AIM: To study the changes of brain - derived neurotrophic factor ( BDNF ) expression in gene modified bone marrow mesenchymal stem cells ( BMSC) .
●METHODS:BMSC were divided into blank control group ( without transfected BMSC ) , negative control group ( empty vector without BDNF gene transfected BMSC) and experimental group ( BDNF gene transfected BMSC) . The expression of BDNF mRNA in BMSC was measured by Realtime PCR, and the expression of BDNF in BMSC was measured by ELlSA.
●RESULTS:The BDNF mRNA expressions of 3, 4, 5, 6, 7 and 8-generation BMSC cells in the experimental group were higher than those in the blank control group and negative control group. The differences were statistically significant (P3: F=491. 788, P<0. 05; P4: F=380. 112, P<0. 05;P5:F=1854. 929, P<0. 05; P6: F=224. 540, P<0. 05;P7:F=619. 155, P<0. 05; P8: F=10. 092, P<0. 05). As the BMSC cells in the experimental group passaging, the BDNF mRNA expressions in the experimental group decreased. The difference of BDNF mRNA expression among different passage cells was statistically significant (F=298. 603, P<0. 05). The BDNF secretion of 3, 4, 5, 6, 7 and 8-generation BMSC cells in the experimental group were higher than those in the blank control group and negative control group. The differences were statistically significant (P3: F=520. 609, P<0. 05; P4: F=734. 520, P<0. 05;P5:F=152. 847, P<0. 05;P6:F=80. 372, P<0. 05; P7:F=96. 083, P<0. 05;P8:F=38. 532, P<0. 05). As the BMSC cells in the experimental group passaging, the BDNF secretion decreased. The difference of BDNF secretion among different passage cells was statistically significant (F=230. 084, P<0. 05).
●CONCLUSION:Long-term expression of BDNF in BMSC can be enhanced by genetic engineering.

Four small GTPase genes which may be relative to sclerotial development were firstly cloned from medicinal fungus Polyporus umbellatus using rapid amplification of cDNA end PCR (RACE) method. The results showed that full-length cDNA of PuRhoA was 698 bp contained 585 bp ORF, which was predicted to encode a 194 amino acid protein with a molecular weight of 21.75 kD with an isoelectric point (pI) of 6.44; the full length cDNA of PuRhoA2 was 837 bp in length and encoded a 194 amino acid protein with a molecular weight of 21.75 kD and an isoelectric point (pI) of 6.33; the full length cDNA of Puypt1 was 896 bp in length and encoded a 204-aa protein with a molecular weight of 22.556 kD and an isoelectric point (pI) of 5.75; the full length cDNA of PuRas was 803 bp in length and encoded a 212-aa protein with a molecular weight of 23.821 kD and an isoelectric point (pI) of 5.2. There are fani acyl transferase enzyme catalytic site and myrcene-transferase enzyme catalytic site in PuRhoA1 while the PuRhoA2 only possess myrcene-transferase enzyme catalytic site. Puypt1 contains the Rab1-Ypt1 conserved domain of small GTPase family and PuRas contains the fani acyl transferase enzyme catalytic site. According to the phylogenetic analysis all these four small GTPase clustered with basidiomycete group. Quantitative real-time PCR analysis revealed that Puypt1, PuRas and PuRhoA1 transcripts were significantly higher in the beginning of sclerotial formation than that in the mycelia, whereas the transcripts levels of PuRhoA2 gene were particularly lower in sclerotia than that in mycelia, suggesting that these four genes might be involved in P umbellatus selerotial development.

Four small GTPase genes which may be relative to sclerotial development were firstly cloned from medicinal fungus Polyporus umbellatus using rapid amplification of cDNA end PCR (RACE) method. The results showed that full-length cDNA of PuRhoA was 698 bp contained 585 bp ORF, which was predicted to encode a 194 amino acid protein with a molecular weight of 21.75 kD with an isoelectric point (pI) of 6.44; the full length cDNA of PuRhoA2 was 837 bp in length and encoded a 194 amino acid protein with a molecular weight of 21.75 kD and an isoelectric point (pI) of 6.33; the full length cDNA of Puypt1 was 896 bp in length and encoded a 204-aa protein with a molecular weight of 22.556 kD and an isoelectric point (pI) of 5.75; the full length cDNA of PuRas was 803 bp in length and encoded a 212-aa protein with a molecular weight of 23.821 kD and an isoelectric point (pI) of 5.2. There are fani acyl transferase enzyme catalytic site and myrcene-transferase enzyme catalytic site in PuRhoA1 while the PuRhoA2 only possess myrcene-transferase enzyme catalytic site. Puypt1 contains the Rab1-Ypt1 conserved domain of small GTPase family and PuRas contains the fani acyl transferase enzyme catalytic site. According to the phylogenetic analysis all these four small GTPase clustered with basidiomycete group. Quantitative real-time PCR analysis revealed that Puypt1, PuRas and PuRhoA1 transcripts were significantly higher in the beginning of sclerotial formation than that in the mycelia, whereas the transcripts levels of PuRhoA2 gene were particularly lower in sclerotia than that in mycelia, suggesting that these four genes might be involved in P umbellatus selerotial development.
Amino Acid Sequence ; Cloning, Molecular ; DNA, Complementary ; Fungal Proteins ; genetics ; GTP Phosphohydrolases ; genetics ; Genes, Fungal ; Mycelium ; Phylogeny ; Polyporus ; enzymology ; genetics ; Real-Time Polymerase Chain Reaction

Objective To characterize the antibiotic resistance,homology and carbapenemase genotypes of imipenem resistant Acinetobac1ter baumannii isolated from our hospital,and analyze the clonal relatedness of the test strains.Methods Ninety five strains of imipenem resistant A.baumannii were isolated from August 2003 to December 2004 in the First Affiliated Hospital, College of Medicine,Zhejiang University.The MICs of 16 antimicrobial agents against these strains were determined by agar dilution and E-test method.The homology of these isolates was analyzed by pulse-field gel electrophoresis(PFGE).The coding gene of carbapenemases was amplified.PCR products were purified,cloned and sequenced.Plasmid DNA was extracted and purified.Conjugation and Southern blot were performed to locate the position of oxa 23 gene.Results The resistance rates to ampicillin-sulbactam and cefoperazone sulhactam were 67.9% and 30.2%.Polymyxin E had the lowest resistance rate of 17%. The resistance rate to other antimicrobial agents was higher than 90%.The 95 strains,isolated from 10 clinical units,were classified into 6 clones.Clones A and B were predominant clones.All strains produced carbapenemases which were confirmed as OXA 23 by PCR and sequencing analysis.No plasmid was extracted and conjugation was not successful.Southern bolt showed that oxa-23 gene was located on Apal-digested chromosomal segments about 220 kb and 200 kb in Clones A and B,re spectively.Conclusions OXA 23-producing A.baumannii has become one of the most important multi-resistant pathogens in our hospital.Clones A and B have widely spread in our hospital.Oxa-23 gene is located on chromosomal DNA.

Objective To investigate the resistant mechanism of imipenem-resistant K. pneumoniae.Methods The minimal inhibitive concentrations (MICs) of the antimicrobial agents were determined by Etest.Isoelectric focusing electrophoresis (IEF),plasmid extraction,conjugation, transformation,PCR amplification,cloning and sequencing were carried out for analyzing the encoding gene of ?-1actamases.Results Three kinds of ?-1actamases were detected with pIs of 7.2,6.7,and 5.4.in a clinical strain of K.pneumoniae.These ?-1actamases were TEM-I (pI,5.4),SHV-12 (pI,8.2) and KPC-2 ( pI,6.7 ) confirmed by sequencing of the PCR products.Only one band of ?-1actamase with pI 6.7 was displayed in the transformant.A 1500 bp segment,which contained the KPC-2 gene confirmed by nucleotide sequence analysis,was cloned from a 60 000 bp plasmid of the transformant.Conclusion The strain of K.pneumoniae resistant to imipenem produces a plasmid-mediated carbapenemase KPC-2 which belongs to Bush group 2f,class A ?-1actamase.

Actins ; metabolism ; Aged ; Breast Neoplasms ; metabolism ; pathology ; surgery ; Carcinoma ; metabolism ; pathology ; surgery ; Diagnosis, Differential ; Fasciitis ; metabolism ; pathology ; Female ; Fibroma ; metabolism ; pathology ; surgery ; Humans ; Keratin-5 ; metabolism ; Mastectomy, Modified Radical ; Neoplasms, Muscle Tissue ; metabolism ; pathology

OBJECTIVEThe same person's pulmonary venous blood volume, left atrial volume and stroke volume were measured by lung CT scans and cardiac CT angiography (CTA). Then their relationships were analyzed in order to investigate the mechanism of breathing control.

METHODSAs we described before, full pulmonary vascular (-0.6mm) volume was accurately calculated by three-dimensional imaging technology from lung CT scan; left atrial volume and stroke volume of left ventricle were calculated from the CTA data. Then the relationships among them were analyzed for estimation of the lung-artery time.

RESULTSThe total volume of lung and pulmonary vascular blood was 3486 ± 783 (2156-4418) ml, and the pulmonary vascular blood volume was 141 ± 20 (105-163) ml. The estimated pulmonary venous volume was 71 ± 10 (52-81) ml. Left atrial volume at the end diastolic was 97 ± 39 (53-165) ml, Stroke volume of left ventricle was 86 ± 16 (60-106) ml. Pulmonary venous volume and the left atrial volume were double of stroke volume(1.7-2.4).

CONCLUSIONThe estimated lung-artery time was three heart beat.

OBJECTIVEBecause the traditional loop of breathing control and regulation effect on blood circulation, there was rare study of pulmonary vein capacity. We need a noninvasive and accurate pulmonary vascular capacity measurement and analysis method.

METHODSTwelve normal volunteers were performed a total lung CT scan, image data analysis processing by computer software, the whole lungs from the apex to the base of lung with 40-50 layers by hand-cut, the connection between adjacent layers automatically by a computer simulation, the full pulmonary vascular (≥ 0.6 mm) were treated by high-accuracy three-dimensional imaging technology after removing the interference, and then calculate the whole lung and pulmonary vascular.

RESULTSThe whole lung of the 12 normal volunteers from the apex to the base of lung CT scan image layers was 530 ± 98 (range, 431-841). The total capacity of lung and pulmonary vascular blood was 3705 ± 857 (range, 2398-5383) ml, and the total volume of the pulmonary vascular blood was 125 ± 32 (range, 94-201) ml. The pulmonary vein vascular blood volume was 63 ± 16 (range, 47-100) ml.

CONCLUSIONThe method of measuring the three-dimensional imaging of pulmonary vascular capacity by analyzing lung CT scan data is available and accurate.

BACKGROUND:Currently, there are few reports on the effect of nickel-chromium (Ni-Cr) al oy porcelain-fused-to-metal (PFM) restoration on subgingival flora of abutment.
OBJECTIVE:To investigate the effect of the Ni-Cr al oy PFM restoration on subgingival flora ratio of abutment.
METHODS:Nine patients (12 teeth) who suspected that Ni-Cr al oy PFM could affect their health and therefore came to hospital to ask for removal of the prosthesis were selected in this study. Their subgingival plaques of abutment were obtained before and 1 month, 3 months after the Ni-Cr al oy PFM restorations were removed, respectively, and the changes of subgingival flora were observed and analyzed by the method of denaturing gradient gel electrophoresis.
RESULTS AND CONCLUSION:The images of denaturing gradient gel electrophoresis in subgingival bacteria of experimental group had significant changes at 1 and 3 months after Ni-Cr al oy PFM restorations removed, furthermore, there were significant differences in the images of denaturing gradient gel electrophoresis at 1 and 3 months. In addition, the specific bands were selected from denaturing gradient gel electrophoresis image that appeared before Ni-Cr al oy PFM restorations removed and weakened or disappeared after the removal of restorations, then 16S rDNA sequence in the specific bands were analyzed. The results showed that the gene sequences of these bands were closest related to Eikenel a corrodens, Campylobacter rectus and Eubacterium saphenu. These findings indicated that the Ni-Cr al oy PFM restorations would result in the changes of the proportion of subgingival microflora and increases in the detection rates of some periodontal pathogens.