthetized with intraperitoneal 3％ pentobarbital 45 mg/kg,tracheally intubated and mechanically ventilated.Myocardial I/R was produced by occlusion of anterior descending branch of left coronary artery for 30 min followed by 120 min reperfusion in groups I/R and Spo.In group Spo the animals inhaled 2.5％ sevoflurane for 5 min starting from 1 min before reperfusion was started.The animals were sacrificed at the end of 120 min reperfusion.Their hearts were removed for measurement of infarct size and the area at risk and determination of apoptotic index (the number of apoptotic cells/the total number of cells) and Bcl-2 and Bax protein and mRNA expression.Results Sevoflurane postconditioning significantly reduced infarct size in group Spo as compared with group I/R.There was no significant difference in area at risk between groups I/R and Spo.Myocardial I/R significantly increased the apoptotic index,Bcl-2 and Bax protein and mRNA expression in group I/R as compared with group S.Sevoflurane postconditioning significantly decreased apoptotic index and Bax protein and mRNA expression but increased Bcl-2 protein and mRNA expression in group Spo as compared with group I/R.Conclusion Sevoflurane postconditioning attenuates myocardial I/R injury by redncing myocardial apoptosis,up-regulating Bcl-2 expression and down-regulating Bax expression.

Objective To evaluate the role of Ras homolog family member A (RhoA)/Rho-associated coiled-coil containing protein kinase 2 (ROCK2) signaling pathway in propofol-induced neuroapoptosis in the hippocampus of newborn rats.Methods Experiment Ⅰ Primarily cultured hippocampal neurons were seeded in 6-well culture plates at a density of 1×106 cells/ml and divided into 2 groups (n=6 each) using a random number table:solvent control group (C group) and propofol group (P group).Propofol was added with the final concentration of 60 μg/ml in group P.Dimethyl sulfoxide was added with the final concentration of 0.04％ in group C.The expression of RhoA and ROCK2 in hippocampal neurons was measured by Western blot at 24 h of incubation.Experiment Ⅱ Primarily cultured hippocampal neurons were seeded in 6-well culture plates at a density of 1 × 106 cells/ml and divided into 3 groups (n =6 each) using a random number table:solvent control group (C group),propofol group (P group) and propofol plus specific RhoA/ROCK2 signaling pathway blocker Y27632 group (P+Y group).Propofol was added with the final concentration of 60 μg/ml in group P.Propofol at the final concentration of 60 μg/ml and Y27632 at the final concentration of 10 μmol/L were added in group P+Y.Dimethyl sulfoxide was added with the final concentrauon of 0.04％ in group C.At 24 h of incubation,the neuroapoptosis in hippocampi was detected by flow cytometry,and the expression of activated caspase-3 in hippocampal neurons was measured by Western blot.The apoptotic rate was calculated.Results Experiment Ⅰ Compared with group C,the expression of RhoA and ROCK2 in hippocampal neurons was significantly up-regulated in group P (P＜0.05).Experiment Ⅱ Compared with group C,the apoptotic rate of hippocampal neurons was significantly increased,and the expression of activated caspase-3 was up-regulated in P and P+Y groups (P＜0.05 or 0.01).Compared with group P,the apoptotic rate of hippocampal neurons was significantly decreased,and the expression of activated caspase-3 was down-rcgulatcd in group P + Y (P＜ 0.05).Conclusion Activation of RhoA/ROCK2 signaling pathway is involved in propofol-induced neuroapoptosis in hippocampi of newborn rats.

Objective To evaluate the role of conventional protein kinase Cγ (cPKCγ)/growthassociated protein-43 (GAP-43) signaling pathway in ketamine-induced apoptosis in hippocampal neurons of developing rats in an in vitro experiment.Methods Primarily cultured hippocampal neurons were seeded in culture plates at a density of 1×10.6 cells/ml and divided into 2 groups (n=10 each) using a random number table:control group (C group) and ketamine group (K group).Group C received no treatment.Ketamine was added with the final concentration of 300 μmol/L in group K.At 12 h of culture or incubation,the apoptosis in hippocampal neurons was detected by flow cytometry.The apoptotic rate was calculated.The expression of cPKCγ,GAP-43 and phosphorylated GAP-43 in hippocampal neurons was measured by Western blot.Results Compared with group C,the apoptotic rates of hippocampal neurons were significantly increased,and the expression of cPKCγ,GAP-43 and phosphorylated GAP-43 was down-regulated in group K (P<0.01).Conclusion The mechanism by which ketamine induces apoptosis in hippocampal neurons of developing rats may be related to inhibition of cPKCγ/GAP-43 signaling pathway activation in an in vitro experiment.

Objective To explore the clinical effect of percutaneous vertebroplasty (PVP) in the treatment of old unstable osteoporotic vertebral fracture.Methods Twenty cases of old unstable osteoporotic single vertebral body fracture were divided into the stable group and unstable group according to the imaging results,10 cases in each group.PVP was performed in all 20 cases.The VAS scores of waist bending activity at preoperative 30 min,postoperative 6 h,3,30 d were observed.The changes of anterior edge height of spinal body in the injured vertebral segment of erect position and horizontal position were compared before and after operation.Results The VAS score of waist bending at preoperative 30 min had statistical difference between the two groups (P＜0.05).Compared with at preoperative 30 min,the VAS scores at postoperative observation points in the two groups were significantly decreased with statistical difference (P＜0.05).The VAS scores at postoperative 6 h,3,30 d had no statistical difference between the two groups(P＞0.05).The changes of posterior edge height of spinal body in the stable groups had no statistical difference before and after operation (P＞0.05);the anterior edge height of spinal body after treatment in the unstable group were significantly changed compared with before operation,and the difference was statistically significant(P＜0.05).Conclusion Preoperative pain in the patients with unstable osteoporotic vertebral fracture is more obvious than that in the patients with stable osteoporotic vertebral fracture.But all have similar effect after PVP therapy;the postoperative height in unstable osteoporotic vertebral fracture can obtain a certain recovery after PVP.

Objective: To investigate the effect of micro RNA (miR)-335-5p regulating bone morphogenetic protein 2 (BMP-2) on the osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs). Methods: hBMSCs were cultured in vitro and randomly divided into control group (group A), miR-335-5p mimics group (group B), miR-335-5p mimics negative control group (group C), miR-335-5p inhibitor group (group D), and miR-335-5p inhibitor negative control group (group E). After grouping treatment and induction of osteogenic differentiation, the osteogenic differentiation of cells in each group was detected by alkaline phosphatase (ALP) and alizarin red staining; the expressions of miR-335-5p and BMP-2, Runt-related transcription factor 2 (Runx2), osteopontin (OPN), and osteocalcin (OCN) mRNAs were detected by real-time fluorescence quantitative PCR analysis; the expressions of Runx2, OPN, OCN, and BMP-2 proteins were detected by Western blot. Results: Compared with group A, the relative proportion of ALP positive cells and the relative content of mineralized nodules, the relative expressions of BMP-2, miR-335-5p, OPN, OCN, Runx2 mRNAs, the relative expressions of Runx2, OPN, OCN, and BMP-2 proteins in group B were significantly increased ( P<0.05); the above indexes in group D were significantly decreased ( P<0.05); the above indexes between groups C, E and group A were not significantly different ( P>0.05). Conclusion: miR-335-5p can up-regulate BMP-2 expression and promote osteogenic differentiation of hBMSCs.

Lung cancer is one of the most common malignant tumors and has higher morbidity and mortality world-wide. The genetic and environmental risk factors play important role in the process of tumorigenesis and cancer development. Cigarette smoking and some other environmental risk factors may be involved in the pathogenesis of lung cancer, including polycyclic aromatic hydrocarbons, particulate matter and occupational exposure. Studies have shown that epigenetic mechanism such as DNA methylation plays an essential role in relationship between lung cancer and the environmental risk factors. Aberrant DNA methylation is considered to be an early event during the process of lung cancer occurrence. This paper summarizes the recent studies on association of lung cancer and aberrant DNA methylation of genes induced by several common environmental factors including smoking, polycyclic aromatic hydrocarbons and particulate matter. Further understanding of DNA methylation in linking mechanism between lung cancer and environmental risk factors will help to clarify the pathogenesis and development of lung cancer. Moreover, abnormal DNA methylation induced by environment factors may be a potential epigenetic biomarker for early diagnosis and prognostic prediction of lung cancer.