Objective To construct eukaryotic expression plasmid pEGFP-N1/EPO and to detect its expression in vitro in order to provide experimental evidence for further study on the treatment of hypoxic-ischemic brain damage.Methods The total RNA was extracted from rat kidney.EPO cDNA fragment was obtained by RT-PCR amplication,inserted into pEGFP-N1 vector and identified by restriction endonuclease digestion and nucleotide sequencing.MSCs cells were transfected with the plasmid using Lipofectamine 2000.The expression of EPO protein was detected by immunohistochemistry technique and RT-PCR.Results Confirmed by restriction endonuclease digestion and sequencing,the recombinant plasmid contained the sequence of rat EPO gene was successfully constructed.After transfection with the plasmid,EPO protein could be expressed in MSCs cells.Conclusion The constructed eukaryotic expression vector pEGFP-N1/EPO can express rat EPO in vitro.

Objective To observe the effects of chemical compounds exposure and hyperthermal environment to pregnant rats on the hippocampus neuronal morphology and neurobehavioral development of their offsprings. MethodsPregnant rats were divided into 4 groups randomly: the control group; chemical compounds exposure group, chemical compounds combined hyperthermal exposure group and hyperthermal group. The control group was living in normal condition; the chemical compounds exposure group in a cabin with benzene (20.26?0.80) mg/m3, toluene (39.66?4.23) mg/m3, dimethylbenzene (42.40?2.85) mg/m3, and formaldehyde (23.13?1.30) mg/m3; the hyperthermal group was kept in an atmosphere of 38.5 ℃; and the combined group was exposed to the chemical compounds and high temperature at the same time. All animals were treated respectively 2 h per day for 10 d since their pregnancy. Their offspring were lived in normal condition. The development of brain and changes of hippocampus neuronal morphology were observed just or 1 month after born. The primary reflex activity of new-born rats was also examined. ResultsThere was no abnormality in the brain of new born rats from 4 groups. In the chemical compounds exposure and the combined exposure groups, the number of degeneration and necrosis neurons in CA1 region of hippocampus were significantly more than those in control and hyperthermal groups (P

Objective To investigate the long-term efficacy of transendocardial injection of vascular endothelial growth factor (VEGF) into ischemic porcine myocardium.Methods Thirty pigs were divided into two groups:control group and treatment group (15 each).A angioplasty balloon was used to occlude the left anterior descending coronary arteries of the animals to reproduce a porcine myocardial ischemic model.Blank plasmid (control group) or constructed pIRES2-EGFP-hVEGF165 eukaryotic expression plasmid (treatment group) was directly injected into the ischemic myocardium through an transendocardial injection catheter of NOGA system,respectively.Before and one year after injection,left ventricular electromechanical mapping (LVEMM) was used to detect local linear shortening (LLS) and unipolar potential (UpV);M-mode local wall motion,cycle variation of integrated backscatter (CVIB) and left ventricular ejection fraction (LVEF) were detected by ultrasonagraphy.One year later,the animals were sacrificed for the observation of the capillary formation in myocardial tissue with immunohistochemical staining.Results One year after injection,as compared with that of control group,local linear shortening (LLS) showed a significant increase (P0.05),and ultrasonagraphy showed better heart function in the treatment group (P

Object To optimize the process for the extraction of the original recipe used in the treatment of eczema to give a new ECZEMA SPRAY dosage form Methods The extraction process was studied by orthogonal experimental design as guided by determining the content of paeonol and baicalin in the extract Results The optimal extraction process was to reflux the original recipe with 80% ethanol twice at a bath temperature of about 90 ℃ for 1 5 and 1 0 h respectively The amount of ethanol used for each extraction was 10 and 8 times of the original recipe respectively Conclusion The above extraction process gave the most rational and satisfactory results

Objective To investigate the influence of CO2 pneumoperitoneum on the adhesive and invasive ability of gastric cancer cells based on the expression of adhesive and invasive molecules. Methods With an artificial CO2 pneumoperitoneum model in vitro, human gastric cancer cells MKN-45, SGC-7901 and MKN-28 were exposed to 3 different CO2 gradients: 9 mm Hg, 15 mm Hg and control group (0 mm Hg). The expression of E-cadherin, ICAM-1, MMP-2 and VEGF-A were measured at 2 and 4 hours exposure by using RT-PCR, CytoMatrixTM kit and ECMatrixTM kit. The pretreated gastric cancer cells were injected into abdominal cavity of nude mice(2×106 cells per mouse). Five mice in each group were sacrificed 4 weeks later to record the number of tumor nodules in abdominal cavity. The remaining mice were kept for observation of survival time. Results The expression of E-cadherin (MKN-45: from 2.26 to 2.19, SGC-7901 :from 2.16 to 2.09、MKN-28 :from 2.06 to 1.99), ICAM-1 (MKN-45 : from 2.20 to 2.28、SGC-7901: from 2.10 to 2.18、MKN-28: from 2.00 to 2.08), MMP-2 (MKN-45:from 2.05 to 2.13、SGC-7901: from 1.95 to 2.03、MKN-28: from 1.85 to 1.93) and VEGF-A(MKN-45 : from 2.10 to 2.16、SGC-7901 :from 2.00 to 2.06、MKN-28: from 1.90 to 1.96) didn't change significantly with increasing pressure and time (P>0.05). The expression of adhesive and invasive molecules didn't change significantly between the experimental groups and the control group. There was no statistical significance of tumor metastasis in abdominal cavity of nude mice(MKN-45:from 22 to 23、SGC-7901 :from 20 to 22、MKN-28:from 21 to 22) and survival time(MKN-45 :from 23 to 21、SGC-7901 :from 22 to 21、MKN-28 :from 22 to 21) among all the groups. Conclusion Under low pressure and short time of CO2 exposure, the adhesive and invasive capacity of gastric cancer cells did not change significantly hence did not increase the possibility of neoplasm metastasis.

Objective To study the risk factors related to recurrence of gastrointestinal stromal tumor (GIST) after discontinuing postoperative adjuvant imatinib mesylate (IM) treatment.Methods We retrospectively analyzed our clinical database of 138 GIST patients who received radical resection and subsequent IM adjuvant treatment at the Renji Hospital,Shanghai Jiaotong University School of Medicine between January 2006 and January 2014.Results For the entire Multivariate analysis study group,the overall 5-year recurrent free survival (RFS) rate was 54.5％.There were two tumor characteristics which were independent prognostic factors of GIST treated by postoperative IM:Ki67 index (P =0.005),and serosal invasion (P =0.026).The accuracy of comprehensive evaluation based on the two weighted variables was better than NIH staging criteria(AUC:0.714 vs.0.631).Furthermore,two risk groups were created according to the risk model with 5-year RFS of 81.3％ and 31.1％ as low-risk and high-risk groups,respectively (P ＜0.05).Conclusions For patients with intermediate or high risk in NIH classification,if there was tumor serosal invasion,or if there was no local invasion but Ki67 index ＞ 8％,extended continuous IM adjuvant treatment should be recommended after the primary tumor was radically resected.

Objective To evaluate the radiation protection of occupational hazards in a hospital Clinac 23EX medical electronic accelerator construction project so as to ensure the health and safety of the relevant people involved.Methods According to the relevant laws,regulations and standards of China,combined with the relevant materials provided by the construction unit,the radiation protection tests and comprehensive assessment of this project were carried out.Results The performance test results of the medical electron accelerator met the requinements of GB/T 19046-2013 The ambient dose equivalent rate in the workplace was between the background dose rate (0.10 μ,Sv/h) and 11.5 μSv/h,which suggested the computer room shielding met the requirements of radiation protection.The total body effective dose,the for 7 radiation workers were 0.85,1.19,1.64 mSv,respectively,which were lower than the dose management control values of the construction unit and the national standards.Radiation protection supplies and the management system of the construction unit met the national requirements.Conclusions The construction project can effectively control the radioactive occupational hazard factors in normal operation,and the radiation protection facilities have reached the completion requirements.

BACKGROUND: Previous studies have confirmed that embryonic stem cells call be induced to differentiate into insulin-producing cells, but the induction process takes a long time. Most of the processes take about one month.OBJECTIVE: Activin A, all-trans retinoic acid (ATRA), basic fibroblast growth factor (bFGF) and nicotinamide were applied in vitro in combination to observe whether mouse embryonic stem cells could be induct to differentiate into insulin-producing cell in a relatively short time.DESIGN: Cell observation experiment.SETTING: Shanghai Institute of Endocrine and Metabolic Diseases, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China.MATERIALS: This study was performed at Shanghai Institute of Endocrine and Metabolic Diseases from October 2004 to February 2006. Two mice of clean grade and of 12.5-14.5 days of gestational age were provided by Shanghai SLAC Laboratory Animal Co., Ltd (Permission No. 2004A034). The protocol was performed in accordance with ethical guidelines for the use and care of animals. Mouse embryonic stem cell lines were supplied by Dr Changxian Zhang (CNRS UMR5641, France). Activin A was the product of the R & D Corporation. ATRA and nicotinamide were supplied by the Sigma Corporation, USA. BFGF was supplied by Gibco Corporaion. METHODS: Head and viscera were removed from embryos of the pregnant mouse. The remaining tissues were cut into pieces and digested with trypsin. Cell suspension was centrifuged and inoculated at 3×108L-1. The cells could be used as mouse feeder layer after 2-3 times of passage. The mouse embryonic stem cells (ESCs) were inoculated onto the feeder layer in knockout Dulbecco's modified Eagle medium (DMEM) supplemented with leukemia inhibitory factors (LIF). ESCs were passaged at 1:3-1:6 after 2-3 days of culture. Culture medium with serum was added into the culture dishes to terminate the digestion. Cell fluid was centrifuged and supernatant was discarded. The sediments were prepared into suspension and inoculated at 2.5×104 with LIF-free culture medium. After 24-48 hours, embryonic bodies (EBs) were collected and replated in 1% Matrigel-coated dishes. When began to adhere to the dishes, EBs were cultured in 10% FBS/DMEM supplemented with 100μg/L activin A for 24 hours. Then EBs were switched to 10% FBS/DMEM for 6-8 hours as an interval. After this interval. EBs differentiated were cultured in 10% FBS/DMEM with 10<-6mol/L RA for another 24 hours followed by culture in 10% FBS/DMEM supplemented with 10μg/L bFGFs for 3-5 days. Finally, EBs differentiated were cultured in DMEM/F12 supplemented with N2 supplement, B27 supplement, 1μg/L laminin, 10μg/L bFGFs, and 10mmol/L nicotinamide for 3-5 days. Dithizone (DTZ) staining, inununofluorescent staining and reverse transcription-polymerase chain reaction (RT-PCR) were applied to detect insulin expression in the differentiated cells.MAIN OUTCOME MEASURES: Induction of ESCs, DTZ staining and immunofluorescent staining as wel as RT-PCR detection.RESULTS: Mouse ESCs growing on a feeder layer formed many colonies with clear boundary and dense structure. However, there was no obvious outer limit between these ESCs. EBs began to adhere to the dishes, which were coated with matrigel, on the 2nd day. After activin A and ATRA interval induction, EBs spread, and most of the living cells were epithelial cell-like when cultured in 10% FBS/DMEM supplemented with 10μg/L bFGFs. After culturing in DMEM/F12 supplemented with N2, B27, nicotinamide, bFGFs and laminin, the cells formed small clusters. The insulin-producing cells were stained dark red with DTZ, and the cells stained with primary antibody to insulin were insulin-positive. After 2 weeks of induction of activin A, ATRA, bFGFs and nicotinamide, the insulin-producing cells expressed insulin 2, Pdxl, Nkx6.1, Nkx2.2, PP, IAPP, Glut2, Somastatin, Hnf3β and Neuro D mRNA but did not express insulin 1 mRNA.CONCLUSION: Mouse ESCs call be induced to differentiate into insulin-producing cells by activin A, ATRA, bFGFs and nicotinamide in vitro. Induction time call be shortened to 2 weeks.

Objective To evaluate the safety and efficacy of bipolar plasmakinetic system in exfoliative tran-surethral resection of bladder tumors .Methods Clinical data of 72 patients with non muscle invasive bladder cancer (NMIBC) were retrospectively analyzed.Transurethral bipolar plasmakinetic system was used ,30°viewer,F27 outer sheath was pushed off and bladder tumor was cut .When the bladder filling state ,pushed off bladder mucosa distance from tumor basal 2cm,then electricity cut the exfoliative bladder tumor .When bladder half filling state electricity cut the base of the bladder muscle layer of bladder tumor .Results This group of 72 cases were successfully completed surgery,surgery time 37~93 min,without intraoperative bladder perforation ,slight obturator nerve reflex in 5 cases. The keeping intact pathologic specimens was good for pathological staging .Conclusion Exfoliative transurethral resection of bladder tumors with bipolar plasma is a safe , practical and effective way of operation , which can avoid severe obturator reflex occurred in the operation , and greatly reduce the occurrence of bladder perforation , without TURS,surgical removal of the pathological specimens is specification .

Objective: To analyze the electrocardiographic (ECG) characteristics for the ifrst diagonal branch of infarction related artery (IRA) in patients with acute ST-segment elevation myocardial infarction (STEMI) in order to ifnd the rule for physician to make quick diagnosis. Methods: A total of 28 STEMI patients with coronary angiography (CAG) confirmed first diagonal branch of IRA were retrospectively analyzed. The patients were treated in our hospital from 2005-01 to 2014-06 and their ECG changes at admission were studied for ST-segment elevation/depression and q wave, Q wave changes during the period of evolution at different leads in all patients. Results: CAG presented that there were 19/28 (67.9%) patients with single vessel disease, 13 (46.4%) with isolated diagonal lesion. From onset of chest pain to AMI graph shown on ECG was about 240 (252 ± 71) min in all patients. All 28 (100%) patients were with ST-segment elevation in lead aVL, 27 (96.4%) in lead I, and 15 (55.6%) patients with ST-segment elevation by (0.5-1.0) mm. The incidence of ST-segment elevation in the chest lead was, in turn as 21 (75.0%) patients in lead V2, 16 (57.1%) in lead V3 and 12 (42.9%) in lead V1respectively; while ST-segment depression was as 28 (100%) patients in lead III, 27 (96.4%) in lead aVF and 22 (78.6%) in lead II respectively. During the period of evolution, the most q wave or Q wave formation were, in turn as 22 (88.0%) patients in lead aVL, 10 (40.0%) in lead V2, 9 (36.0%) in lead V3 and 7 (28.0%) in lead I respectively. Conclusion: The ECG changes in STEMI patients with diagonal branch of IRA have the high prevalence of ST-segment elevation in lead aVL and lead I, while there is an important feature that the ST-segment elevation < 1 mm in about half amount of relevant patients.