BACKGROUND: Reactive oxygen species (ROS), including superoxide anion (O2) and hydrogen peroxide (H2O2), have been recognized as specific second messengers in signaling cascades involved in the growth and differentiation of cells.The generation of excessive ROS initiates cardiomyocyte apoptosis. This paper is aimed to corroborate the hypothesis that excessive amounts of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) 4-dependent ROS can suppress cardiomyocyte differentiation from embryonic stem (ES) cells by induction of apoptosis.OBJECTIVE: To investigate the role of NOX4 ROS on cardiomyocyte differentiation.DESIGN: Randomized and controlled trial observation.SETTING: Beijing Institute of Geriatrics, Beijing Hospital, Key Laboratory of Geriatrics, Ministry of Health.MATERIALS: ES cells were preserved by the laboratory of Beijing Institute of Geriatrics, Beijing Hospital, Key Laboratory of Geriatrics, Ministry of Health, while other reagents were purchased from Sigma Company without specific notification.METHODS: The experiment was carried out in the Beijing Institute of Geriatrics, Beijing Hospital, Key Laboratory of Geriatrics, Ministry of Health from October 2003 to August 2005. ①Mouse ES cells were differentiated into embryoid bodies (Ebs). At day 4, Ebs were managed for one hour with different concentrations (1, 10, 100, 1 000 nmol/L) of H2O2, and generation of cardiomyocytes was observed at day 8 to analyze the effect of ROS on cardiomyocyte differentiation.② CGB8 cells transfected with pcDNA3.1 and pcDNA3.1-NOX4 respectively were adopted, while those untransfected CGB8 cells were taken as controls. A quantitative NBT (nitro blue tetrazolium) test was used to measure NOX4 ROS generation. The levels of NOX4 mRNA and MLC2v protein were assayed by RT-PCR and western blotting, respectively.③ROS scavenger N-acetylcysteine (5 mmol/L) and catalase (200 U/mL) managed for 2 hours before transcription were taken as controls to observe the apoptosis of CGB8 cell after NOX4 overexpressed and the expressions of p21, p53 and Bcl-2 of NOX4-transfected CGB8 cells simultaneously. And the apoptosis of ES cells was detected by Hoechst staining, TUNEL assay and DNA fragmentation.MAIN OUTCOME MEASURES: ①the role of ROS on cardiomyocyte differentiation.②the expression of NOX in ES cells.③ the production of ROS, differentiation of cardiomyocyte and apoptosis of ES cells after NOX4 overexpressed.④the feasibility that p53, p21 and Bcl-2 are involved in the apoptosis induced by the overexpression of NOX4.RESULTS: ①Different concentrations of ROS play different roles on cardiomyocyte differentiation. The exposure of Ebs to 1-100 nmol/L H2O2 for 2 hours at day 4 of culture leaded to an enhanced beating activity (P ＜ 0.01 or 0.001), whereas 1 μmol/L H2O2 depressed cardiomyocyte differentiation as compared with control conditions (P ＜ 0.001). ②NOX4 was highly expressed in ES cells, while NOX1 and NOX2 were absent and only a weak band of NOX3 was detected. The results from RT-PCR revealed that NOX4 overexpressed in CGR8 transfected with pcDNA3.1-NOX4 as compared to the control.③NBT result proved that highly-expressed NOX4 produced the excessive amounts of ROS (P ＜ 0.05). The beating activity was remarkably reduced in NOX4-overexpressing Ebs as compared to wild-type or mock-transfected Ebs (P ＜0.01). Moreover, a severe reduction of MLC2v protein in NOX4-overexpressing Ebs was also observed by western blot analysis.④p21 and p53 may be responsible for the NOX4-induced apoptosis. P53-/- cells R72D27 transfected by NOX4 did not induce apoptosis. NOX4 overexpression induced ES cell apoptosis could be prevented by overexpression of Bcl-2.CONCLUSION: The findings of this paper highlight the role of NADPH oxidase NOX4 in cardiomyocyte differentiation from ES cells. P53, p21 and Bcl-2 could be involved in the apoptotic pathway.

Large amount of data including chemical composition and size information of individual particles would be generated in the measurement of aerosol particles using atmospheric aerosol time-of-flight mass spectrometry ( ATOFMS ) . Our home-made ATOFMS was used to measure the indoor individual aerosol particles in real-time for 24 h, and the obtained mass spectrometric data were clustering analysis by self-organizing map ( SOM ) because of its ability of vector quantization and data dimensionality reduction. 20 classification results were got which includedCalcium-Containing,Salt+Secondary particles,Secondary particles,Organic Amines,K+-Rich Organics andSoil particles, etc. Compared with previous mass spectrometric methods, SOM is a natural visualization tool, more classification results can be obtained. This classification information would be useful to assess the response and toxicity of atmospheric aerosol particles and identify the origin of atmospheric aerosol particles.

Objective To investigate the existence of pulmonary vascular remodeling after left pneumonectomy in rats and the role of hypoxia inducible factor-lα( HIF-1α) and vascular endothelial growth factor ( VEGF) in pulmonary vascular remodeling.Methods Twenty-four healthy male Sprague-Dawley rats were randomly divided into experimental and control groups, 12 in each group.The rat models of pulmonary vascular remodeling were created by open-chest left pneumonectomy.After 12 weeks of feeding, the mean pulmonary artery pressure ( mPAP) and partial pressure of arterial oxygen ( PaO2 ) of each rat were measured.The ultrastructure of small arteries in the lung specimens were examined by e-lectron microscopy.Muscularized degree of three kinds of small pulmonary vessels ( muscularized artery MA, partially mus-cularized artery PMA, and non-muscularized artery NMA) were observed by light microscopy, and the percentage of each kind of pulmonary arteries ( MA%, PMA%, NMA%) were calculated.Arterial external diameter, media thickness of ves-sel ( MTV) , total vascular area, media area of vessel ( MAV) , MTV%and MAV%were calculated as indicators of pul-monary vascular remodeling.The expressions of HIF-1αand VEGF in artery were detected by immunohistochemistry.Re-sults The values of mPAP, MA%, PMA%, MTV, MAV, MTV% and MAV% in the experimental group were signifi-cantly higher than those in the control group (P<0.01), but the value of PaO2 and NMA%were significantly lower than those in the control group (P<0.01).The IOD value of HIF-1αand VEGF expressed in the pulmonary arterial wall of the experimental group were 26.47 ±4.16 and 42.04 ±3.79, respectively, significantly higher than those in the control group (6.12 ±2.14 and 11.53 ±2.29, P<0.01).Linear correlation analysis showed that the expression of HIF-1αand VEGF was positively correlated with MTV% and MAV%, negatively correlated with PaO2 , and the HIF-1αexpression was posi-tively correlated with VEGF expression.Conclusions A rat model of pulmonary vascular remodeling can be successfully established by left pneumonectomy.Hypoxia is a key factor in the development of pulmonary vascular remodeling, HIF-1αand VEGF may play an important role in its pathogenesis.

Objective To evaluate the preoperative nutritional status of patients with gastric cancer by using patient-generated subjective global assessment (PG-SGA) its relationship with postoperative results.Methods Make a preoperative nutrition assessment by using PG-SGA for 87 patients with gastric carcinoma who can be treated with operation.Analyze the effects of the postoperative complication,survival rate and the hospitalization time on patients.Results The number of patients of this group who can process the preoperative nutrition assessment by using PG-SGA accounts for 100％ of the total.There are 37 patients with moderate or severe malnutrition before operation (account for 39.1％).The complication incidence of patients with moderate or severe malnutrition before operation and the patients with no or mild malnutrition are 52.9％ and 5.7％ separately (P ＜ 0.01).The sensitivity and specificity of PG-SGA score for predicting complications are 85.7％ and 75.8％ respectively.And we should implement the analysis of regression to verify that the PG-SGA grade is the independent risk factor of postoperative complication by applying multiple Logistic(P ＜ 0.01).The average hospital stays of patients with moderate or severe malnutrition before operation and the patients with no or mild malnutrition are (21 ± 6) d and (16 ± 4) d separately (P ＜ 0.01).Conclusion According to the PC-SCA results,patients with moderate or severe malnutrition before operation have increased susceptibility to complication and the extension of hospital stays after the operatiou.Therefore,it is important to supply the proper nutrition support to these patients.

Both motor therapy and functional electrical stimulation was used to treat the patients with hemiplegia. In 43 patients 17 cerebral haemorrhages, 26 cerebral infarctions and 7 traumatic cerebral injuries and haematomas male 28, female 15,were included in our studies. Therapy began in 1 to 2 weeks after lesion in 23, 2 to 5 weeks in 14, 3 -4 months in 3,6 months in 1,8 months in 1 and 1 year in 1. The results were evaluated by Barthel Index before and after the treatment,increasing from 41.02 ±20.58 pretreatment to 85.70±16.46 post-treatment with 22.58±15.10 times

Objective To develop mycobacterial inducible expression vectors which permit to overexpress Mycobacterium tuberculosis (Mtb) immunodominant antigen, and to analyze its immunogenicity after purification by affinity chromatography. Methods The regulatory region of M. smegmatis (Ms) acet-amidase(pACE) was obtained by PCR amplification, and was used as promoter to construct the mycobacteri-al inducible expression vectors, pMF series. The coding gene of Mtb chimeric antigen Ag856A2 which is a recombinant Ag85A with 2 copies of ESAT-6 inserted in its Acc Ⅰ site and showed excellent immunogenicity in the animal experiments we described previously, was cloned into the pMF vector series, and was induced to express by the addition of acetamide. The recombinant protein expressed in the Ms was purified by the Ni~(2+)-NTA affinity chromatography, the resulted homologous recombinant antigen was added into the spleen cells separated from BCG vaccinated mice, and the immunogenicity was analyzed by the IFN-γ ELISPOT as-say. Results The mycobacterial inducible expression vectors, pMF series was constructed successfully, target antigen could be. induced to express in the Ms by the addition of 0.02% acetamide, and could be puri-fied by the Ni~(2+) -NTA affinity chromatography due to the addition of 6×His tag in the vector pMF406. Fur-thermore, the mycobactefial homologous antigen could induce more IFN-γ secretion than the heterogonous one. Conclusion The mycobacterial inducible expression system based on the regulatory region of Ms acet-amidase as promoter could permit the Mtb target antigen of interest overexpression and purification, and the immunogenicity of the homologous antigen from Ms is better than that of be expressed from E. coli, which may be more potential for immunological detection of tuberculosis.

Objective To study the organ damage in patients with systemic lupus erythematosus (SLE) and its significance in prognosis. Methods The SLE damage index (SDI) was used to evaluate 121 female and 15 male patients. Results Among these patients, the lowest and the highest SDI was 0 or 9, separately (average: 1.79 ? 2.22). Most damaged organ was renal (39.0%), followed by muscle/skeletal system (19.9%) and cardiovascular system (18.4%). There was statistically significant difference of SDI in different groups of prognosis, life condition or disease duration ( 10 years). However, there was no significant difference of SDI in different sex groups. The correlation analysis revealed that the age at onset or diagnosis was not related to SDI. Conclusion SDI is effective in evaluating the organ damage in predicting prognosis of SLE patients.

Objective To express and purify mouse interleukin 17A(mIL-17A) in E. coli and to analyze its ability of stimulating macrophage inflammatory factors expression. Methods The coding gene of mouse mIL-17A mature protein was amplified from mouse spleen cells by RT-PCR. PCR product was cloned into the prokaryotic expressing vector pET28a, and the resulting recombinant plasmid pET28a/mIL-17a was then transformed into the host E. coli strain BL21(DE3) for expression. The mIL-17A protein was identified by SDS-PAGE and Western blot. The recombinant protein was purified by the Ni-NTA affinity chromatography, and was further tested on the stimulation of cytokine and chemokine of RAW264.7 cells by ELISA and real-time quantity PCR in vitro. Results The mIL-17A with bioactivity was over-expressed and purified successfully, and the results of real-time PCR and ELISA showed that recombinant mIL-17A stimulated macrophage mRNA upregulation of IL-6, defensin β2 and Cxcl3 and secretion of defensin β2, Ccl3, Cxcl3,IFN-γ, IL-6 and IL-4. Interestingly, these effects could be blocked by the addition of anti-IL-17A neutralizing antibody partly. After treatment with mIL-17, 74. 87-fold of defensin β2 mRNA expression was increased comparing with that of untreated cells( P ＜0.01 ), while blocking with anti-IL-17A antibody the increase was only 5.4-fold(P ＜ 0.01 ). Conclusion The recombinant mIL-17A has a strong stimulation on secretion of cytokine and chemokine of macrophage, that maybe result to the enhancement of anti-infection ability of macrophage.

Objective To investigate the influence of the hematoma involving the aortic arch in endovascular aortic repair of complicated type B intramural aortic hematoma. Methods A total of 69 patients[58men; mean age(58.1±8.9)years; range 38-77]underwent endovascular repair between February 2011 and June 2015 were retrospectively reviewed. Patients with hematoma involving about the left subclavian artery level were categorized as group A(n=28) and patients without hematoma involvement to the aortic arch were categorized as group B (n=41). Results All the patients were treated with coverd aortic stents. The success rate was 97.1％ with complete isolation of lesion in 67 patients. The average follow-up period was(19.6±14.1)months. During perioperative period, no procedure related deaths was recorded. Perioperative complications include paraplegia in 1case(1.4％) in group B and stent graft-induced new entry in 2 cases(2.9％) in group A. During the follow-up period 1 case in group A within 1 month and another 1 case in group B within 1 year developed new entries at proximal end of stents. 1 case (1.4％) in group B had asymptomatic type Ⅰ endoleak 2 years after TEVAR. Conclusions Type B aortic intramural hematoma with arch involvement is not a risk factor of stent-induced new entry in perioperative period after endovascular treatment and further studies are needed. Strict control of blood pressure is essential for the prevention of stent-related complications.

Objective To prepare a conjugate vaccine by linking Haemophilus influenzae type b (Hib)polysaccharide to PsaA protein carrier and evaluate the immunogenicity and efficacy of the conjugate vaccine. Methods A recombinant protein rPsaA,expressed by using the genetic engineering technology, was used as a protein carrier to prepare conjugate vaccine together with Hib polysaccharide. Ten mice at age of 3 weeks were immunized with the conjugate vaccine,while another 10 age-matched mice were immunized with Hib-tetanus toxoid(Hib-TT)vaccine which was produced formerly as a control. The mice treated with equal volume of PBS were set up as the negative control. The IgG antibodies in serum samples against PsaA and Hib polysaccharide were detected in two weeks after the final immunization. A suspension of Pneumococ-cus was injected into the middle ears of mice from experiment and control group. Histopathological analysis was performed to measure the clearance of bacteria in the middle ears and the severity of infection on days 3 and 7 after bacterial challenge. Results The rPsaA protein was prepared by the genetic engineering tech-nology and purified successfully with anion-exchange column. The Hib polysaccharide-PsaA protein conju-gate vaccine was prepared through a series of amide condensation reactions. The detection of IgG antibodies against PsaA protein and Hib polysaccharide in the immunized mice demonstrated that there was no signifi-cant difference with the titer of IgG against Hib polysaccharide between the mice immunized with the Hib-PsaA conjugate vaccine and those immunized with the Hib-TT vaccine. Less Pneumococcus strains were de-tected in the middle ears of mice immunized with the conjugate vaccine than those mice immunized with the Hib-TT vaccine three days after challenge. The mice from control group showed severe inflammation in the middle ears than those from experiment group. The Hib polysaccharide-PsaA protein conjugate vaccine im-proved protection against Pneumococcus infections as compared with the Hib-TT vaccine. Conclusion The rPsaA protein could be produced by genetic engineering technology and purified by anion-exchange column. The Hib polysaccharide was successfully conjugated with the rPsaA protein through amide condensation reac-tion. Both anti-PsaA and anti-Hib immune responses were induced in young mice by the injection of Hib pol-ysaccharide-PsaA protein conjugate vaccine. Apart from providing protection against Hib infection,the con-jugate vaccine might also be used for the prevention of acute otitis media caused by Pneumococcus infection.