The B.subtilis ywtD gene,encoding a ?-polyglutamic acid(?-PGA)depolymerase,was amplified from the genome of B.subtilis NX-2 by PCR.The comparability between the cloned ywtD gene sequence to the reported sequence is high to 99.0%.Only one of the substituted nucleotide base caused the change to the amino acid sequence.The recombinant plasmid pET-15b-ywtD was then transformed into E.coli Rosetta(DE3)and the ywtD gene product could be expressed with the induction of 0.5mmol/L IPTG.The YwtD protein exhibited a remarkable activity in ?-polyglutamic acid degradation.The molecular weight of ?-PGA could be reduced from 700kDa to 20kDa after 72h through the enzymatic hydrolysis and consequently trended to be constant.

Objective The change of thyroid hormones affects male sexual function, but the underlying physiological mechanism is not yet clear.The aim of this study was to explore the association of thyroid hormones with abnormal penile neuro-electrophysiology in patients with premature ejaculation (PE).Methods This retrospective study included 52 acquired PE (APE) patients aged 35.36 ± 8.39 years and 24 healthy males aged 32.87±7.45 years.We measured the levels of thyroid hormones in all the subjects and, based on the latency time of penile sympathetic skin responses (PSSR), classified the APE patients into a normal (n=31) and an abnormal PSSR type (n=31).Results The concentration of total tetraiodothyronine (TT4) in the serum was significantly higher in the APE patients than in the normal control ([102.81±18.37] vs [92.68±11.56] nmol/L, P=0.018).The levels of serum TT4 and free tetraiodothyronine (FT4) were both markedly increased in the APE patients of the abnormal PSSR type as compared with those of the normal PSSR type ([113.28±20.89] vs [95.72±12.42] nmol/L, P<0.001 and [12.02±0.88] vs [10.81±1.63] nmol/L, P=0.003).The FT4 concentration was correlated negatively with the latency time of PSSR (r=-0.363, P=0.008) and positively with the Premature Ejaculation Diagnostic Tool (PEDT) scores (r=0.455, P=0.001).Conclusion Abnormal PSSR may be associated with elevated levels of thyroid hormones in APE patients, and regulation of thyroid hormone levels may help improve the symptoms of PE with abnormal PSSR.

Background One of the features of the pathogenesis of primary angle-closure glaucoma(PACG) is anterior synechia of peripheral iris.Goniosynechialysis and combination of phacoemulsification and goniosynechialysis have been applied for the treatment of the disease recently,but the selection of operative types has great impact on clinical efficacy.Objective This study was to investigate the long-term efficacy of goniosynechialysis and combination of phacoemulsification and goniosynechialysis for the management of chronic PACG.Methods A non-randomized clinical controlled trial was designed.This clinical trial complied with Declaration of Helsinki and was approved by Medical Ethic Committee of Nanjing Medical University.Written informed consent was obtained from each patient.One hundred and ten eyes of 110 patients with chronic PACG were assigned to the goniosynechialysis group and combined operative group from March,2008 to February,2011 in Wuxi People's Hospital.180° goniosynechialysis was performed on 34 patients in the goniosynechialysis group,and phacoemulsifieation +intraocular lens (IOL) implantation + goniosynechialysis were carried out in 78 patients of the combination operative group.All the patients were followed-up for 2 years.Vision acuity,intraocular pressure (IOP),anterior chamber depth(ACD),unltrasound biomicroscopy and perimetry were recorded and compared between before and after operation.Results No significant difference was found in vision acuity between preoperation and postoperative 2 years in the goniosynechialysis group ([0.65 ± 0.15] vs.[0.45 ± 0.15]) (t =1.57,P＞0.05),but in the combination operative group,the vision acuity was significantly different between the before and after operation ([0.25±0.15] vs.[0.85 ±0.05]) (t =9.12,P＜0.001).The lOPs at 2 years after operation were (14.2±4.1) mmHgand(13.7±4.8) mmHg,respectively in the goniosynechialysis group and combination operative group and were significantly lower than(47.2 ±6.3) mmHg and(46.9±7.0) mmHg before operation(t =4.95,P＜0.001 ;t=5.03,P＜0.001).The ACD values in the goniosynechialysis group and combination operative group were(3.38±0.02)mm and (3.54±0.03) mm 2 years after operation,which were significantly increased in comparison with (1.33 ±0.24)mm and (1.56±0.37) mm before operation(t=7.65,P＜0.001;t=6.76,P＜0.001).Conclusions Both combination of phacomulsification with goniosynechialysis or 180° goniosynechialysis are effective for the treatment of PACG.Suitable operation should be alternated depending on the indicators of PACG patients.

Objective:To clone the human cell death-inducing DFF45-like effectors 3(CIDE3) full length cDNA for construction the eukaryotic expression vector pcDNA3.l(+)-CIDE3 and to study its bioactivity.Methods:① Total RNA was extracted from human white adipose tissues and the desired cDNA fragment was obtained by RT-PCR.After the fragment had being inserted into an eukaryotic expression vector pcDNA3.l(+),the resulted recombinant plasmid pcDNA3.l(+)-CIDE3 was transformed into DH5?.The positive clone was selected and confirmed to contain full length of CIDE3 cDNA by agarose gel and DNA sequence analysis.②After the pcDNA3.l(+)-CIDE3 plasmid was transfected into HeLa cells under lipofectamine mediation,the effect of target gene expression on growth of HeLa cells was analysed by TUNEL staining. Results:① The recombinant eukaryotic expression plasmid pcDNA3.l(+)-CIDE3 was constructed successfully and the sequence of CIDE3 was consistent with that of genebank.②After transfected pcDNA3.l(+)-CIDE3,HeLa cells presented distinguished apoptosis(about 15%),compared with that of transfected plasmid pcDNA3.l(+)(

BACKGROUND:Development of new coronary thrombolytic agents is hot in the market. A new drug, mutated recombinant tissue-type plasminogen activator (rtPAm), is the product of mutation of tPA by changing binding loci with plasminogen activator inhibitor (PAI)-1 to reduce the degradation. In vitro test has demonstrated that the activity of rtPAm is much higher than rtPA in the absence of PAI. The present study is to observe the efficacy of mutated recombinant tissue-type plasminogen activator (rtPAm) in coronary thrombolytic therapy. METHODS:A total of 30 adult beagles were equally divided into 5 groups after thrombi:vehicle group, urokinase group, rtPAm low-dose group, rtPAm medium-dose group, and rtPAm high-dose group. Thrombolytic effect and myocardial infarction were observed after thrombolytic therapy. RESULTS:In the urokinase group, time to reperfusion was (15.8±3.8) minutes. TIMI 2 flow was demonstrated in 4 beagles, TIMI 3 flow in 2, and re-occlusion in 4 after 90 minutes respectively. In the low-dose rtPAm group, time to reperfusion was (15±4.5) minutes; TIMI 2 flow was demonstrated in 2 beagles, TIMI 3 flow in 4, and re-occlusion in 2 after 90 minutes. In the high-dose rtPAm group, time to reperfusion was (7.5±2.6) minutes. None of the beagles showed re-occlusion after 90 minutes. The infarction areas were (2.1+0.9)％ in the medium-dose rtPAm group and (0.7+0.4)％ in the high-dose rtPAm group, which decreased significantly than those in the low-dose rtPAm group. The aggregation rate in the medium-dose and high-dose rtPAm groups decreased significantly than that in the urokinase group. CONCLUSION:rtPAm may serve as a thrombolytic agent with platelet-targeted fibrinolysis and antiplatelet aggregation activities.

In this paper, the chloroplast psbK-psbI intergenic spacers of 18 species of Dendrobium and their adulterants were amplified and sequenced, and then the sequence characteristics were analyzed. The sequence lengths of chloroplast psbK-psbI regions of Dendrobium ranged from 474 to 513 bp and the GC contents were 25.4%-27.6%. The variable sites were 71 while the informative sites were 46. The inter-specific genetic distances calculated by Kimura 2-parameter (K2P) of Dendrobium were 0.006 1-0.058 1, with an average of 0.028 4. The K2P genetic distances between Dendrobium species and Bulbophyllum odoratissimum were 0.093 2-0.120 4. The NJ tree showed that the Dendrobium species can be easily differentiated from each other and 6 samples of the inspected Dendrobium species were identified successfully through sequencing the psbK-psbI intergenic spacer. Therefore, the chloroplast psbK-psbI intergenic spacer can be used as a candidate marker to identify Dendrobium species and its adulterants.
Chloroplasts ; DNA, Chloroplast ; genetics ; DNA, Plant ; genetics ; DNA, Ribosomal Spacer ; genetics ; Dendrobium ; classification ; genetics ; Plants, Medicinal ; classification ; genetics

In order to identify Peucedani Radix, Peucedani Decursivi Radix and their adulterants, the internal transcribed spacer 2 (ITS2) regions of Peucedani Radix, Peucedani Decursivi Radix and their adulterants were amplified and bidirectionally sequenced based on the Principles for Molecular Identification of Traditional Chinese Materia Medica Using DNA Barcoding, which has been promulgated by Chinese Pharmacopoeia Commission. Sequences were analyzed and assembled by Codon Code Aligner V3. 7.1. The relevant data were analyzed by MEGA 5. 0. Species identification analyses were performed by using the nearest distance methods and neighbor-joining (NJ) methods. The result showed that the ITS2 sequence lengths of Peucedani Radix were 229-230 bp and the average intra-specific genetic distances were 0.005. The ITS2 sequence lengths of Peucedani Decursivi Radix were 227 bp and the sequences contained no variation site. The average inter-specific K2P genetic distance of Peucedani Radix, Peucedani Decursivi Radix and their adulterants species were 0.044 and 0.065 respectively. The minimum inter-specific divergence is larger than the maximum intra-specific divergence of Peucedani Decursivi Radix. The nearest distance methods and NJ trees results indicated that Peucedani Radix, Peucedani Decursivi Radix and their adulterants species could be identification clearly. The ITS2 regions can stably and accurately distinguish Peucedani Radix, Peucedani Decursivi Radix and their adulterants.
Apiaceae ; classification ; genetics ; DNA Barcoding, Taxonomic ; methods ; DNA, Ribosomal Spacer ; Drug Contamination

Objective To investigate the value of two-dimensional speckle tracking imaging (STI) in assessing regional myocardial dysfunction and myocardial fibrosis in a mouse model of myocardial infarction.Methods Twenty C57/B6 mice were randomly divided into two groups:myocardial infarction (MI) group (n =10) and sham-operation (SO) group (n =10).Echocardiography was performed four weeks after surgery.High frame rate two dimensional images were recorded in the left ventricular short axis views at the papillary muscle level and analysised at EchoPac workstation.Peak radical strain (PRS) and peak radical strain rate (PRSR) of each segment were measured at systolic period.Percent segmental fibrosis (PSF) was assessed from histological left ventricular cross sections stained by Masson trichrome.Results Compared with SO group,PRS and PRSR decreased significantly in all segments of MI group (P ＜ 0.01),especially in anterosepetal,anterior and lateral segments (P ＜0.05).PSF of anterosepetal,anterior,lateral and posterior segments in MI group increased significantly than those in SO group(P ＜0.01),and were negatively correlated with PRS and PRSR of these segments(r =-0.88,P ＜0.001 ; r =-0.77,P ＜0.001).Conclusions STI could accurately quantify regional myocardial function in a mouse model of myocardial infarction.Segmental radial strain and strain rate measured by STI were significantly correlated with PSF,which can be a non-invasive tool for monitoring myocardial fibrosis after myocardial infarction.

Objective To investigate the relationship between SOCS-3 gene polymorphism and abnormal glucose metabolism in Xinjiang Uygur population.Methods According to different plasma glucose levels,1232individuals in Xinjiang Hetian area were divided into 3 groups,451 patients with pre-diabetes,252 patients with type 2 diabetes mellitus,and 529 healthy people as normal controls.Based on HapMap,the polymorphisms rs12953258,rs4969168,rs9914220 were selected as haplotypes,tagging SNP (htSNP) sufficiently covering the genetic variation of the whole gene.Associations of rs12953258,rs4969168,and rs9914220 within SOCS-3 with abnormal glucose metabolism in three groups of Xinjiang Uygur population were examined ; The genotype and allele frequencies and relative clinic data were compared among groups.Results The type 2 diabetes mellitus study with 451 individuals showed that the homozygosity for the C allele of rs12953258 polymorphism of SOCS-3 was associated with increased diabetes risk(OR=1.756,95％ CI 1.168-2.640).In addition,association between rs4969168 or rs9914220 and abnormal glucose metabolism in the Xinjiang Uygur population was not found.Age,total cholesterol,and body mass index were risk factors of diabetes mellitus,total cholesterol and low high density lipoprotein-cholesterol were risk factors of pre-diabetes in Uygur people.Conclusions The C allele of rs12953258 polymorphism of SOCS-3 gene may be an independent risk factor for abnormal glucose metabolism in Xinjiang Uygur population.

Objective To evaluate the imaging quality of the non-contrast enhanced MR angiography of sampling perfection with application optimized contrasts using different flip angle evolutions (SPACE)in showing portal system and compared it with that of the contrast enhanced MR angiography of volumetric interpolated breath-hold examination (VIBE),and study its diagnostic ability in the detection of portosystemic and portohepatic collaterals.Methods Thirty consecutively cirrhotic patients with suspected of portosystemic and portohepatic collaterals were enrolled,and underwent SPACE followed by VIBE at 1.5 T MR scanner.The diagnostic accuracy of SPACE for the portal vein disease was evaluated by two doctors and compared it with that of VIBE.The contrast-to-noise ratio(CNR) and signal-to-noise ratio(SNR) of two MRA techniques were compared by using the Wilcoxon signed rank test.The quality assessment including scores of the portal vein segments and overall image quality were used the paired t test.Results Twenty-one patients were diagnosed as portal hypertension,including five types of portosystemic collaterals: esophageal varices (n =5),gastric fundic varices (n =11),splenic varices (n =5),paraumbilical varices (n =5) and cavemous transformation (n =2),and one patient was diagnosed as portal vein tumor thrombus.The diagnostic efficiency of SPACE was equivalent to that of VIBE.In SPACE,the SNR were 291 ± 57,301 ± 74,344 ±76 and the CNR were 231 ±59,242 ±73,286 ±76 at main portal vein,the left branch of portal vein and the right branch of portal vein,respectively.However in VIBE,the SNR were 185 ± 56,176 ± 52,182 ±52 and the CNR were 57 ±23,50 ±21,57± 19 at,respectively.Both SNR and CNR of portal vein segments in the former were better than those in the latter (t values were 7.691,7.418,7.946,15.746,13.508 and 13.880,respectively,P ＜ 0.05).There were no significant difference for the scores of displaying main,left branch and right branch portal vein and overall image quality in VIBE and SPACE (Z values were -1.496,-1.895,-1.496,-2.138,-2.324 and-1.328,respectively,P ＞ 0.05).The scores of displaying the distal branches of left and right portal vein were 2.08 ± 0.78,2.08 ± 0.78 in SPACE,and 1.75 ± 0.53,1.71 ± 0.55 in VIBE,respectively.It was better (Z =-2.138,-2.324,P ＜ 0.05) in SPACE than that in VIBE.Conclusion The SPACE has better visualization of portal vein distal branches than VIBE,and it can be applied for the diagnosis of the portal vein disease.