OBJECTIVEIntravenous administration of basic fibroblast growth factor (bFGF) is effective to reduce the volume of cerebral infract due to ischemia. This study was designed to investigate the molecular mechanism, especially the signal transduction pathways, involved in this protective role of bFGF.

METHODSAnoxia-reoxygenation treated astrocytes were used to study the role of mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MAPK/ERK kinase, MEK)-ERK signaling pathway after exogenous bFGF administration by Western blot. Electrophoretic mobile shift assay was used to detect the binding activity of early growth response factor-1 (Egr-1), an important transcription factor for endogenous bFGF.

RESULTSbFGF could protect some signal transduction proteins from the oxygen-derived free radicals induced degradation. ERK1/2 was activated and involved in Egr-1 binding activity enhancement induced by exogenous bFGF.

CONCLUSIONMEK-ERK MAPK cascade may be an important signal transduction pathway contributed to bFGF induced enhancement of Egr-1 binding activity in anoxia-reoxygenation injured astrocytes.

Animals ; Animals, Newborn ; Astrocytes ; drug effects ; metabolism ; Cells, Cultured ; Early Growth Response Protein 1 ; metabolism ; Electrophoretic Mobility Shift Assay ; methods ; Fibroblast Growth Factors ; pharmacology ; Mitogen-Activated Protein Kinase Kinases ; metabolism ; Oxygen ; metabolism ; Protein Binding ; drug effects ; Rats ; Signal Transduction ; physiology ; Time Factors

Objective To investigate the polymorphism of human cytomegalovirus（HCMV）UL146 gene in clinical strains,and to evaluate its clinical diagnostic and therapeutic value of gene.Methods The UL146 gene of clinical strains was examined by quantitative polymerase chain reaction（Q-PCR）or general polymerase chain reaction（PCR）.Positive samples of PCR amplification were sequenced and analyzed.Results High variability of UL146 gene was found among 28 HCMV strains.According to phylogenetic analysis,all sequences of UL146 in clinical strains could be divided into three types and four subtypes.Chemokine ELRCXC region was highly conserved in all sequences.Conclusion HCMV-UL146 genes showed a high degree of polymorphism,and its encoded chemokine ELRCXC region was highly con-served.The relationship between HCMV-UL146 gene′s polymorphism and different clinical symptoms of HCMV infection was unclear.

OBJECTIVETo study the gap junction mechanisms for synergistic effects of liuwei di-huang pill (LDP) containing serum on HSV-tk/GCV suicide gene therapy of mouse malignant melanoma B16 cells.

METHODSThe LDP containing serum (2.5% LDP serum group, 5.0% LDP serum group, and 10.0% LDP serum group) and the control serum group were set up. The effects of LDP on mRNA expressions of Cx26 and Cx43 in mouse malignant melanoma B16 cells were detected by RT-PCR assay. The effects of LDP on protein expressions of Cx26 and Cx43 in B16 cells were detected using Western blot and indirect immunofluorescence assay. The interference efficiencies of Cx26-309, Cx26-337, Cx26-367, Cx26-2098 SiRNA on Cx26 gene in B16 cells were detected using RT-RCR technique. Cx26-2098 SiRNA, due to the optimal interference efficiency, was chosen to interfere Cx26 gene, and the bystander effect of LDP + HSV-tk/GCV was observed. The effects of the gap junctional intercellular communication (GJIC) inhibitor glycyrrhetinic acid on the killing of LDP + HSV-tk/GCV system to B16 cells were detected by MTT assay. In this experiment, the control serum group, 2.5% LDP serum group, 5. 0% LDP serum group, 10.0% LDP serum group, the control serum combined GCV group, 2.5% LDP serum combined GCV group, 5.0% LDP serum combined GCV group, 10.0% LDP serum combined GCV group, 2.5% LDP serum combined GCV + glycyrrhetinic acid group, 5.0% LDP serum combined GCV + glycyrrhetinic acid group, 10.0% LDP serum combined GCV + glycyrrhetinic acid group were set up. The final concentration of GCV was 20 micromol/L. The final concentration of glycyrrhetinic acid was 50 micromol/L.

RESULTSLDP containing serum could increase the protein and mRNA expressions of Cx43 in B16 cell in a dose-dependent manner. It had bi-directional regulation on the Cx26 protein and mRNA expressions. The low dose LDP had inhibition while high dose LDP could up-regulate its expression. After SiRNA interfered Cx26 gene, there was no obvious change in the bystander effect of LDP combined suicide gene therapy between before and after interference. There was significant reduction in the inhibition rate between before (48.75%, 59.39%, and 69.28%) and after blockage (29.14%, 38.71%, and 58.13%) of glycyrrhetinic acid in the 2. 5%, 5.0%, 10.0% LDP serum combined GCV groups (P <0.01).

CONCLUSIONThe synergistic effects of LDP containing serum on HSV-tk/GCV suicide gene therapy in mouse malignant melanoma B16 cells were correlated with the gap junction mechanisms, which might be achieved through increasing the expressions of Cx26 and Cx43.

Animals ; Cell Line, Tumor ; Connexin 26 ; Connexin 43 ; metabolism ; Connexins ; metabolism ; Drugs, Chinese Herbal ; therapeutic use ; Gap Junctions ; metabolism ; Genetic Therapy ; Male ; Melanoma ; therapy ; Mice ; Rats, Sprague-Dawley ; Serum

OBJECTIVETo explore the effects of rearing patterns on diet and temperament traits among infants in urban areas.

METHODSA total of 480 25-30-month-old infants were randomly selected from the birth cohort in Hefei Maternal and Child Health Care Center in 2008. A household survey was conducted using China Toddler Temperament Scale (CTTS), Dietary Characteristics Questionnaire and Family Environment Questionnaire.

RESULTSOf the 430 surveyed households, there were three main rearing patterns including parents rearing pattern (Group A), grandparents rearing pattern (Group B) and joint rearing pattern (Group C), accounting for 33.0%, 21.2% and 45.8%, respectively. Infants in Group A tended to adopt processed food pattern, with poor rhythmicity and adaptability; infants in Group B tended to adopt fruit, vegetable, and cereals-based food pattern, with relatively poor rhythmicity; infants in Group C tended to adopt aquatic products and fruit/vegetable-based food pattern, with good rhythmicity and adaptability. Linear regression model showed that infants who consumed more aquatic products, high-protein food, and fruits/vegetables had more positive temperamental traits, whereas infants who consumed more processed foods had more negative temperamental traits.

CONCLUSIONSA joint rearing pattern may be a favorable rearing style for infants aged 25-30 months in urban areas in terms of diet and temperament traits.

Child Rearing ; Child, Preschool ; Diet ; Humans ; Infant ; Infant Nutritional Physiological Phenomena ; Temperament

OBJECTIVETo evaluate the impacts of maternal weight gain during pregnancy on offspring weight and obesity from birth to 24 months of age.

METHODSThe information on maternal pre-pregnancy body mass index (BMI), gestational weight gain and demographic characteristics were collected from 317 pregnant women. The information on offspring weight, BMI and breastfeeding data was obtained from various follow-up examinations from 0 to 24 months of age.

RESULTSThe logistic regression analysis showed that excessive gestational weight gain resulted in an increased risk of obesity in children at age of 6 months (adjusted RR=3.56, 95% CI:1.31-8.35) and 9 months (adjusted RR=2.87, 95% CI: 1.04-3.28) after adjustment for potential confounding factors. The linear regression model showed that there were significant correlations between gestational weight gain and Z score of weight in offsprings at birth (β=0.032, 95% CI: 0.008-0.057), 3 months (β=0.037, 95% CI: 0.013-0.062), 6 months (β=0.043, 95% CI: 0.017-0.068), 9 months (β=0.038, 95% CI: 0.013-0.063) and 12 months (β=0.034, 95% CI: 0.009-0.059), but not at 18 months and 24 months.

CONCLUSIONSExcessive gestational weight gain may affect offspring weight and increase the risk of obesity in children from birth to 12 months of age. During their second year of life, this effect will temporarily disappear.

Body Mass Index ; Body Weight ; Female ; Follow-Up Studies ; Humans ; Linear Models ; Logistic Models ; Obesity ; etiology ; Pregnancy ; Weight Gain

Objective To observe the efficacy and safety of the rat nerve growth factor combined with αlipoic acid in the treatment of diabetic periph-eral neuropathy (DPN).Methods Eighty -eight patients with DPN were randomly divided into two groups and they were given diabetes diet and insu -lin or oral hypoglycemic agents to control blood sugar.The control group (n =43) was given αlipoic acid 0.6 g,qd; the trial group (n =45) were given rat nerve growth factor injection 20 μg based on the control group,im, qd.Twenty -one days as a treatment cycle.Before and after treatment, the total symptom score(TSS) ,the nerve conduction velocity(NCV),microalbu-minuria and high sensitivity C -reactive protein were tested and recorded respectively before and after treatment in 2 groups.Results The NCV of the trial group was higher than those of the control group (P <0.01); after treatment,TSS was significantly lower in the trial group than in the control group (P <0.01); the microalbuminuria and high sensitivity C -reactive protein of the trial group descended more apparently than those of the control group (P <0.05).The total effective rate of the trial group was higher than that of the control group (P <0.05).Conclusion Rat nerve growth factor combined with αlipoic acid treatment is safe and effective for the treatment of diabetic peripheral neuropathy.

Objective: To construct and identify lentiviral vector containing human ILK-shRNA and mda7 gene.Methods:Based on the human ILK gene sequences , RNAi target sequences were designed and cloned into the lentiviral vector pSicoR -eGFP by restriction endonuclease HpaⅠand XhoⅠdouble digestion and T 4 DNA ligase ligation . Based on the human mda7 gene sequences , PCR primers were designed to clone the full-length mda7, and were cloned into the lentiviral vector pLVX-Puro.After the candidate clones were identified by DNA sequencing , the recombinant plasmid and the three packaging plasmids were co-transfected into the human embryonic kidney 293T cells by lipofectamine 2000 to produce the lentiviral particles .Human prostate cancer PC-3 cells were infected with the constructed lentiviral vector .The ILK and mda7 expression levels in PC-3 cells were quantified by qPCR and Western blot , respectively .The effect of ILK and mda7 on proliferation and migration of PC-3 cells were assessed by MTT method and Transwell assay, respectively.Results: ILK-pSicoR-eGFP and mda7-pLVX-Puro lentiviral vectors were successfully constructed .Strong green fluorescence was observed in the 293T cells under the fluorescent microscope after co-transfection of 293T cells with 4 plasmids of lentiviral vector .The transfection efficiency of the collected virus exceeded 90%in the 293T cells and the PC-3 cells were infected with the lentiviral particles with high efficiency .The A and B lentiviral vector inhibited the expression of ILK at both the mRNA and protein levels in PC-3 cells significantly .The mda7-pLVX-Puro lentiviral vector increased the expression of mda 7 in PC-3 cells, and the ability was maintained for one month.Within 96 h, ILK and mad7 significantly inhibited the proliferation and migration of PC-3 cells ( Ps <0.05 ) . Conclusion: The lentiviral vectors of ILK knockdown and mda 7 over-expression have been successfully constructed and identified . The recombinant lentivirus can efficiently infect human prostate cancer PC -3 cells, in which ILK expression is inhibited and mda 7 is over-expressed .

OBJECTIVESelecting variable-number tandem-repeat (VNTR) loci for different serogroups of Shigella spp to explore and establish multilocus variable-number tandem-repeat analysis (MLVA) method, in order to study the molecular characteristic of the isolated strains.

METHODSOf the Shigella strains found by dysentery surveillance in Beijing from 2001 to 2009, 180 strains were selected for this study, according to the number and serotypes of the surveillant strains, at the ratio of 15%; including 50 strains of Shigella sonnei and 130 strains of Shigella flexneri. After screening the polymorphism of the 18 VNTR loci, 10 VNTR loci (sh1-sh10) were retained and constructed three groups of multi-PCR methods to detect all he 180 strains and analyze MLVA molecular subtypes using capillary segments.

RESULTSA range of 2 to 11 alleles were found on the 10 VNTR loci among the 180 Shigella strains, with a diversity index value between 0.158 and 0.766. The 10 loci showed diversity in different serogroups, such as only one allele found in sh6 of Shigella flexneri, sh2 and sh3 of Shigella sonnei individually. The isolated 180 strains were divided into 84 MLVA subtypes, with a resolution ratio D value at 0.967 (95%CI: 0.956 - 0.978). The 130 strains of Shigella flexneri were divided into 63 subtypes, named as TF001-TF063; among which TF001, TF002 and TF 005 were the dominant subtypes, accounting to 17, 16 and 15 strains respectively. The 50 strains of Shigella sonnei were divided into 21 subtypes, named as TS001-TS021; among which TS002 (14 strains) and TS001 (7 strains) were the dominant subtypes.

CONCLUSIONMLVA subtyping method including 10 VNTR loci was preliminarily developed. The MLVA cluster analysis revealed that the subtypes of Shigella strains isolated in Beijing were diverse, and suggested the possibility of multiple-clone source.

Alleles ; Bacterial Typing Techniques ; methods ; China ; DNA, Bacterial ; genetics ; Genotype ; Minisatellite Repeats ; Polymorphism, Genetic ; Shigella ; classification ; genetics ; isolation & purification

Acquired Immunodeficiency Syndrome ; epidemiology ; Adult ; China ; epidemiology ; Cross-Sectional Studies ; Female ; HIV Infections ; epidemiology ; Humans ; Middle Aged ; Poverty Areas ; Young Adult

OBJECTIVETo evaluate the anticoagulant and antineoplastic activities of chemically modified low-molecular-weight heparin (LMWH).

METHODSLMWH obtained by splitting unfractionated heparin (UFH) with sodium periodate oxidation and sodium borohydride reduction was subjected to acetylation catalyzed by DCC and DMAP to produce acetylated LMWH (ALMWH). The anticoagulant activity of ALMWH was determined in mice, and its antiproliferative and anti-invasion activities was assessed in human breast cancer cells MDA-MB-231 and MFC-7.

RESULTSThe anticoagulant activity of LMWH was decreased significantly after acetylation. The concentrations of commercial LMWH* and ALMWH for doubling the coagulation time (CT) were 33.04 µmol/L and 223.56 µmol/L, respectively, and the IC(50) of ALMWH for doubling CT was 6 times of that of LMWH*. ALMWH and LMWH at 0.1, 0.3, 0.9, 2.7 and 8.1 mmol/L both significantly inhibited the proliferation of MDA-MB-231 and MCF-7 cells in a concentration-dependent manner, but ALMWH produced stronger inhibitory effects. The IC(50) of LMWH and ALMWH for inhibiting cell proliferation was 3168.4 µmol/L and 152.6 µmol/L in MCF-7 cells, and 12299.6 µmol/L and 22.2 µmol/L in MDA-MB-231 cells, respectively. ALMWH and LMWH all markedly suppressed the invasion of MDA-MB-231 cells with comparable effects.

CONCLUSIONChemical modification of structure can endow LMWH with a low anticoagulant and high antiproliferative activities.

Animals ; Anticoagulants ; chemistry ; pharmacology ; Antineoplastic Agents ; chemistry ; pharmacology ; Blood Coagulation ; Blood Coagulation Tests ; Cell Line, Tumor ; Heparin ; chemistry ; Heparin, Low-Molecular-Weight ; chemistry ; pharmacology ; Humans ; Mice