Objective To observe effect of TNF-? on PPAR?2 mRNA expression in 3T3-L1 cells transfected with human recombinant adiponectin. Methods The 3T3-L1 preadipocytes were transfected with the recombinant plasmid pcDNA3.1+-hADPN.The PPAR?2 mRNA expression was quantitated by semi-quantitative RT-PCR. Results (1)Compared with controls(the 3T3-L1 cells and the 3T3-L1 cells with plasmid), the PPAR-?2 mRNA expression of 3T3-L1 cells with human recombinant adiponectin was higher(P

Objective To study the expression and cellular localization of STAT3 and SOCS-3 in the motor neurons of normal rat spinal cord. Methods Immunocytochemistry and morphometry methods were used in the present study. Results STAT3 immunocytochemical staining was mainly distributed in the cytoplasm of the motor neurons of the ventral horns, SOCS-3 immunoreactive products were extensively distributed in the neurons of the ventral and dorsal horns, glia and fibers of the spinal cord. In the ventral horn, SOCS-3 immunoreactivity was found in the nuclei and/or cytoplasm of the motor neurons.Conclusions STAT3 and SOCS-3 are extensively distributed in the normal adult rat spinal cord and SOCS-3 is existence in the forms of nuclear or cytoplasmic protein.

Objective To study the basic expression and cellular localization of SOCS-3 in normal rat retina. Methods Neuro-immunocytochemistry techniques were used. Results SOCS-3 positive cells were widely distributed in the ganglion cell layer (GCL) and inner nuclear layer (INL) in the retina. In the GCL, SOCS-3 immunoreactivity was mainly in the neuclei of the ganglion cells.Some of the SOCS-3 positive cells in INL were M?ller cells.Conclusion Basal expression of SOCS-3 is widely present in the neurons and glia in normal adult rat retina and mainly in the form of nuclear protein.

Objective To study the effects ofautologous bone marrow mononuclear cells (BMMCs)transplantation during coronary artery bypass gafting (CABG) on myocardial infarct size. Methods Forty myocardial infarction patients diagnosed by coronary angiography (CAG) and SPECT and confirmed at surgery were enrolled and randomly assigned CABG alone (group Ⅰ) or CABG with intramyocardial or intracoronary injection of autologous BMMCs (group Ⅱ), 20 cases in each group. Baseline and followed up evaluations included SPECT and NYHA-FC before and after 6 months operation, recorded the major adverse cardiac events (MACE) at the same time. The number of autologous BMMCs injected was (6.84 ± 2.88) ×107 in group Ⅱ. Results There was no procedure-related complication during 6 months followed up in all patients. After 6 months operation,left ventricular ejection fraction in group Ⅱ was significantly higher than that in group Ⅰ [(57.40 ±5.21)% vs. (50.75 ±5.88)%,t =3.79,P＜0.05],NYHA-FC in group Ⅱ was significantly improved than that in group Ⅰ [(1.30 ± 0.47) grades vs. (1.85 ± 0.59) grades, t = 3.27, P ＜0.05],SPECT showed myocardial infarct size in group Ⅱ was significantly lower than that in group Ⅰ[(14.57 ±5.20)% vs. (20.45 ±5.18)% ,P ＜0.05]. Conclusion Autologous BMMCs transplantation during CABG is safe and feasible, which can reduce the myocardial infarct size in patients with myocardial infarction.

Objectives:To examine whether lipopolysaccharide (LPS) induced apoptosis correlates with TNF ? release by type Ⅱ alveolar epithelial cells (AEC Ⅱ), whether TNF ? knockout (TNF KO) abrogates the induction of apoptosis by LPS and whether TNF ? is sufficient to induce apoptosis in this cell type. Methods:AEC Ⅱ was isolated from wild type mice and TNF KO mice. Cells were stimulated with LPS or recombinant murine TNF ? for 24 h. TNF ? in culture supernatant was determined by ELISA following LPS stimulation. Apoptosis was determined by the TUNEL assay after treatment with either LPS or TNF ?. Results:LPS induced apoptosis in wild type AEC Ⅱ in a concentration dependent manner. LPS induced AEC Ⅱ apoptosis was accompanied by a 11 fold increase from (0.073?0.065) ng/ml in controls to( 0.94?0.14)ng/ml in 50 ?g/ml of LPS( P

OBJECTIVETo study the acireductone dioxygenase (designated as SmARD) gene of Salvia miltiorrhiza through bioinformatics and characterization of its tissue expression and response expression on stress in shoot.

METHODSmARD gene was obtained by sequencing cDNA library constructed by us. BLAST was used for alignment, ORF finder software was applied to find open reading frame, prosite was used to analyze the protein characterization. Semi-quantitative RT-PCR was used to detect the gene expression level.

RESULTThe full -length cDNA of SmRAD was 688 bp long with a 591 bp ORF (open reading frame) that putatively encoded a polypeptide of 196 amino acids; with a predicted molecular mass of 23.27 kDa. The deduced amino acid sequence of SmRAD of gene shared high homology with other known RADs. Semi-quantitative RT-PCR analysis indicated that SmRAD was constitutively expressed in roots, stems, flower and leaves of S. miltiorrhiza, with the high expression in roots. In addition, SmRAD expression level under different stress condition was also analyzed in root, including signaling components for plant defence responses, such as methyl jasmonate, salicylic acid and ABA, as well as drought, cold and salt abiotic stress. The expression of SmRAD was suppressed by water deficit treatment for 3 d, 150 mmol x L(-1) NaCl, 4 degrees C cold and 100 mmol x L(-1) ABA treatment for 1 d, but induced by 100 mmol x L(-1) MJ and 10 mmol x L(-1) ETH.

CONCLUSIONA novel SmARD gene was cloned from S. miltiorrhiza. This study will enable us to further understand the role of SmARD in the defense response under different abiotic stress and in synthesis of active cmpounds in S. miltiorrhiza at molecular level.

Amino Acid Sequence ; Cloning, Molecular ; Dioxygenases ; genetics ; metabolism ; Gene Expression Regulation, Plant ; Molecular Sequence Data ; Phylogeny ; Plant Roots ; genetics ; metabolism ; Salvia miltiorrhiza ; genetics ; metabolism ; Sequence Alignment ; Sequence Analysis, DNA ; Sequence Homology, Amino Acid ; Stress, Physiological

Glycerol from oil hydrolysis industry is being considered as one of the abundent raw materials for fermentation industry. In present study, the aerobic and anaerobic metabolism and growth properties on glycerol by Esherichia coli CICIM B0013-070, a D-lactate over-producing strain constructed previously, at different temperatures were investigated, followed by a novel fermentation process, named temperature-switched process, was established for D-lactate production from glycerol. Under the optimal condition, lactate yield was increased from 64.0% to 82.6%. Subsequently, the yield of D-lactate from glycerol was reached up to 88.9% while a thermo-inducible promoter was used to regulate D-lactate dehydrogenase transcription.
Aerobiosis ; Anaerobiosis ; Escherichia coli ; genetics ; metabolism ; Fermentation ; Glycerol ; metabolism ; L-Lactate Dehydrogenase ; metabolism ; Lactic Acid ; biosynthesis ; Promoter Regions, Genetic ; genetics ; Temperature

OBJECTIVETo examine whether lipopolysaccharide (LPS)-induced apoptosis correlates with TNF-alpha release by type II alveolar epithelial cells (AEC II), whether TNF-alpha knockout (TNF KO) abrogates the induction of apoptosis by LPS and whether TNF-alpha is sufficient to induce apoptosis in this cell type.

METHODSAEC II were isolated from wild type mice and TNF KO mice. Cells were stimulated with LPS or recombinant murine TNF-alpha for 24 h. TNF-alpha in culture supernatant was determined by ELISA following LPS stimulation. Apoptosis was determined by the terminal deoxynucleotidyl transferase end-labeling (TUNEL) assay after treatment with either LPS or TNF-alpha.

RESULTSLPS induced apoptosis in wild type AEC II in a concentration-dependent manner. LPS-induced AEC II apoptosis was accompanied by an 11-fold increase (from 0.073 +/- 0.065 ng/ml in control to 0.94 +/- 0.14 ng/ml in 50 micro g/ml of LPS, P < 0.01) in TNF-alpha release. However, increasing concentrations (5 or 25 ng/ml) of recombinant murine TNF-alpha failed to induce AEC II apoptosis. In addition, apoptosis did occur in AEC II isolated from TNF KO mice following LPS stimulation.

CONCLUSIONSThis study confirms that LPS induces TNF-alpha release and apoptosis in murine AEC II in vitro. Exogenous TNF-alpha failed to induce AEC II apoptosis, and apoptosis occurred following LPS stimulation in cells lacking the ability to produce TNF-alpha. Taken together, these results suggest that LPS-induced AEC II apoptosis occurs by a TNF-alpha-independent mechanism.

Animals ; Apoptosis ; drug effects ; Cells, Cultured ; Epithelial Cells ; drug effects ; Lipopolysaccharides ; pharmacology ; Mice ; Mice, Inbred C57BL ; Pulmonary Alveoli ; cytology ; drug effects ; Tumor Necrosis Factor-alpha ; pharmacology

When Escherichia coli CICIM B0013-030 (B0013, ack-pta, pps, pflB) was used for D-lactate production, succinate and acetate were the main byproducts (as much as 11.9 and 7.1% the amount of lactate respectively). In order to decrease the byproduct levels, we inactivated succinate and acetate synthesis in B0013-030. Two recombinant plasmids containing mutation cassettes of frdA::difGm and tdcDE::difGm respectively were constructed first. The mutation cassettes were used to delete the target genes on the chromosomal by Red recombination. Subsequently, the antibiotic resistance gene was excised from the chromosomal by Xer recombination. Thereby, mutants B0013-040B (B0013-030, frdA) and B0013-050B (B0013-040B, tdcDE) were produced. D-lactate producing abilities of the engineered strains were tested both in shake flasks and in bioreactors using two-phase fermentation (aerobic growth and anaerobic fermentation) with glucose as the sole carbon source. When fermentation was carried out in shake flasks, inactivation of frdA in B0013-030 to produce B0013-040B reduced succinate accumulation by 80.8%. When tested in a 7-liter bioreactor, B0013-040B accumulated 114.5 g/L D-lactate of over 99.9% optical purity. However, 1.0 g/L succinate and 5.4 g/L acetate still remained in the broth. Further inactivation of tdcD and tdcE genes in B0013-040B to produce B0013-050B decreased acetate and succinate accumulation to 0.4 g/L and 0.4 g/L respectively, and lactate titer was as much as 111.9 g/L (tested in the 7-liter bioreactor). In lightof the lower byproduct levels and high lactate production, strain B00 13-050B may prove useful for D-lactate production.
Acetates ; metabolism ; Escherichia coli ; genetics ; metabolism ; Fermentation ; Genetic Engineering ; Lactic Acid ; biosynthesis ; Metabolic Networks and Pathways ; genetics ; Mutation ; Plasmids ; genetics ; Succinic Acid ; metabolism

Objective To investigate the puncture effect of radiofrequency thermocoagulation of semilunar ganglion through the foramen ovale with ultrasound guidance and CT verification in the treatment of V3 branch trigeminal neuralgia(TN).Methods Forty-eight V3 branch TN patients,19 males and 29 females,aged＞18 years,BMI≤28 kg/m2,undergoing radiofrequency thermocoagulation of semilunar ganglion through the foramen ovale were selected.Patients were divided into two groups by random number table method:ultrasound guidance combined with CT verification group(group U)and CT guidance group(group C),24 patients in each groups.According to grouping result,patients received radiofrequency ther-mocoagulation of semilunar ganglion with foramen ovale puncture performed under the ultrasound guidance and CT verification or CT guidance respectively.The success cases of puncture,the number of puncture,ra-diation dose,puncture time,operation time and the occurrence of treatment complications were recorded.Numerical rating scale(NRS)scores and Barrow neurological institute(BNI)scores were recorded before surgery,2,4,12,and 24 weeks after surgery,and good postoperative pain relief rate(BNI Ⅰ or Ⅱ)was calculated.Results All patients in both groups completed puncture operationand radiofrequency thermo-coagulation.Compared with group C,group U had fewer number of puncture,lower radiation exposure,and shorter puncture and surgical times(P＜0.05).Compared with baseline before operation,NRS scores were significantly lower in both groups at 2,4,12,and 24 weeks after surgery(P＜0.05).There was no signif-icant difference in the rate of good pain relief 2,4,12,and 24 weeks after surgery between the two groups.There was no significant difference in the incidence of facial hematoma between the two groups.No other se-rious complications were found in both groups.Conclusion Radiofrequency thermocoagulation of semilunar ganglion through the foramen ovale with ultrasound guidance and CT verification is a safe and feasible method for the treatment of the V3 branch TN.Compared with CT guidance,ultrasound guidance combined with CT verification can significantly improve the puncture accuracy and reduce the radiation exposure related to treatment.