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Biologic effect of heavy ion radiation on human peripheral blood derived T lympho-cytes

Fengjun XIAO ; Yuefeng YANG ; Hua WANG ; Huiyan SUN ; Qunwei ZHANG ; Lisheng WANG

Military Medical Sciences.2014;(8):630-632,637. doi:10.7644/j.issn.1674-9960.2014.08.015

Objective To study the effect of heavy ion radiation on proliferation and apoptosis of human peripheral blood derived T lymphocytes and the mechanism .Methods T lymphocytes were isolated from heparinized whole blood samples by density gradient centrifugation using Ficoll before being irradiated with heavy ion beams 12 C.The accumulated absorbed dose (dose-rate values=0.5 Gy/min, and meanLET=29 keV/μm).12 h and 24 h post-infection, total RNA of T lymphocytes was isolated , and the apoptosis related gene expression , including Bcl-2, Bax, Caspase3, Caspase8 and Caspase9, was detected by RT-RT-PCR.24 h and 48 h after irradiation, the proliferation was analyzed by CCK 8 kit.The cell apoptosis was detected by flow cytometry after being labeled with AnnexinV-PE/7-AAD or AnnexinV-FITC/PE.The expression of Bcl-2, Bax and Caspase3 was also assayed by RT-PCR.Results Data showed that heavy ion radiation could inhibit the proliferation of T lymphocytes obviously , and the inhibition ratio in cells that received 2 Gy dose was much high-er than in cells that received 1 Gy dose.Furthermore, heavy ion radiation promoted the apoptosis of T lymphocytes signifi-cantly.The results of RT-PCR showed that the mRNA expression of Bcl-2 was down-regulated in heavy ion radiation T lym-phocytes while the expression of Bax and Caspase 3 was up-regulated.Conclusion Heavy ion radiation can inhibit the pro-liferation and promote the apoptosis of human peripheral blood derived T lymphocytes .

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Biological activity and application study of a monoclonal antibody against human Tim-3

Chunmei HOU ; Xiaoling LANG ; Yan XIAO ; Mingke ZHENG ; Xingwei JIANG ; He XIAO ; Guojiang CHEN ; Renxi WANG ; Jiannan FENG ; Gencheng HAN ; Beifen SHEN ; Yan LI

Military Medical Sciences.2014;(8):617-620,625. doi:10.7644/j.issn.1674-9960.2014.08.012

Objective To develop a human Tim-3 specific monoclonal antibody and evaluate its biological activity and possible use in clinical diseases associated with dysregulated Tim-3 expression .Methods The BALB/c mice were immu-nized by conventional method, and positive clones were used to develop anti-human Tim-3 antibody, the binding and neutralization activities of which in vitro and in vivo were investigated.Results ①A monoclonal antibody (clone L3D) which could specifically bind to human Tim-3 protein in ELISA assay was obtained and the subtype of the monoclonal antibody was IgG2a .②Flow cytometry indicated that the monoclonal antibody could bind to Tim-3 expressed in human U937 cells.This antibody also showed a cross activity to mice′Tim-3.③The monoclonal antibody inhibited the apoptosis of THP1 cells induced by Gal-9, the ligand of Tim-3.④Injection of Tim-3 antibody exacerbated sepsis in mice as marked by the decreased survival rate and increased expression of pro-inflammatory cytokines .Conclusion An anti-human Tim-3 monoclonal antibody is successfully obtained.The excellent binding and neutralization activities of this antibody enable it to be widely used in clinical diseases associated with deregulated Tim-3 expression .

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Enhanced erythromycin production in Saccharopolyspora erythraea by tandem expres-sion of metK, vhbS and adpA

Lun PAN ; Yongrong MAO ; Meng CHEN ; Panpan WU ; Li YUAN ; Xunduan HUANG ; Hang WU ; Zhongdong XU ; Buchang ZHANG

Military Medical Sciences.2014;(8):608-611,616. doi:10.7644/j.issn.1674-9960.2014.08.010

Objective To construct erythromycin-overproducing mutants by tandemly expressing S-adenosylmethionine synthetase gene metK, Vitreoscilla hemoglobin gene vhbS and pleiotropic regulatory gene adpA in Saccharopolyspora eryth-raea.Methods Through PEG-mediated protoplast transformation , the integrative plasmid carrying metK, vhbS and adpA was respectively introduced into erythromycin-producing wild-type strain S.erythraea A226 and industrial strain WB .The engineered strains were generated by apramycin resistance screening and PCR identification .The erythromycin production was compared in original strains and their mutants by the inhibition test of Bacillus subtilis and HPLC analysis .Results and Conclusion Four A226-derived mutants A226-P1-P4 and three WB-derived mutants WB-P1-P3 were independently obtained.Compared with wild-type strain A226, the relative erythromycin titer of the four engineered strains A 226-P1-P4 was increased from 8%to 25%by scoring the growth-inhibition zones .Further HPLC analysis showed that the four mutants had increased erythromycin A yield by 64%-94%.Likewise, the relative erythromycin titer and erythromycin A yield of the three engineered strains WB-P1-P3 were enhanced by 6%-10%and 31%-62%, respectively, in comparison with the original strain WB.The results show the universality of enhancing erythromycin productionvia tandem expression of metK, vhbS and adpA in S.erythraea.

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Effect of PPAR-δ on lung injury of rats induced by hyperbaric oxygen exposure

Fangfang WANG ; Yiqun FANG ; Xiaochen BAO ; Jun MA ; Pu YOU ; Shi ZHANG

Military Medical Sciences.2014;(8):591-593,601. doi:10.7644/j.issn.1674-9960.2014.08.006

Objective To investigate the effect of PPAR-δ on the lung injury of rats induced by hyperbaric oxygen (HBO2) exposure.Methods Sixty male Sprague-Dawley rats were randomly divided into six groups:air+vehicle, air+GW0742, and air+GSK0660, HBO2 +vehicle, HBO2 +GW0742, HBO2 +GSK0660.Lung injury was induced in rats by HBO2exposure (2.3 ATA, 100%O2, 8 h).Rats were injected with vehicle[10%DMSO in 0.3 ml NaCl 0.9%(v/v)] or GW0742 (0.3 mg/kg, ip) or GSK0660 (1 mg/kg, ip) at 1, 6 and 12 hours before either air or oxygen exposure .Protein levels in the bronchoalveolar lavage fluid ( BALF) , wet/dry ratio of the lung and the pathological changes in the lung tissue were detected 30 min after rats′egress.Results and Conclusion For the HBO2 +GW0742 group, the protein levels in BALF, the wet/dry ratio of the lung and the pathological changes in lung tissues all significantly decreased compared with those of the air group .These changes in HBO 2 +GSK0660 group tended to increase the level of lung injury .PPAR-δhas a protective effect on pulmonary oxygen toxicity induced by HBO 2 .

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Informationization of military medicine

Wei WANG ; Dongsheng ZHAO ; Junwen MAO

Military Medical Sciences.2015;(8):629-632,636. doi:10.7644/j.issn.1674-9960.2015.08.014

Military medicine has to restructure itself to meet military needs and e-Science patterns of the information age.This restructuration is known asinformationization of military medicine ( IMM).IMM is a process of vision refresh-ing into information thinking, mission expansion to the information field and paradigm shift to normal forms of information. The result is the emergence of military med-information technologies, military med-information research fields, and military med-informatics disciplines.

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Diagnostic value of plasma tissue factor pathway inhibitor in patients with severe acute pancreatitis

Rongjian CHEN ; Jingchun SONG ; Yongchun XU ; Hongliang ZHU ; Zili CHEN ; Hongjun QIAN ; Qingwei LIN

Military Medical Sciences.2015;(8):626-628,651. doi:10.7644/j.issn.1674-9960.2015.08.013

Objective To examine the levels of plasma tissue factor pathway inhibitor (TFPI) in patients with acute pancreatitis (AP) to assess the clinical value of diagnosis for severe acute pancreatitis ( SAP).Methods Sixty-eight patients were divided into mild acute pancreatitis (MAP)group (n=36) and SAP group (n=32), and twenty volunteers were chosen into normal group ( n=20 ) .Clinical data of these patients were collected, including APACHEⅡscore and Ranson score.Plasma levels of TFPI were measured by ELISA.Results The plasma levels of TFPI in SAP group, MAP group and control group were (4274.25 ±639.83),(3026.81 ±465.76) and (2468.73 ±262.39)pg/ml, respectively(P<0.05).There were significant positive correlations between TFPI and WBC, AST, ALT, TBIL, Cr, PT, APTT, PCT, APACHEⅡscore and Ranson score (P<0.05).The area under the curve (AUC) of TFPI for SAP was 0.902(95%CI=0.845 -0.959, P<0.05 ) .The cutoff value was 4028.83 pg/ml for plasma TFPI with a sensitivity of 87% and a specificity of 78%.Conclusion Plasma levels of TFPI in patients with SAP are significantly increased, which maybe help diagnose SAP.

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Key optimization of light source parameters of lidar system for bioaerosol detection

Zijian YANG ; Feng CHEN ; Chao LI ; Ming YU ; Taihu WU

Military Medical Sciences.2015;(8):577-581,586. doi:10.7644/j.issn.1674-9960.2015.08.002

Objective There are three categories of bioaerosol laser telemetry systems according to the light source configuration parameters:namely traditional lidar(light datection and ranging,laser radar), micro-pulse lidar and pseudo-random modulation(lidar).The system source parameters,which impact the degree of danger and detection sensitivity of the system, need to be optimized.Methods With reference to the USA laser product safety standards and by establishing the corresponding mathematical model of a lidar, the three categories of lidar source configuration were compared according to signal to noise ratio(SNR) and security before the repetition rate, pulse energy, divergence angle, distance and other dangerous impact factors were calculated.Results The results showed that to ensure eyesafety, the use of the pulse frequency should be set at 55 kHz for the highest SNR under the micro-pulse lidar excitation mode.Conclusion The eyesafety requirements impact the excitation of light source of a bioaerosol telemetry system.

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Role of resistin in insulin resistance in vascular endothelium cells by endoplasmic reticulum stress

Na ZHANG ; Weihong LI ; Yuhan LIN

Military Medical Sciences.2015;(8):607-609,625. doi:10.7644/j.issn.1674-9960.2015.08.008

Objective To identify the role of resistin in insulin resistance(IR) by endoplasmic reticulum(ER) stress in human umbilical vein endothelial cells ( HUVECs) and in rats.Methods HUVECs were cultured in vitro and disposed by resistin (R) or tauro ursodesoxycholic acid (tudca).Expressions of GRP78, P-akt and P-eNOS were determined using Western blotting.Thoracic aortic rings were made and their dilation function exposed to different concentrations of insulin was detected.Changes of vascular morphology were observed by HE staining.Results Results of Western blotting showed that expression of GRP78 was remarkably increased,but P-akt and P-eNOS were markedly decreased in R group.However, there was no difference in expressions of GRP78, P-akt and P-eNOS between tudca group and control group.The insulin induced vasodilation was decreased in R group and there was no difference between tudca group and control.Using HE staining, the R group showed significant medial thickening and proliferation of smooth muscle.Conclusion Resistin can induce insulin resistance in vascular endothelium cells by ER stress.

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Construction of a glycoengineering yeast overexpressing N-glycosyltransferase

Yinping MAO ; Xin GONG ; Shaohong CHANG ; Xiyong SONG ; Jun WU ; Bo LIU

Military Medical Sciences.2015;(8):602-606,617. doi:10.7644/j.issn.1674-9960.2015.08.007

Objective To obtain a strain of glycoengineering yeast with higher N-glycosylation efficiency by overexpressing N-glycosyltransferase.Methods Through the selecting marker URA3 gene, a new glycoengineering yeast strain named 4-32-STT3D was constructed, which could overexpress the Leishmania major N-glycosyltransferase staurosporine and temperature sensitivity3 D subunit(STT3D) under the control of an inducible alcohol oxidase 1(AOX1) promoter.We analyzed the N-glycosylation status of anti-human epidermal growth factor receptor 2 ( HER2 ) antibody and granulocyte macrophage colony stimulating factor (GM-CSF) expressed in 4-32-STT3D using SDS-PAGE,Western blotting and peptide-N-asparigineamidase F(PNGase F).Finally the effect of STT3D on the growth rate of glycoengineering yeast was detected.Results SDS-PAGE showed that anti-HER2 antibody expressed in 4-32-HL had two components:the first one with a relative molecular mass 55 ×103 was glycosylated,while the second one with 50 ×103 was non-glycosylated,but anti-HER2 antibody expressed in 4-32-HL-STT3D had the component of 55 ×103 only without any non-glycosylated 50 ×103 .The above components became 50 ×103 with the digestion of PNGaseF.All of them proved to be antibodies by Western blotting.As a report protein,GM-CSF expressed in 4-32-GM-CSF had two components: 22 ×103 and 20 ×103, while in 4-32-GM-CSF-STT3D there was only one with 22 ×103 .All these components became 18 ×103 with the digestion of PNGase F.Statistical analysis showed that without induction,STT3D had no effect on the growth rate of glycoengineering yeast, while great effect was observed when STT3D was induced.Conclusion Glycoengineering yeast with the overexpression of N-glycosyltransferase has higher N-glycosylation efficiency.

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ALKBH5 inhibits proliferation of HepG2 and L-02 cells

Ranran MENG ; Ying ZHANG ; Gangqiao ZHOU

Military Medical Sciences.2015;(8):593-596,601. doi:10.7644/j.issn.1674-9960.2015.08.005

Objective To investigate the effect of AlkB homologue 5 ( ALKBH5 ) on proliferation, cell cycle, and apoptosis of HepG2 and L-02 cells.Methods Recombinant plasmid vector containing the CDS region of ALKBH5 (pEGFP-C1b-ALKBH5) was stably transfected into HepG2 and L-02 cells.Western blotting was used to detect the expression of green fluonescence protein ( GFP )-ALKBH5.There were two groups in our experiment: GFP-ALKBH5 lentivirus group and GFP lentivirus group.Characteristics, such as proliferation, cell cycle and apoptosis of HepG2 and L-02,were detected through Cell Counting Kit-8 (CCK-8) assay, flow cytometry and clone formation, respectively.Results The result of Western blotting revealed that ALKBH5 was efficiently up-regulated at protein levels.Despite apoptosis, phenotypic analysis revealed that the proliferation and cell phases were significantly inhibited in ALKBH5 overexpressed stable cell strains compared with the control cells (both P<0.05).Conclusion ALKBH5 can restrain fetal liver cell (L-02) and hepatocellular carcinoma cell (HepG2) from proliferating.Taken together, our results strongly suggest that ALKBH5 can play a key role in the generation and progression in HCC as a tumor suppressor.

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