Purpose To clone and express Trypanosoma Leucyl-tRNA synthetase (LeuRS) gene and to complete the purification and activity assay of LeuRS. Methods We cloned the LeuRS gene from T. Brucei genomic DNA,and cloned it into pBS-T and then into the expression vector pET21a( + ) .The expression of T. Brucei LeuRS was carried out in E. coli BL21 (DE3) RIPL host by optimizing the expression conditions. The products are purified by His-Bind affinity column and verified by Western blot . Enzymatic activity was detected by isotope labeling. Results A DNA fragment at length of 3.2 kb was amplified. Both restriction analysis and sequencing analysis proved that recombinant plasmid pET21a( + )/LeuRS was correctly constructed. The expressed product contained about 20% of total somatic soluble protein and reached a purity of 85% after purification . It was verified by the Western blot. Enzyme unit per millilitre of purified products was about 72. Conclusion Here we report the successful clone, expression and purification of LeuRS from Trypanosoma Brucei ( T. Brucei) in bacterial host. In addition, we tested its enzymatic activity by isotope labeling.This work would be helpful to the design and in vitro screening of Trypanosoma LeuRS inhibitors.

Objective To explore the value of intraoperative contrast-enhanced ultrasonography in radiofrequency ablation completeness evaluation in patients with hepatic carcinoma of differently sized nodules.Methods Fifty five patients (69 lesions) with hepatic carcinoma were included and were divided into the experimental group (20 cases,30 lesions in whom CEUS were used) and the control group (35 cases,39 lesions,conventional ultrasound was used).After RAF,the treatment effect of the two groups were evaluated by enhanced CT regularly.Differences between conventional ultrasound and CEUS were compared and their judgement on ablation residual tumor tissue was analyzed.Results After 1 to 24 months of follow-up,the total residual rate of the experimental group and the control group was 13.3％ (4/30) and 35.9％ (14/39) respectively.The significant difference was found in the residual rate between the experimental group and the control group (x2 =5.366,P ＜ 0.05).For the two groups (＞ 5 cm and ＜3-5 cm),the residual rate of the experimental group was 30.0％ and 10.0％,the residual rate of the control group was 87.5％ and 46.2％,there were statistically significant difference (respectively x2 =5.951,4.790,all P ＜ 0.05).The significant difference was found in the recurrence rate (the experimental group:20.0％ (4/30),the experimental group:75.0％ (6/8)),when the lesion was larger than 5 cm in diameter(x2 =5.445,P ＜ 0.05).Conclusions CEUS has advantage over conventional ultrasound for the detection of residual tumor tissues after radiofrequency ablation for hepatic carcinoma,it reduces after ablation tumor recurrence especially in large tumors.

AIM:To establish a HPLC fingerprint of Stellera chamaejasme L. from Inner Mongolia Region. METHODS: The RP-HPLC method was used with Akzonobel Kromasil C_ 18 (250 mm?4.6 mm, 5 ?m) the acetonitrile-0.5% phosphoric acid (gradient elution) was used as mobile phase, analytic time was 60 min, and detective wavelength was at 297 nm, the column temperature of 15℃ were adopted. RESULTS: The HPLC fingerprint of Stellera chamaejasme L. set up showed that 14 peaks were co-possessing in different sources. The results of method validation met technical standard of fingerprint, the similarities of Stellera chamaejasme L. were 0.9 to 1.0. CONCLUSION: The method is stable and reliable with a good reproducibility and provides a reference standard for the quality control of Stellera chamaejasme L. from Inner Mongolia Region.

Objective: In T_(739) inbred line mice,bladder cancer model was established via tumor source transplant.In order to get insights of the biological effects of implanting ~(125)I seed to treat bladder cancer,the development and the pathological modality of tumor tissues were observed through the experiment of implanting ~(125) I seed.The experiment was to provide theoretical gist and practical guidance for clinical(utilization) of ~(125)I seed for brachytherapy of bladder cancer. Methods: 16 T_(739) inbred line mice were divided into two groups(A:experimental group,n=8;B:control group,n=8) in random;bladder cancer model was established via tumor source transplant.When the diameter of the transplanting tumor was about 0.7 cm,~(125)I seeds were implanted at the tumor of experimental group while the empty seed was penetrated into the control group at the same position as the experimental group and the experimental mice were executed after 24 days.The longest vertical diameters(a) and(b) of tumor body were observed every 3 days and the volume of tumor body(V=ab~(2)/2) was calculated.Combining with the weight of mouse(W) and the weight of tumor body after execution of mice(w),the growth curve of tumor was protracted and the colony redouble time and tumor suppression rate were calculated.After fixation of formaldehyde,the pathological modality of tumor tissues were observed with HE staining. Results: ① The transplantation was 100% successful and there was no natural fade-away phenomenon.②After implantation of seed,the weight of group A mice increased by(4.93?0.60)g,and group B increased by(13.60?1.15)g(P

0.05),but it was associated with the depth of invasive cervical and lymph node metastases(P

OBJECTIVEStudy the current quality management situation of enterprises marketing corneal contact lens via systemic investigations and explore effective administration countermeasures in the future.

METHODSThe quality management indicators of sixty-two corneal contact lens marketing enterprises in Xuhui district of Shanghai were systematically investigated and enterprises of different operation models was compared and analyzed.

RESULTSWholesale enterprises and retail chain enterprises are apparently better than independent enterprises almost in all facets.

CONCLUSIONFacilitate market accession of corneal contact lens marketing enterprises, encourage the business model of retail chain, enhance supervision of corneal contact lens marketing enterprises, especially independent franchisors.

Trypanosoma is a human parasite severely affecting poor tropical areas.However,current frontline drugs for Trypanosoma treatment have severe side-effects with decreased effectiveness.Based on the fact that aminoacyl-tRNA synthetase is a bonafide drug target for several microorganisms,including bacteria and fungi,it is plausible that it may also be effective target of Trypanosoma.The Trypanosoma brucei leucyl-tRNA synthetase(tbLeuRS)was cloned,expressed and purified to develop an in vitro enzymatic assay system.The assay conditions were further optimized for the effective screening of tbLeuRS inhibitors thus establishing an anti-Trypanosoma drug screening system targeting tbLeuRS.The results indicated that this system can be employed for the effective screening of anti-Trypanosoma drugs with satisfactory specificity.In addition,this system can also be used for compound optimization,as well as IC50 testing.Using this system a series of compounds are identified that are effective Trypanosoma inhibitors without toxicity to human cells.Therefore,targeting tbLeuRS may represent a new venue for the development of anti-Trypanosoma drugs.

Objective To investigate the relationship between the level of serum uric acid(SUA) and carotid atherosclerosis in male patients with type 2 diabetes mellitus(T2DM). Methods A collection of 579 male T2DM patients with or without carotid atherosclerosis were grouped based on quartiles of SUA. Age, SUA, smokers, duration, body mass index(BMI), blood pressure, total cholesterol(TC), triglyceride(TG), high-density lipoprotein cholesterol(HDL-C), low-density lipoprotein cholesterol(LDL-C), blood urea nitrogen(BUN), creatinine(Cr), and HbA1C were determined in all subjects. The plaques in carotid arteries and intima-media thickness(IMT) were measured with high-resolution ultrasound. Results BMI, systolic and diastolic blood pressures, TG, and Cr showed a gradual increase, while HDL-C and HbA1C showed a gradual decrease according to the higher SUA quartiles in male T2DM(P＜0.05). Nevertheless, the detectable rates of smokers, duration, age, TC, LDL-C, and BUN had no relationships with the SUA quartiles(P＞0.05). The detectable rate of carotid atherosclerosis and the thickness of carotid plaque were positively associated with the levels of SUA in male patients with T2DM(P＜0.05). However, intima-media thickness of carotid arteries did not illustrate the correlation with the levels of SUA in male T2DM patients(P＞0.05). Age, HbA1C, and SUA were independent factors of carotid atherosclerosis in these patients by logistic regression(P＜0.05). Conclusion The levels of SUA seems to be associated with the occurrence and development of carotid atherosclerosis in male patients with T2DM.

Objective To analyze the clinical characteristics of autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC), primary sclerosing cholangitis (PSC) and AIH/PSC or/PBC overlap syndrome in order to further understand the autoimmune liver diseases (AILD). Methods One hundred and nine patients with AILDs confirmed pathologically were collected between Jan. 2004 and June 2006. Of 109 patients, AIH was found in 27 eases, PBC in 67 cases, PSC in 4 eases, AIH-PSC overlap syndrome in 1 case and AIH-PBC overlap syndrome in 10 cases. The clinical and laboratory data of all patients were assessed retrospectively. Results The AILD was predominantly found in middle-aged women (73.3% ,80/109), and the main clinical manifestations were jaundice, malaise, anorexia and pruritus. The age distribution of patients with AIH showed a single peak at 50 years. Elevated serum gamma globulin and IgG were found in patients with AIH, of whom 62.9% (17/27) were positive for anti-nuclear antibody (ANA) and 3 were positive for liver-kidney microsomes type 1 antibody. The main histological changes in severe AIH cases included interface hepatitis (77.7 %) and bridging necrosis. Most of the PBC patients were presented with elevated serum alkaline phosphatase, glutamyl transpeptidase and IgM. Fifty patients (74.6%) were positive for anti-mitochondrial antibody (AMA) and AMA-M2. The pathological examination showed that 28. 3% of the cases were in Ⅰ or Ⅱ stage and 71.7% in Ⅲ or Ⅳ stage in patients with PBC who received liver biopsy. The pathologic change of reduction or even disappearing of bile ducts was found in 62. 6% patients with PBC. The clinical and pathological manifestations in patients with AIH-PBC overlap syndrome had both characteristics of PBC and AIH. Three out of 10 patients with AIH-PBC overlap syndrome were positive for ANA and AMA/AMA-M2. Conclusion Since AILD is not rare in Chinese, its diagnosis should be based on the clinical presentation, biochemical, immunological and histologic changes.

Objective To make a comparison for the neutrophils prepared either by induction of differentiation of myeloid leuke-mia cell line,or by separation and purification of peripheral blood cells,or by induction of myeloid differentiation of peripheral blood stem cells.Methods NB4 cells were induced differentiation by 1μmol/L all-trans retinoic acid (ATRA)to mature granulo-cytes;neutrophils were separated and purified from peripheral blood by lysis of red blood cells followed by negative selection using magnetic bead-labeled antibodies;hematopoietic stem cells were separated and purified from peripheral blood by Percoll gradient centrifugation followed by negative selection using magnetic bead-labeled antibodies,and were induced to myeloid differentiation by GM-CSF and G-CSF.Morphology and purity of neutrophils prepared by these three methods were studied by means of MGG stai-ning.CD18 protein expression and subcellular distribution were studied by means of immunofluorescence staining.Results Purity of neutrophil was above 40% by induction of differentiation of NB4 cells,and was about 90% if purified from peripheral blood,and was above 70% if induced by myeloid differentiation of peripheral blood stem cells.There was no obvious difference for CD18 ex-pression in neutrophils prepared by these three methods,and staining of CD18 had a dotted pattern distributed in these cells.Con-clusion Peripheral blood neutrophils prepared by lysis of red blood cells followed by negative selection using magnetic bead-labeled antibodies are with high purity and viability which is suitable for immediate test of neutrophils from fresh blood.Neutrophils pre-pared by myeloid differentiation of hematopoietic stem cell are with high viability and last for days,which can be used in long test for neutrphils.