BACKGROUND: Troponin I (Tn I ) could inhibit the growth of vascular endothelial cells, inhibit neovascularization,through which to inhibit the development and metastasis of solid tumor. Similar to Tn I, TnC also exists in non-muscular tissue, but does it has the analogous activity of anticancer like Tn I ?OBJECTIVE: To explore the effect of recombinant human TnC (rhTnC) on the growth of human umbilical vein endothelial cells (HUV-EC) and mouse xenograft tumor.DESIGN: Controlled observation in vivo and in vitro.SETTING: Research Institute of Medicine, Chongqing K.E.W Pharmaceutical Co., Ltd. and Department of Biochemistry of Chongqing Medical University.MATERIALS: The experiment was conducted in the Biochemical Laboratory of Research Institute of Medicine,Chongqing K.E.W Pharmaceutical Co., Ltd. from March 2003 to December 2004. 100 Kunming mice either male or female of 15-22 g purchased from Chongqing Academy of Chinese Materia Medica. E.coli BL21 (DE3)pLysS/pET3b-TnC provided by Chongqing K.E.W Pharmaceutical Co., Ltd. HUV-EC (Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences).METHODS: Human TnC cDNA was obtained from human thymus cDNA library using PCR. The colony was cloned in E.coli and a bacterial strain of gene engineering E.coli BL21 (DE3) pLysS/pET3b-TnC was obtained, which could express hTnC. The recombinant human TnC (rhTnC) was purified with affinity chromatography of Ni-NTA agarose. ①In vitro cell experiment: HUV-ECs were seeded in the 96-well plates at density of 2×103 cells per well and co-cultured with rhTnC of 1, 5, 10, and 50 mg/L for 3 days. The absorption (A value) was detected with microplate reader at 540 nm and the inhibition rate of cell growth was calculated. Meanwhile, the 50% inhibiting dose (IC50 value) was assayed by LOGIT method. ②In vivo animal experiment: Ascites tumor (S-180) that had been inoculated for 7-8 days was harvested. The tumor cells were diluted to 1 ×1010 L-1 and 0.2 rnL was subaxillarily and intraperitoneally injected into each mouse (50 mice in each group). The next day, the mice were randomly divided into 5 groups: rhTnC 20 mg/kg group, rhTnC 10 mg/kg group, rhTnC 5 mg/kg group, Cyclophosphamide (Cy) group and control group with 10 mice in each group. The rhTnC 20,10 and 5 mg/kg groups were given administration at the corresponding doses, once a day for 7 days; 50 mg/kg Cy was given the Cy group one after an interval of day, and the same volume normal saline was given to the control group. One day after the last time of administration, all mice were killed and the tumor was harvested and weighed. The inhibition rate of tumor growth was calculated: tumor inhibition rate=[(Average weight of tumor in control group-Average weight of tumor in drug group)/Average weight of control group]×100%.MAIN OUTCOME MEASURES: Inhibition rate of rhTnC to HUV-EC proliferation in cell experiment in vitro and mouse xenograft tumor in animal experiment in vivo.RESULTS: ①In vitro cell culture showed that rhTnC suppressed HUV-EC proliferation in a dose-dependent manner (IC50=7.5 mg/L). ②Similar to the result of in vitro cell experiment, after intraperitoneal administration, the inhibition rate of rhTnC 5, 10, and 20 mg/kg groups was higher than that of control group (P ＜ 0.01); after subaxillary administration, the inhibition rate of rhTnC 5, 10, and 20 mg/kg groups was also higher than that of control group (P ＜ 0.05-0.01). There was no significant difference in the inhibition rate between two administration approaches (P ＞ 0.05).CONCLUSION: rhTnC is capable of inhibiting the proliferation of HUV-EC dose-dependently, and displays the activities of inhibiting the proliferation of HUV-EC and anti-tumor.

Objective To investigate the role of nuclear factor ?B activation in the onset of acute coronary syndrome(ACS).Methods 76 patients with acute myocardial infraction(AMI),41 patients with unstable angina pectoris(UAP),43 patients with stable angina pectoris(SAP)and 20 normal controls were enrolled.NF-?B activation in monocytes in peripheral blood monocyte was determined by ELISA with the NF-?B p65 Kit the at 3 and 5 days after admission.Results The activity of NF-?B in monocytes of peripheral blood in AMI patients and UAP patients was significantly higher than that in SAP patients and normal controls(P

BACKGROUND: It demonstrated that combined application of thymosin α 1 (TM-α1) and interferon (IFN) can enhance the antiviral activity of IFN.OBJECTIVE: To obtain recombinant fusion protein of TM- α1 and consensus IFN α (IFN α -con) with double activity of antiviral activity and immunity enhancement.DESIGN, TIME AND SETTING: The in vitro contrast experiment was conducted in the Biochemical Laboratory of Research Institute of Medicine, Chongqing K.E.W Pharmaceutical Co., Ltd. from March 2003 to December 2004.MATERIALS: The fusion gene fragment (TM- α 1+IFN α -con) was synthesized by Shanghai Sangon Biological Engineering Technology And Service Co., Ltd, WISH cell, and vesicular stomatitis virus (VSV) was purchased from Institute of Biochemistry and Cell Biology, commercial products of IFN α 1b, IFN α 2a and TM- α 1 was served as reference substance.METHODS: The preference for E. coli of fusion sequence coding TM- α 1 and IFN α -con were cloned into expression vector of pET-22b(+) and expressed in BL21(DE3)-Codon plus-RP-X, which was purified by precipitation of (NH4)2SO4, hydrophobic interaction chromatography, anion-exchange chromatography, cation-exchange chromatography and molecular exclusion chromatography. The antiviral activity of fusion protein was tested by cytopathic-effect inhibition assay, and effect of fusion protein on lymphocyte proliferation was tested by cell proliferative assay.MAIN OUTCOME MEASURES: The specific activity of fusion protein and its biological activity in promoting lymphocyte proliferation.RESULTS: The fusion protein was expressed as a soluble form, accounting for over 20% of the total cell protein in E. coli, which approached 96% after purification. The antiviral activity of fusion protein was superior to IFN α 1b and IFN α 2a. However, the activity of fusion protein for promoting lymphocyte proliferation was similar to TM- α 1.CONCLUSION: The fusion protein of TM- α 1 and IFN α -con expressed in E. coli has both effects on anti-virus and promoting lymphocyte proliferation.

To develop a safe and efficient recombinant subunit vaccine to foot-and-mouth disease virus(FMDV)type Asia 1 in sheep,a tandem repeated multiple-epitope gene consisting of residues 137-160 and 197-211 of the VP1 gene of FMDV was designed and artificially synthesized.The biologically functional molecule,the ovine IgG heavy constant region(oIgG)as a protein carrier was introduced for design of the multiple-epitope recombinant vaccine and recombinant expression plasmids pET-30a-RE and pET-30a-RE-oIgG were successfully constructed.The recombinant proteins,RE and RE-oIgG,were expressed as a formation of inclusion bodies in E.coli.The immune potential of this vaccine regime in guinea pigs and sheep was evaluated.The results showed that IgG could significantly enhance the immune potential of antigenic epitopes.The recombinant protein RE-oIgG could not only elicit the high levels of neutralizing antibodies and lymphocytes proliferation responses in the vaccinated guinea pigs,but confer complete protection in guinea pigs against virus challenge.Although the recombinant protein RE could not confer protection in the vaccinated animals,it could delay the appearance of the clinical signs and reduce the severity of disease.Inspiringly,the titers of anti-FMDV neutralizing antibodies elicited in sheep vaccinated with RE-oIgG was significantly higher than that for the RE vaccination.Therefore,we speculated that this vaccine formulation may be a promising strategy for designing a novel vaccine against FMDV in the future.

AIM: To observe and study the influence of cardiac sympathetic nerve on QT dispersion (QTd) and the circadian variations of QTd in experimental rabbits. METHODS: The rabbits were divided into experimental group (without cardiac sympathetic control by operation) and control group (with retained cardiac sympathetic control by operation, sham operation). QTd of both groups were measured and compared before and after the operation. The circadian variations of QTd were also observed in both groups. RESULTS: QTd in experimental group decreased significantly after the cardiac sympathetic nerves were excised (P

Objective To analyze the status of human metapneumovirus in children with respiratory tract infection in Shenz-hen and Shantou and the clinical characteristics of the diseases with hMPV infection.Methods The viral nucleic acid of 1 217 samples were amplified by RT-PCR(1 137 from children with respiratory tract infection,80 from health children),and five positive PCR products were chosen randomly for sequencing.Nucleotide sequences alignment and phylogenetic analysis were performed with DNAstar software.The clinical data of the children with hMPV infection were analyzed.Results The positive rate of hMPV infection identified from 1 137 children wtith respiratory tract infection was 4.49% (51/1 137), hMPV was significantly prevalent in February,March and April.Most of the positive cases was under 3 years old.Five hMPV gene fragments of sequence were closely related to hMPV in the GenBank.Similarity of hMPV partial N gene with which published at nucleotide levels varied from 80.8%~98.4%.Phylogenetic tree of nucleotide revealed two different gen-otypes.All hMPV positive patients suffered from fever,cough and wheezing.The clinical diagnoses were wheezing pneumo-nia (17),bronchiolitis (9).None of the nasal swabs from 80 healty subjects were tested hMPV specific gene fragment.Con-clusion hMPV is an important viral pathogen in children with respiratory tract infection in Shenzhen and Shantou.Two genotypes may circurate in these areas.

We analyzed the best light intensity for callus induction and maintenance in Vitis vinifera and explored the mechanism of grape callus browning. Tender stem segments of grape cultivar "gold finger" were used to study the effects of different light intensities (0, 500, 1 000, 1 500, 2 000, 2 500, 3 000 and 4 000 Lx) on the induction rate, browning rate and associated enzyme activity and gene expression during Vitis vinifera callus formation. The callus induction rate under 0, 500, 1 000 and 1 500 Lx was more than 92%, significantly higher than in other treatments (P < 0.05). A lower browning rate and better callus growth were also observed during subculture under 1 000 and 1 500 Lx treatments. We found that chlorogenic acid, caffeic acid, p-hydroxybenzoic acid and coumaric acid contents were correlated with the browning rate of callus, among which chlorogenic acid content was positively correlated with the browning rate (P < 0.05). Peroxidase (POD) and polyphenol oxidase (PPO) activities were negatively correlated with the browning rate of callus (P < 0.01). The POD, PPO and phenylalanine ammonialyase (PAL) expression levels were positively correlated with the browning rate at P < 0.05 or P < 0.01. An appropriate light intensity for the tissue culture of Vitis vinifera was 1 000-1 500 Lx, higher or lower light intensities significantly impaired normal callus growth.
Caffeic Acids ; chemistry ; Catechol Oxidase ; chemistry ; Culture Media ; chemistry ; Gene Expression Regulation, Plant ; Light ; Peroxidase ; metabolism ; Phenylalanine Ammonia-Lyase ; metabolism ; Plant Stems ; enzymology ; radiation effects ; Tissue Culture Techniques ; Vitis ; enzymology ; radiation effects

Objective To explore the behavioral dimensions and validity of the light-dark box(LDB),as an model of state anxiety in Kunming mice.Methods The behavior of adult,male,Kunming mice in LDB was recorded twice,for five minutes,with a one-week inter-trial interval.The following parameters were evaluated by factor analysis:latence for the first entry into the dark area(Latence);percentage of time exploring in the light area (Ltime%);percentage of squares crossing in the light area(Lcross%);percentage of rears in the light area (Lrear%);transitions between two areas(Transition);total number of squares crossing in the whole apparatus (Cross);total number of rears in the whole apparatus(Rear);and number of fecal boli(FB).Results There was a good intercorrelation both among Ltime%,Lcross%,Lrear%,and among Transition,Cross,Rear.A good correlation also was found between test and retest for Transition,Rear and FB.Ltime%,Lcress%and Lrear%had significant loadings on Anxiety factor(Loading%:39.54%,35.40%);Transition loaded on Locomotor factor(Loading%:23.78%,19.66%),combined with Cross;Rear loaded on Exploration factor(Loading%:13.75%,22.60%);and FB loaded on Emotionality factor(Loading%:14.42%,13.02%);Latence was not stable.Conclusion Light-dark box,as an animal model of state anxiety in Kunming mice,comprised three behavioral dimensions:anxiety factor,LDB-activity factor(locomotor factor and exploration factor)and emotionality factor.

The vitamin-mineral enriched flour (VMF) was a byproduct obtained from a new processing technique of flour milling, and rich in vitamin B group, minerals and dietary fibers.The addition of 10 or 20% VMF to the stock diet composed mainly of white flour failed to improve PER and growth rate of young rats. When the rats were fed the tested diet for 6 weeks, in which the VMF served as the only diet source ot riboflavin, blood glutathione reductase activity coefficient was 1.13 and 1.09 respectively. This indicated that the amount of riboflavin supplied by VMF could maintain an adequate level in the body of the rats.Adult rats were fed high cholesterol plus 10, 20 and 30% VMF, the contents of serum cholesterol, liver lipids and liver cholesterol were similar to those of the normal diet group, these contents, however, in control group were increased obviously. The results suggested that VMF could protect the rats from the increment of serum cholesterol, liver lipids and cholesterol.

Objective To explore the test-retest reliability of the open field test,as a method for ethology in Kunming mice. Methods The behavior of adult male Kunming mice in the open field test was recorded twice,once for five minutes, with a one-week inter-trial interval. The following parameters were evaluated:number of central squares; number of peripheral squares; percent of central squares; number of horizontal squares; number of rearings; total score; and number of fecal boli. Subsequently,the intraclass correlation coefficient (ICC) and the agreement index (Kappa) were calculated for each of these parameters. Results Both ICC and Kappa for total score( ICC =0.902, P＜0.01; Kappa =0.680, P＜0.01; r=0.815, P＜0.01 ) showed a good reliability and correlation between the test trials. Only ICC for number of central squares ( ICC = 0. 578, P ＜ 0.01 ), number of peripheral squares( ICC =0.615, P＜0.01 ) ,percent of central squares(ICC=0.523, P＜0.01) ,number of horizontal squares( ICC = 0. 607, P ＜ 0.01 ) and number of rearings ( ICC = 0. 606, P ＜ 0.01 ) indicated a fair reliability between the test trials. Neither ICC nor Kappa for number of fecal boli displayed a significant reliability between the test trials. The difference of these parameters between test and retest, such as number of peripheral squares( t =3.01, P＜0.01 ),number of horizontal squares( t = 3.34, P ＜ 0.01 ) and total score( t = 7.86, P ＜ 0.01 ) were significant. Conclusion The parameter,total score for Kunming mice in the open field test,is found to be stable over time, and indicates a good reliability with a one-week inter-trial interval.