Objective To study the genotypes of Mycobacterium tuberculosis(M.tuberculosis)strains isolated from Jiangsu province and to explore the relationship between the 'Beijing family' and the drug resistance of M.tuberculosis.Methods Two hundred and sixty M.tuberculosis strains were isolated from 30 drug surveillance sites in Jiangsu province.Susceptibility of the isolates to the first-line antituberculosis drugs(isoniazid,streptomycin,rifampicin and ethambutol)was tested by using the proportion method.Molecular typing of M.tuberculosis strains was determined by Spoligotyping and analyzed with BioNumerics software.Results Based on Spoligotyping fingerprint,260 strains showed 34 different genotypes,including 27 exclusive genotypes and 7 shared genotypes.These strains could be clustered into two groups:the Beijing family(80.4％,209/260)and the Non-Beijing family(19.6％,51/260).Data from logistic regression analysis revealed that infection by the Beijing family was related to an increased risk of multi-drug resistant M.tuberculosis,with the OR(95％ CI)of 11.07(1.45-84.50).Non-Beijing families including T1,T2,H3,H4,CAS,LAM,U and MANU2 families were also found.Among them,the CAS,LAM and MANU2 families were first reported in Jiangsu province.Conclusion It was revealed that the marked gene polymorphisms did exist in M.tuberculosis strains.The Beijing family had been the predominant strain circulating in Jiangsu province,which might be related to multi-drug resistant M.tuberculosis strains.

A cDNA sequence coding for ovine inhibin N terminal 1-33 AA residue fragment (INH) was inserted between BamHI\SacI sites in plasmid pRSET-A to generate plasmid pR-INH. By utilizing a pair of isocaudamer BamHI and Bgl II sites and another downstream Hind III site, following simple double digestions and combination ligation of the resultant products, 2 to 6-repeat INH genes were constructed respectively. Each plamids containing 3 to 6 repeated INH fragment genes, pR-3INH, pR-4INH, pR-5INH and pR-6INH, directed expression of the target proteins in E. coli. BL21 (DE3) under induction of ITPG, which respectively accounted for 6%, 6%, 7% and 8% of the total bacterial protein. The expressed target proteins were all in the form of inclusion bodies. The above results implied that utilization of isocaudamer restriction disgetion sites in expression plasmid is capable of rapidly and correctly constructing repeat fragment polymer of short peptides, which may become a new method in construction of high immunogenic recombinant vaccines of short peptides.
Animals ; Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Inhibins ; biosynthesis ; genetics ; Peptide Fragments ; biosynthesis ; genetics ; Plasmids ; genetics ; Polymers ; chemistry ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; isolation & purification ; Repetitive Sequences, Nucleic Acid ; genetics ; Sheep

OBJECTIVETo investigate the effect of electroporation-mediated gene transfect on the expression of cyclins during mandible distraction in rabbit.

METHODSBilateral mandibular osteotomy was performed in 45 New-Zeland rabbits. After a latency of 3 days, the mandibles were elongated using distractors with a rate of 0.8 mm/day for 7 days. After the completion of distraction, the rabbits were randomly divided into 5 groups. 2 microg (0.1 microg/microl) of pIRES-hVEGF165-hBMP2, recombinant plasmid pIRES-hBMP2, recombinant plasmid pIRES-hVEGF165, pIRES and the same volume of normal saline (NS) was injected into the distraction area in each group, respectively. After injection, electroporation was performed in every group. Three animals in each group were sacrificed at 7, 14, and 28 days after completion of distraction, respectively. The lengthened mandibles were harvested and processed for immunohistochemical examinations. The expression of cyclins A, D1 ,E in positive cells were measured by CMIAS-2001A computerized image analyzer. The data were analyzed with the single factor analysis of variance and q test.

RESULTSCyclins A, D1, E staining was mainly located in inflammatory cells, granulation tissue monocyte, fibroblast, osteoblasts, osteocyte and the connective tissues around the new bone. The expression reached to the peak at 7th day of consolidation, and decreased at 14th day, and weak at 28th day. Image analysis results showed that, at 7th day, the expression absorbance A in group C (0.59 +/- 0.14) was the strongest, compared to group A (0.41 +/- 0.13), B (0.38 +/- 0.14), D (0.34 +/- 0.12) and E (0.31 +/- 0.10), showing a significant difference (P < 0.05, P < 0.01). There was no significance difference between group A and B (P > 0.05), but the difference between group A/B and group D/E (P < 0.05). At 14th and 28th day, there was no significant difference among group A (0.39 +/- 0.11), B (0.34 +/- 0.10) and C (0.33 +/- 0.09) (P > 0.05), but there was significant difference between group A/B/C and group D (0.19 +/- 0.12) or E (0.14 +/- 0.04) (P < 0.05 or P < 0.01).

CONCLUSIONSElectroporation-mediated gene transfection can promote cyclins A, D1, E expression effectively, which may promote cell differentiation and proliferation, stimulate extracellular matrix synthesis and new bone formation in distraction gap.

Animals ; Cyclins ; metabolism ; Electroporation ; Genetic Therapy ; Mandible ; surgery ; Osteogenesis, Distraction ; methods ; Plasmids ; Rabbits ; Transfection

OBJECTIVETo assess the effects of penehyclidine hydrochloride on patients with acute lung injury (ALI), to observe the expression of Toll-like receptor 4 (TLR4) on the peripheral monocytes of ALI patients and changes of inflammatory and anti-inflammatory cytokines and to investigate the mechanism of TLR4 in ALI.

METHODSForty-five patients with ALI were randomly divided into penehyclidine hydrochloride treatment group (P group, n equal to 21) and conventional treatment group (control group, C group, n equal to 24). Patients in both groups received conventional treatment, including active treatment of the primary disease, respiratory support, nutritional support and fluid management therapy, while those in P group were given penehyclidine hydrochloride (1 mg, im, q. 12 h) in addition. The TLR4 expression of 20 healthy volunteers were detected. The clinical effect, average length of stay in ICU and hospital, values of PaO2 and PaO2/FiO2, expression of TLR4 on the surface of peripheral blood mononuclear cells and some serum cytokines were evaluated for 48 h.

RESULTSThe general conditions of the two groups were improved gradually and PaO2 increased progressively. Compared with 0 h, PaO2 and PaO2/FiO2 at 6, 12, 24 and 48 h after treatment were significantly increased (P less than 0.05). The improvement in P group was obviously greater than that in C group (P less than 0.05). The average length of hospitalization showed no difference between the two groups, but penehyclidine hydrochloride significantly decreased the average length of stay in ICU (t equal to 3.485, P less than 0.01). The expression of TLR4 in two groups were both obviously higher than that of healthy volunteers (P less than 0.01). It decreased significantly at 24 h (t equal to 2.032, P less than 0.05) and 48 h (t equal to 3.620, P less than 0.01) and was lower in P group than in C group. The patients who showed a higher level of TLR4 expression in early stage had a worse prognosis and most of them developed acute respiratory distress syndrome (ARDS). The incidence of ARDS was 23.8% in P group and 29.17% in C group at 24 h. Untill 48 h, there were other two patients developing ARDS in control group. Serum IL-1, IL-8 and TNF-alpha expressions reduced after 24 h in both groups. The reduction in P group was more obvious than that in C group (P less than 0.05). IL-13 increased gradually from 0 h to 24 h, and decreased slightly at 48 h, which showed no difference between two groups (t equal to 1.028, P larger than 0.05).

CONCLUSIONSPenehyclidine hydrochloride improves the arterial oxygen pressure, down-regulates the expression of TLR4 and restrains the inflammatory cytokines in the downstream of TLR4 signaling pathway. It prevents the development of ALI and can be considered as an important drug in ALI treatment.

Acute Lung Injury ; drug therapy ; etiology ; physiopathology ; Cytokines ; blood ; Heart Rate ; drug effects ; Humans ; Oxygen ; blood ; Prognosis ; Quinuclidines ; therapeutic use ; Toll-Like Receptor 4 ; genetics ; physiology

To establish a bioassay method and quality standard of Banlangen granula, agglutinated activity assay was used in the analysis of the traditional Chinese medicine, Banlangen granula. It showed that masculined effect could be picked up effectively and the products quality of different pharmaceutical factories and different batch numbers from the same factory could be revealed conveniently, accurately, quickly and directly with this method (valence value was between 2 and 11). The established bioassay method had a good reproducibility with RSD = 2%. The dependablity of the activity of red cell agglutination and restrainting influenza virus NA was conspicuous (r2 = 0.878 3). In conclusion, this bioassay method is suitable to control and evaluate the quality of Banlangen granula. Thus the method may provide a simple and effective technique in supervising and examining the quality of other traditional Chinese medicine.
Animals ; Antiviral Agents ; administration & dosage ; isolation & purification ; pharmacology ; standards ; Biological Assay ; Dosage Forms ; Drugs, Chinese Herbal ; administration & dosage ; isolation & purification ; pharmacology ; standards ; Hemagglutination ; drug effects ; Male ; Neuraminidase ; metabolism ; Orthomyxoviridae ; drug effects ; enzymology ; Plants, Medicinal ; chemistry ; Quality Control ; Rabbits ; Reproducibility of Results

A biothermal activity detection method has been established to determine the potency of colistin. The biothermal activity fingerprints of E. coli with colistin were determined. There was a good linear relationship (r = 0.993) between logarithm concentration of colistin (lgC) and lag rate of growing time (Deltat%) when the concentrations of colistin ranged from 17.0 to 41.6 u x mL(-1). The average recovery rate was 100.3% (n = 9). Using this method, there was no significant difference between results of colistin potency measurement and those using cup-plate method (P > 0.05). As a result, biothermal activity detection method is sensitive, accurate, rapid, convenient and feasible to determine the potency of colistin. This method can also be applied in real time and online to monitor the process of bacterial growth and could be complementary to the cup-plate method.
Anti-Bacterial Agents ; administration & dosage ; pharmacology ; Calorimetry ; methods ; Colistin ; administration & dosage ; pharmacology ; Dose-Response Relationship, Drug ; Escherichia coli ; drug effects ; growth & development ; Microbial Sensitivity Tests ; methods ; Thermodynamics

OBJECTIVETo assess the differences between the main parameters obtained by sperm quality analyzer V (SQA-V) and computer-aided sperm analysis system (CASA), and to investigate their application to sperm quality analysis for fertile and infertile men.

METHODSTwelve fresh semen samples from fertile volunteers and 73 from infertility patients were detected with SQA-V and CASA for sperm concentration and motility, the percentage and concentration of motile sperm, sperm motility index (SMI), amplitude of lateral head displacement (ALH), beat cross frequency (BCF), curvilinear velocity (VCL), straight line velocity (VSL), average path velocity (VAP), linearity (LIN = VSL/VCL) and straightness (STR = VSL/VAP). The correlation between the parameters obtained by the two devices were analyzed.

RESULTSSignificant differences were observed in the above parameters between the fertile and infertile groups. An obvious consistency was noted between the results from SQA-V and those from CASA in sperm concentration (r = 0.58, P < 0.01), motile sperm concentration (r = 0.75, P < 0.01) and average sperm velocity (r = 0.59, P < 0.01). Significant correlations were found between the SMI from SQA-V and STR, LIN, BCF, VCL, VSL and VAP from CASA (P < 0.05).

CONCLUSIONThere is a consistency between the results from SQA-V and those from CASA. Both the devices can detect the seminal differences between different cohorts of patients.

Adult ; Diagnosis, Computer-Assisted ; Humans ; Infertility, Male ; diagnosis ; Male ; Semen Analysis ; methods ; Sperm Count ; Sperm Motility

BACKGROUNDMicroRNAs (miRNAs) function as essential posttranscriptional modulators of gene expression, and are involved in a wide range of physiologic and pathologic states, including cancer. Numerous miRNAs are deregulated in hepatocellular carcinoma (HCC). This study aimed to investigate the role of miR-27a in the development of HCC.

METHODSThe expression of MiR-27a was measured by quantitative real-time polymerase chain reaction (qRT-PCR). 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide was used to examine changes in the viability of HepG2, Bel-7402, Bel-7404 hepatoma cell lines associated with up-regulation or down-regulation of miR-27a. A dual-luciferase activity assay was used to verify a target gene of miR-27a. Immunohistochemistry, qRT-PCR, Western blotting analysis, and cell cycle and apoptosis flow cytometric assays were used to elucidate the mechanism by which miR-27a modulates liver cancer cell proliferation.

RESULTSThe expression of miR-27a was significantly increased in HCC tissues and HepG2, Bel-7402, Bel-7404 hepatoma cell lines (P < 0.05). We also found that the down-regulation of miR-27a in HepG2 cells dramatically inhibited proliferation, blocked the G1 to S cell cycle transition and induced apoptosis (P < 0.05). In addition, miR-27a directly targeted the 3'- untranslated region of peroxisome proliferator-activated receptor γ (PPAR-γ), and ectopic miR-27a expression suppressed PPAR-γ expression on the mRNA and protein levels. The rosiglitazone-induced overexpression of PPAR-γ attenuated the effect of miR-27a in HCC cells.

CONCLUSIONSOur findings suggested that miRNA-27a promoted HCC cell proliferation by regulating PPAR-γ expression. MiR-27a may provide a potential therapeutic strategy for HCC treatment.

Carcinoma, Hepatocellular ; genetics ; metabolism ; Cell Proliferation ; genetics ; physiology ; Gene Expression Regulation, Neoplastic ; Hep G2 Cells ; Humans ; Liver Neoplasms ; genetics ; metabolism ; MicroRNAs ; genetics ; physiology ; PPAR gamma ; metabolism

Plant virus diseases are one of the major diseases restricting erop production.Timely identification of their pathogen and development rules is the prerequisite for effective control of their large- scale spread.However, long cycle, tedious steps and strict detection environment were the disadvantages existing in the detection technology of plant virus disease.In this study, Tobacco Mosaic virus (TMV) was used as a model to he extract UNA based on CMBs-ACPtmv , which was design based on the principle of complementary base pairing.Meanwhile, the experimental conditions were optimized and analyzed, including the preparation conditions of functionalized magnetic beads, the reaction conditions during extraction, and the sensitivity, stability and other properties of the method.The results showed the ability to capture RNA of CMBs-ACPtmv were best when prepared with 4 fxmol capture probe (ACPTMV ) and 0.08 mg carboxyl magnetic beads (CMBs) ; After 3 min of extraction, CMBs-ACPtmv has the best RNA extraction effect, but when the extraction temperature of CMBs-ACPtmv was changed, its extraction capacity showed no significant change; In the comprehensive performance evaluation, the sensitivity of CMBs-ACPjjjv can reach 2.5 ng/fxL, and the detection stability is good.Compared with conventional RNA extraction technology, CMBs-ACPimv has outstanding advantages in detection time and sample consumption.The functional magnetic beads extraction method established in this study is fast, safe and simple.It can achieve rapid extraction of plant virus RNA with simple equipment, which has a broad application prospect.

OBJECTIVE: To explore the influencing factors of low back pain and the relationship of the influence of bad working posture, weight load and frequency of load and the dose-response relationship among the occupational workers of key industries in China. METHODS: A total of 57 501 employees from 15 key industries in China were selected as research subjects using stratified cluster sampling method. The occurrence of low back pain in the past one year, as well as occupational factors such as job type, labor organization and work posture were investigated by using the Chinese version Musculoskeletal Disorders Questionnaire. RESULTS: The prevalence of low back pain in the occupational population of key industries in China was 16.4%(9 448/57 501). Multivariate Logistic regression analysis showed that the risk of low back pain in females was higher than that in males(P<0.01). Married, obese, occasional and frequent smokers, and a history of lower back disease were associated with increased risk of low back pain(all P<0.05). The risk of low back pain was associated with older age, higher education level, and lower frequency of physical exercise(all P<0.01). The risk of low back pain was higher with longer working time, greater back curvature, and the high frequency of long standing and sitting position work, uncomfortable working posture, repeated operation per minute, and lifting>5 kg weight(all P<0.01). CONCLUSION: The influencing factors of low back pain in the occupational population of key industries in China include bad working posture, high frequency load, weight load and other individual factors. There is a dose-response relationship with low back posture load and frequency of load.