Objective To investigate the effects of intrathecal transplantation of bone marrow stromal cells (BMSCs) on inter cellular cell adhesion molecule-1 (ICAM-1) expression and blood spinal cord barrier following spinal cord ischemia reperfusion injury.Methods Ninety Sprague Dawley rats were randomly divided into three groups:sham (Sham group),ischemia-reperfusion injury (I/ R group),and BMSCs transplantation (BMSCs group).Spinal I/R injury was induced by clamping the aortic arch between left common carotid artery and left subclavian artery for 14 min in I/R group and BMSCs group.Sham group was subjected to exposure of aortic arch but without occlusion.I/R group and BMSCs group were intrathecally injected with phosphate buffered saline (PBS) or BMSCs (2 × 106) two days before injury.At 1 d,3 d,and 7 d after injury,neurological function was evaluated and damaged lumbosacral seg ment was removed for measurement of blood spinal cord barrier permeability and ICAM-1 protein expression.Results Compared with Sham group,neurological function score was significantly lower:1 d (F =38:59,P =0.001),3 d (F =31.34,P =0.001),and 7 d (F =27.71,P =0.001) ; ICAM-1 expression was increased 1 d (F =34.33,P =0.001),3 d (F =29.76,P =0.001),and 7 d (F =23.65,P =0.001),and blood spinal cord barrier permeability was higher:1 d (F =42.57,P =0.001),3 d (F =32.75,P =0.001),and 7 d (F =26.89,P =0.001) in I/R group.Compared with I/R group,neurological function score was increased:1 d (F =16.62,P =0.001),3 d (F =21.54,P =0.001),and 7 d (F =12.84,P =0.002) ; ICAM-1 expression was decreased:1 d (F =19.84,P =0.018),3 d (F =17.38,P =0.008),and 7 d (F =22.46,P =0.007),and blood spinal cord barrier permeability was lower:1 d (F =22.38,P =0.016),3 d (F =27.59,P =0.009),and 7 d (F =23.25,P =0.001) in BMSCs group.Conclusions Intrathecal transplantation of BMSCs inhibited ICAM-1 expression and decreased blood spinal cord barrier permeability,and then attenuated spinal cord ischemia-reperfusion injury.

Objective To investigate the expression of Notch1 and its ligand DLL4 in human gastric carcinoma tissues and its correlation with tumor angiogenic metastasis.Methods Immunohistochemical SP method was used to detect the expression of Notch1,DLL4 and VEGF in 45 gastric carcinoma tissues and paired adjacent normal gastric mucosa,and the relationship between them and clinico-pathological parameters were analyzed.Results The positive expression rate of Notch1,DLL4 and VEGF in gastric carcinoma were higher than that in normal gastric mucosa(P

ObjectiveTo evaluate the effect of low glycemic index(LGI)diet and exercise on plasma glucose and lipid profiles in newly diagnosed type 2 diabetic patients. MethodsSeventeen newly diagnosed type 2 diabetic patients with FPG ≤ 10mml/L treated by LGI diet and exercise only for two months.Fasting plasma glucose (FPG),2 hours postprandial glucose(2hPG),glycosylated hemoglobin A1 C(GHbA1C),and lipid profiles were measured.The results of FPG,2hPG,GHbA1C,and lipid profiles were compared. ResultsTwo months after treatment,the level of fasting glucose(6.19 ± 0.60)mmol/L,postprandial 2h plasma glucose(8.59 ± 0.90)mmol/L,TG(1.15 ± 0.45)mmol/L,TC(4.98 ± 0.77)mmol/L,LDL(3.20 ± 0.71)mmol/L were significantly lower than (7.84 ± 1.19)mmol/L,(13.97 ± 3.35)mmol/L,TG(1.79 ± 0.75)mmol/L,TC(5.46 ± 0.27)mmol/L,LDL (3.57 ± 0.28)mmol/L,HDL(1.59 ± 0.30)mmol/L was significantly higher than(1.42 ± 0.26)mmol/L,the differences were statistically significant(all P＜0.05);HbA1c(6.49 ± 0.57)% was slightly lower than(7.29 ±0.77)%,but the difference was not significant(P＞0.05);No hypoglycemia was observed during the treatment. ConclusionThe exellent glycemic control and improvement of lipid profile could be achieved by low glycemic index diet and exercise only.Furthermore,no hypoglycemia occurred during the treatment.

Objective To investigate the effect of glucose-based peritoneal dialysis fluids and icodextrin-based peritoneal dial-ysis fluids on the expression of TLR2 and TLR4 on huamn peritoneal mesothelial cells .Methods Human peritoneal mesothelial cell line 5 - 10 generations(HMrSV5) was cultured in DMEM /F12 medium supplemented with 10% (v/v) fetal calf serum (FCS) .Cell viability and cell proliferation were assessed using M TT method .The experiment were divided into 5 different groups :group A (control group) ,1 .5% dextrose group ,2 .5% dextrose group ,4 .25% dextrose group and 7 .5% Lcodextrin group .Icodextrin group (aikau dextrin) ,TLR2 and TLR4 expression were detected by Western blot .Results Treatment with different concentrations of glucose-based peritoneal dialysis fluids for 24 h did not affect the expression of TLR2 and TLR4 protein .In addition ,after stimula-tion for 48 h ,1 .5% dextrose ,2 .5% dextrose ,4 .25% dextrose decreased TLR2 expression by (5 .5 ± 2 .8)% ,(31 .4 ± 7 .5)% , (54 .9 ± 1 .9)% respectively ,TLR4 expression by (32 .9 ± 17 .6)% ,(47 .7 ± 13 .5)% ,(66 .4 ± 13 .5)% respectively .Stimulation for 72 h ,decreased TLR2 expression by (29 .4 ± 14 .7)% ,(38 .9 ± 9 .9)% ,(63 .5 ± 16 .5)% respectively ,TLR4 expression by(59 .5 ± 16 .8)% ,(63 .1 ± 9 .5)% ,(79 .2 ± 14 .0)% respectively .There was no significant change in TLR2 and TLR4 protein expression on 7 .5% icodextrin group .Conclusion Glucose-based peritoneal dialysis fluids ,but not icodextrin-based peritoneal dialysis fluids downregulates expression of TLR2 and TLR4 by HM rSV5 .

Objective To investigate the protective effect of hypoxic-preconditioned bone marrow mesenchymal stem cells (BMSCs) on spinal cord tissue after ischemia reperfusion injury.Methods Healthy adult Sprague Dawley (SD) rats weighing 200 ～ 250 grams (g) were randomly divided into 3 groups with 6 animals in each group:The sham group received simple surgical manipulation without ischemia/reperfusion treatment;The spinal cord ischemia/reperfusion group (Control group) only received spinal cord ischemia/reperfusion surgery.The hypoxic preconditioned BMSC transplantation group (HP-MSCs group) was injected with hypoxic preconditioned BMSCs 2 days before ischemia/reperfusion.The control group,HP-MSCs group received spinal cord ischemia/reperfusion for 10 min and observed for 48 h.The permeability of the blood-spinal cord barrier was examined with Evans blue (EB),and the histomorphology changes were observed with hematoxylin and eosin (HE) staining.Results EB red fluorescence was significantly weakened in the HP-MSCs group than that in the Control group (P ＜ 0.05),and more intact motor neurons were found in the lumbar spinal cords in the HP-MSCs group than that in the Control group (P ＜0.05).Conclusions The hypoxic-preconditioned BMSCs could effectively attenuate spinal cord ischemia reperfusion injury,it may be associated with protective effect of the blood-spinal cord barrier integrity.

OBJECTIVE:To establish the method for determining 10 residual organic solvents in norvancomycin hydrochloride raw material. METHODS:Headspace gas chromatography was performed on the column of nitro modified polyethylene terephthal-ate glycol as stationary phase capillary column;the oven temperature program started at 40 ℃ for 3 min and increased at a rate of 8 ℃/min up to 150 ℃ for 10 min;the temperature was 200 ℃ with carrier gas of high-purity nitrogen gas,the constant flow rate was 5 ml/min with split ratio of 15∶1;the headspace vial equilibrium temperature was 85 ℃ with equilibrium time of 40 min,and the volume was 1 ml. RESULTS:The concentration of n-pentane,acetone,ethanol,benzene,acrylonitrile,toluene,xylene,chlo-robenzene,styrene,divinylbenzene had good linear relationship with its peak area values(r=0.995 7-0.999 9);the RSDs of preci-sion,repeatability tests was ≤6.6%;average recovery was in the range of 94.3%-106.6%(RSD=0.5%-4.5%,n=9). CONCLU-SIONS:The method is fast,sensitive and accurate,and can be used for the determination of residual organic solvents in norvanco-mycin hydrochloride raw material.

Objective To investigate the effects of allele-31 C>T on the binding activity to IL-1βpromoter of the nuclear transcription factor C/EBPβand PU.1 induced by Mycobacterium tuberculosis infection.Methods The electrophoretic mobility shift assay ( EMSA) was performed to explore whether the nuclear transcription factor C/EBPβand PU.1 could bind to -31 region in IL-1βpromoter.The C/EBPβ-and PU.1-expressing vectors were constructed and co-transfected into HeLa cells with IL-1βpromoter luciferase vector.The expression of C/EBPβand PU.1 was confirmed using Western blotting assay, and the promoter activity was determined using Dual-Glo Luciferase system under various transfection conditions. Lentivirus-mediated RNA interference was used to explore the effects of C/EBPβand PU.1 on IL-1βexpression.GraphPad Prism 5.0 was used for data analysis.Results EMSA results showed that both C/EBPβand PU.1 could bind to -31 region in IL-1βpromoter.Both C/EBPβand PU.1 induced by Mycobacterium tuberculosis infection could increase IL-1βpromoter activity, especially for the -31 T allele (t=22.33 and 7.98,P<0.01), and there was a synergy on the promoter activity between C/EBPβand PU.1.The promoter activity decreased significantly when C/EBPβand/or PU.1 were silenced by lentivirus-mediated RNA interference (q=5.79, 6.23 and 11.66,P<0.01).Conclusion The allele-31 C>T can induce IL-1βpromoter activity and gene transcription through regulation of binding activity to C/EBPβand PU.1 induced by Mycobacterium tuberculosis infection.

Abstract: Objective To construct a shuttle vector pHT315-AaCPR100A with two spore-producing-dependent promoters and the target gene AaCPR100A in Escherichia coli-Bacillus thuringiensis. Methods The forward promoter of Cry3A, named Pro-1 (+), was amplified by PCR using pSVP27A plasmid as the template, and the target gene AaCPR100A was amplified using Aedes aegypti RNA reverse conversion cDNA as the template. The plasmid pHT315 was linearized by digestion with Hind Ⅲ and Sal Ⅰ. The forward promoter and the target gene were inserted into the linearized vector pHT315 successively by in-fusion cloning according to the transcription direction. The synthesized plasmid containing the Cry3A reverse promoter sequence was used as the template, and the Pro-1 (-) reverse promoter was amplified by PCR. The intermediate vector containing the forward promoter and the target gene was linearized by EcoR I restriction enzyme, and the reverse promoter was inserted downstream of the target gene by in-fusion cloning in the direction of transcription. Results By agarose gel electrophoresis, the forward promoter, target gene AaCPR100A and reverse promoter bands were clear and of good quality, which could be used for in-fusion cloning experiments. The two spore-producing-dependent promoters and target gene fragments were connected by In-fusion cloning. The recombinant vector pHT315-AaCPR100A was verified by PCR. The forward promoter, target gene fragment and reverse promoter were successfully amplified in the recombinant vector. Nucleotide sequencing verified that the sequencing results of the bidirectional promoter sequence and the target gene sequence were basically consistent with the sequence alignment results, which met the requirements of the construction of vector elements and proved that the recombinant vector was successfully constructed. Conclusions Based on the above results, this study proves that the recombinant shuttle vector with two spore-producing-dependent promoters can be successfully constructed by in-fusion cloning technology, laying the foundation for the construction of engineered Bacillus thuringiensis expressing dsRNA of AaCPR100A.

Preparation of maternal strain A/PR/8/34 HA antiserum for influenza virus classical reassortment. A/PR/8/34 virus was digested by bromelain after inactivation and purification. 5%-20% sucrose continuous density gradient centrifugation method was used to purify HA protein. SIRD method was used to select the target protein. SDS-PAGE method was used to identified HA protein. High Immunogenic A/PR/8/34 HA protein was successfully prepared and HI titer reached 10240. High purity HA antiserum was identified by SIRD method. The key reagent in the classical reassortment of influenza virus was prepared, and the complete set of technical methods were explored, which laid the foundation for the independent research and development of seasonal influenza vaccine strains of China.
Animals ; Antibodies, Viral ; immunology ; Electrophoresis, Polyacrylamide Gel ; Female ; Hemagglutination Inhibition Tests ; Hemagglutinin Glycoproteins, Influenza Virus ; analysis ; immunology ; Humans ; Influenza A Virus, H1N1 Subtype ; genetics ; immunology ; Influenza, Human ; immunology ; virology ; Rabbits ; Reassortant Viruses ; genetics ; immunology

Objective To evaluate the effect of hypoxia-preconditioned bone marrow mesenchymal stem cells (BMSCs) on the expression of HO-1 during spinal ischemia-reperfusion (I/R) in rats.Methods Thirty healthy adult Sprague-Dawley rats,weighing 200-250 g,were randomly divided into 5 groups with 6 animals in each group using a random number table:sham operation group (group S),I/R group,PBS group,BMSC transplantation group (BMSC group) and hypoxic preconditioning group (group HP).In group HP,BMSCs were exposed to 3％O2-5％CO2-92％N2 for 24 h for hypoxia preconditioning.In PBS,BMSC and HP groups,PBS,BMSCs and BMSCs for hypoxic preconditioning were injected intrathecally,respectively,and 60 min later spinal I/R was induced by clamping the aorta in the rats anesthetized with chloral hydrate.At 6,12,24 and 48 h and 7 days of reperfusion,the neurological function was evaluated and scored.At day 7 of reperfusion,the rats were sacrificed,and spinal cord tissues were obtained for determination of HO-1 expression (by Western blot) and HO-1 mRNA expression (using real-time PCR).Results Compared with group S,the neurological function score was significantly decreased at each time point of reperfusion,and the expression of HO-1 mRNA and protein in spinal cord tissues was up-regulated in the other groups.Compared with group I/R,the neurological function score was significantly increased at each time point of reperfusion,and the expression of HO-1 mRNA and protein in spinal cord tissues was upregulated in BMSC and HP groups,and no significant change was found in group PBS in the parameters mentioned above.Compared with group BMSC,the neurological function score was significantly increased at each time point of reperfusion,and the expression of HO-1 mRNA and protein in spinal cord tissues was up-regulated in group HP.Conclusion The mechanism by which hypoxic preconditioning enhances protection of spinal cord by BMSC transplantation may be related to up-regulation of HO-1 expression in rats.