Biocontrol bacteria 308R(pKSH) with hrpN of Erwinia amyloyora. It can secrete the Harpin protein which can induce plant resistance. After successive growth in antibiotic-free LB medium for 50 generations, only 23.1% of the cell still retained the plasmid pKSH, in comparison with 4.75% of the cosmid pCPP430. Sprayed onto the leaves of tomato, the recombinant strain maintained a population density of over 10~ 5 cfu/cm~ 2 when kept under high humidity in thirteen days, but if without humidity, the strain keep 10~ 4 cfu/cm~ 2 over in five days. On the tomato leaves, the stability of the recombinant strain 308R(pKSH) was higher than the strain 308R(pCPP430). The experiment indicated that the recombinant strain 308R(pKSH) had higher stability compared with the strain 308R(pCPP430), but that was not enough. In this paper reasons for unstability and strategies for improving were discussed for the recombinant strain.

Objective To investigate the distribution and antimicrobial susceptibility of the bacteria isolated from bloodstream infections during 2013-2014 in Changhai Hospital for rational use of antibacterial agents.Methods The bacterial strains from blood samples were collected during the period from January 2013 through December 2014,and subjected to antimicrobial susceptibility testing by using automated system or Kirby-Bauer method.The results were interpreted according to CLSI M100-S24 breakpoints or FDA breakpoints.The data were analyzed by WHONET 5.6 software.Results A total of 1 048 nonduplicate isolates were collected,of which Escherichia coli,coagulase-negative Staphylococcus (CNS) and Klebsiella pneumoniae accounted for 29.5％,15.8％ and 13.8％,respectively.Gastroenterology,Hematology,General surgery,Urology and Department of Infectious Diseases are the top 5 departments according to their total number of bacterial isolates.The results of antimicrobial susceptibility testing showed that ESBLs-producing E.coli and K.pneumoniae accounted for 63.8％ and 38.6％,respectively.The prevalence of methicillinresistant CNS (MRCNS) was 77.6％.The E.coli strains isolated from Urology showed higher resistance rates to cephalosporins than the total E.coli strains,while the E.coli strains isolated from Gastroenterology showed higher resistance rates to betalactarn/beta-lactamase inhibitor combinations and carbapenems than the total E.coli strains.Higher prevalence of MRCNS was found in departments of Hematology,Urology and Neurosurgery.All the CNS strains isolated from Neurosurgery were resistant to methicillin.The K.pneumoniae strains isolated from Bum ICU had higher resistance rates to all the antibacterial agents tested than the total K.pneumoniae strains,while the K.pneumoniae strains isolated from Gastroenterology showed higher resistance rates to carbapenems and tigecycline than the total K.pneumoniae strains.Conclusions The pathogenic bacteria isolated from bloodstream infections vary with departments in terms of species distribution and antimicrobial susceptibility profile.It is necessary to strengthen the surveillance of antimicrobial resistance in hospital for rational use of antibiotics.

In order to study the analgesic effect of Shaoyao Gancao Decoction, this paper discussed material basis and mechanism from the perspective of macromolecules in traditional Chinese medicine. Inspired by the phenomenon of turbidity after boiling Chinese medicine, this experiment took Shaoyao Gancao Decoction as the research object to study the formation process of precipitation during boiling. The results showed that aggregates with a certain shape were formed in the solvent during the boiling process, and the precipitate was obtained by standing and centrifuging. Analysis found that the precipitation was mainly composed of small molecules such as paeoniflorin, albiflorin, liquiritin, glycyrrhizic acid, isoliquiritin and gallic acid, and macromolecules such as protein and polysaccharide. The composition of precipitate was consistent with that of Shaoyao Gancao Decoction, but the analgesic effect of Shaoyao Gancao Decoction after removing the precipitate was significantly reduced. Based on these results, we isolated small molecular compounds, polysaccharides and protein from Shaoyao Gancao Decoction and their contents are 60.4, 700.7 and 207.2 mg·g^-1 respectively. We get the ratio, polysaccharide: small molecule = 11.6∶1, protein: small molecule = 3.4∶1, the precipitate is prepared in the state of boiling. The characterization results showed that the particle size of the precipitate will change significantly after co-heating, and the content determination results showed that the content of the six small molecular compounds which was free in solvent was significantly reduced after the formation of the precipitate. The acetic acid writhing experiment proved that the precipitate has a good analgesic effect, and effectively reduced the levels of inflammatory factors prostaglandin E2 and nitric oxide, and increased the level of anti-inflammatory factor interleukin-10. These results proved that the precipitate in Shaoyao Gancao Decoction is an important material basis for analgesic effect, and macromolecules such as protein and polysaccharide are the main components of the precipitate. The study of macromolecules in the precipitate of Shaoyao Gancao Decoction not only provides new ideas and methods for elucidating the pharmacodynamic material basis of Shaoyao Gancao Decoction, but also provides a reference for analyzing the scientificity of traditional decoction.

Objective To clone and express the encoding sequence of glyceraldehydes-3-phosphate dehydrogenase(GAPDH)from periodic Brugia molayi(Bm).Methods Total RNA was extraeted from periodic Brugic malayi.The BmGAPDH gene was amplified by RT-PCR.The PCR product was cloned and then subeloned into pcDNA3.1(+)vector.The recombinant plasmids were screened and identified by digestion with restriction enzyme and PCR amplification,and were transformed into COS-7 cell subsequently.The expressed protein was identified by SDS-PAGE.Results BmGAPDH mRNA was highiy expressed in transfected COS-7 cell.The deduced amino acid sequence was identical with that of BmGAPDH.The recombinant pnotein wag about Nr 43 000.Conclusion The recombinant plasmid peDNA3.1(+)-BmGAPDH has been constructed and the protein has been expressed correctly.

Purpose To explore the immunohistochemical ( IHC) expression of ALK antibodies with different clones in anaplastic lym-phoma kinase ( ALK) gene fusion non-small cell lung cancer ( NSCLC) . Methods ALK expression in 60 NSCLCs were detected by IHC including autostainer (D5F3, Ventana+BenchMark) and manual staining using 4 different antibodies of D5F3 (Ventana), D5F3 (Cell Signaling), 1A4/1H7 (OriGene), 5A4 (Abcam), and all cases were verified with ALK FISH. Their expressions with 4 anti-bodies were compared with those by D5F3 (Ventana+BenchMark). Results 32 ALK gene rearrangement NSCLCs and 28 negative cases were identified by FISH and D5F3 (Ventana+BenchMark). The sensitivity of D5F3 (Ventana), D5F3 (Cell Signaling), 1A4/1H7 (OriGene), 5A4 (Abcam) was 93. 8%, 84. 4%, 93. 8%, 56. 3%, and all the speciticity was 100%. The consistency with D5F3 (Ventana+BenchMark) was 96. 7%, 91. 7%, 96. 7% and 76. 7%, respectively. The validity of immunohistochemical staining in surgical resection specimens was better than in small biopsies. Conclusion Effective routine manual immunohistochemistry with high-affinity antibody clone may provide a more economic and widespread pre-screening technique.

To study the potential molecular mechanism of tumor angiogenesis in its microenvironment, we investigated the effects of HepG2 conditioned medium on the proliferation of vascular endothelial cell and vascular angiogenesis in our laboratory. Human umbilical vein endothelial EA. hy926 cells were co-cultured with HepG2 conditioned medium in vitro. The proliferation and the tubulogenesis of EA. hy926 cells were detected by teramethylazo salt azole (MTT) and tube formation assay, respectively. The results showed that the survival rate of the EA. hy926 cells was significantly increased under the co-culture condition. HepG2 conditioned medium also enhanced the angiogenesis ability of EA. hy926 cells. In addition, the expressions of intracellular VEGF and extracellular VEGFR (Flk-1) were regulated upward in a time-dependent manner. In conclusion, the proliferation of vascular endothelial cells and Vascula angiogenesis were improved under the condition of indirect co-culture.
Carcinoma, Hepatocellular ; pathology ; Cell Proliferation ; Coculture Techniques ; Culture Media, Conditioned ; Endothelial Cells ; cytology ; Hep G2 Cells ; Human Umbilical Vein Endothelial Cells ; Humans ; Liver Neoplasms ; pathology ; Neovascularization, Pathologic ; Tumor Microenvironment ; Vascular Endothelial Growth Factor A ; metabolism ; Vascular Endothelial Growth Factor Receptor-2 ; metabolism

Objective:To explore the effect on sex hormone functional of extracts of Herba Epimedii /Fructus Ligustri Lucidi in osteoporosis rats.Methods:50 cases of 3 months Wistar rats were rando mly divided into normal group ,model group,extract low dose group(extract-L),extract middle dose group(extract-M),extract high dose group(extract-H),10 cases in each group,in addition to the normal group,other 4 groups were bulided OP model by filling and feeding retinoic acid for 15 d,then normal group and model group were filling physiological saline;extract-L,extract-M,extract-H were respectively feeding extracts of Herba Epimedii /Fructus Ligustri Lucidi at 50 mg/kg,100 mg/kg,200 mg/kg,both for 3 weeks.Measured the bone mineral density (BMD),observed the femoral pathology change of left femur with hematoxylin and eosin ( HE ) staining, measured trabecular bone tissue morphology related parameters ,analyzed the hormone receptor protein expression with immunohistochemistry .Results:The BMD of the whole femur ,femur head,and femur neck in normal group were all significantly higher than that of model group ,the BMD of the whole femur ,femur head, and femur neck in extract-M and extract-H group were all significantly higher than that of model group ,the BMD of the whole femur in extract-L group was significantly higher than that of model group ,the differences were statistically significant (P<0.05 or P<0.01).The bone trabecular number had significantly increased ,integrity,continuity improved significantly ,bone histomorphometry such as Tb.Ar, Tb.Th,and Tb.N in model group were decreased ,and Tb.Sp was increased,extract-M and extract-H group had remarkable effect on each index ,while extract-L group only had a remarkable effect on Tb .Ar and Tb.Th ( P<0.05 or P<0.01 ) , compared with model group.The expression of AR , ER in normal groups were significantly higher than that of model group , the expression of AR , ER in extract-H group was significantly higher than that of model group , the differences were statistically significant ( P<0.05 or P<0.01 ) . Conclusion:This study demonstrated that the combination with TFE and TIFL had potential protective effects on osteoporosis rats induced by retinoic acid .The combination with TFE and TIFL is able to increase BMD ,reverse pathological changes of bone tissue ,co-ordinate hormones of the hypothalamic-pituitary-gonadal axis ,and upregulate the expression of sex hormone receptors .

Objective To compare the accuracy rate of talar osteochondral lesions between the small field of view (FOV) surface coil MR imaging and 2D FSE proton density weighted imaging (2D-FSE-PD),3D-SPACE sequences MR imaging with boot-shaped coil.Methods Totally 43 patients (45 ankles) underwent 3D-SPACE sequence after 2D-FSE-PD sequence scanning using the boot-shaped coil,then the PDWI sequence was scanning after the boot-shaped coil was replaced by the small FOV surface coil at 1.5T MRI.The results of the arthroscopic examinations was regarded as the standard for assessing the accuracy rate of the three methods.Results The accuracy rate of 2D-FSE-PD sequence with small FOV surface coil was 86.67％ (39/45),the accuracy rate of 2D-FSE-PD sequence with boot coil was 60.00％ (27/45),the accuracy rate of 3D-SPACE sequence with boot coil was 68.89％ (31/45).The accuracy rate of small FOV surface coil was higher than those of boot coil used 3D-SPACE sequence (x2 =4.114,P =0.002) and boot coil used 2D-FSE-PD sequence (x2 =8.182,P＜0.001).There was not significant difference between 3D-SPACE sequence using boot coil and 2D-FSE-PD sequence using boot coil (x2 =0.776,P=0.125).Conclusion Compared with the 2D-FSE-PD and 3D-SPACE sequences with boot-shaped coil,the small surface coil scanning with routine sequence can show talar osteochondral lesions better.

Objective:To analyze the change of bone mineral density (BMD) in postmenopausal women,and to study the relationship between BMD and the estrogen,cytokines,and serum levels of bone metabolism.Methods: 135 postmenopausal women were divided into the OP group (n=54),Osteopenia group (n=43) and normal group (n=38) according to the results of lumbar spine bone mineral density (BMD) scan.To detected the concentration of serum estradiol (E2),bone alkaline phosphatase (BALP) and osteocalcin (BGP),tartrate resistant acid phosphatase-5b (TRAP-5b),interleukin-6 (IL-6),tumor necrosis factor alpha (TNF-α),transforming growth factor beta (TGF-β1) and IL-10 of research objects in the three groups.And to analyze the relationship between BMD and the levels of E2,BALP,BGP,TRAP-5b,IL-6,TNF-α,TGF-β1 and IL-10.Results: In the OP group and the Osteopenia group,BMD and the levels of E2,TGF-β1 and IL-10 were obviously lower than that in the normal group (P<0.05),and the levels of BALP,BGP,TRAP-5b,IL-6 and TNF-α were obviously higher than that in the normal group (P<0.05).When compared with the Osteopenia group,the BMD and E2 level in the OP group was more lower (P<0.05),and the levels of BALP,BGP,TRAP-5b,IL-6 and TNF-α were more higher (P<0.05).E2 and TGF-β1 showed the significant positive correction with BMD (P<0.05),and the BALP,BGP,TRAP-5b,IL-6 and TNF-α were all negatively corrected with BMD (P<0.05).However,no significant difference of relevance was seen between IL-10 and BMD (P>0.05).The low serum E2 levels and high serum levels of BALP,BGP,TRAP-5b,IL-6 and TNF-α were the independent factors affecting the reduction of BMD (P<0.05).Conclusion: The E2 level in patients with PMOP decreased obviously,which caused the imbalance of bone remodeling and inflammatory reaction,and which was related to the decrease of BMD.

Objective To study the effects of butylphthalide on cognitive function,apoptosis and pp38MAPK in hippocampus of rat model of vascular dementia.Methods The vascular dementia (VD) model was established by two vascular (2VO) method,and then sixty male Wistar rats were randomly divided into VD group,sham operation group and NBP (butylphthalide) group.Rats in NBP group were given 120 mg · kg-1 · day-1 dose butylphthalide by gavage,and rats in VD group and sham operation group were given the same dose vegetable oil.The cognitive function of each rat was tested by Morris water maze.The expression of p-p38MAPK in the hippocampus was observed by Western blot;and the apoptosis was observed in hippocampal CAl region by TUNEL staining.Results The hidden platform escape latency of NBP group ((48.72 ± 7.01) s,(42.41 ± 4.06) s,(40.34 ± 2.46) s)was significantly shortened compared with those of VD group((82.71±8.27) s,(80.36±9.65) s,(77.74±6.33) s)(P＜ 0.01) ; and the former platform quadrant time and the number of passing through the platform of NBP group ((26.45±4.66)s,(1.84±0.82) times) were significantly prolonged (P＜0.01) compared with those of VD group ((18.67±5.39) s,(1.32±0.61) times);the apoptosis and the expression of p38MAPK phosphorylation in hippocampus in NBP group ((153.65±9.85),(0.42±0.04)) significantly reduced (P＜0.01) compared with those of VD group ((209.46±11.49),(0.88±0.10)).Conclusion Butylphthalide can improve the learning and memory ability of VD rats,the reduce apoptosis in the hippocampus by the inhibition of the P38MAPK pathway.This may be one of the ways by which butylphthalide can treat vascular dementia.