Hirudin (HV) is known as the most potent and specific inhibitor of thrombin. Although hirudin has many advantages , it has the bleeding side effect and this is the great shortage of hiudin for clinical application. In order to alleviate bleeding side effect of hirudin, fusion protein, named as FHV (fusion hirudin linked with FXa recognition peptide) was designed. The fusion protein gene ( fhv) was cloned into plasmid pPIC9K. FHV engineered Pichia pastoris containing high copies was chosen for fermentation and purification at 30 L fermentor scale, finally, FHV with purity of above 97% was obtained. To investigate the function of FHV in vivo, mouse tail thrombosis model was used. In the mice thrombus tail model induced by carrageenan, FHV decreased the length of tail thrombus significantly, similar to that of HV control, and had no obvious effects on the TT, PT and APTT. In conclusion, FHV is constructed and expressed in yeast. FHV fusion proteins is obtained by fermentation and purification. FHV has antithrombotic effects not influencing IT, PT and APTT after administration immediately in animal models. Therefore, FHV is a promising anticoagulant and antithrombotic drug.

Objective To investigate the molecular epidemiology of methicillin-resistant Staphylococcus aureus in China in 2005.Methods From January to December 2005,395 consecutive and non-repetitive isolates of methicillin-resistant Staphylococcus aureus were collected from 17 teaching hospitals in 14 cities.The genotypes of SCCmec were determined by multiplex PCR pulsed-field gel electrophoresis (PFGE)was used to type the chromosome DNA of MRSA.Muhilocus sequence typing(MLST)was used to type the housekeeping genes.Fifty-three strains were selected for MLST typing according to the antimicrobials susceptibility patterns,PFGE types,SCCmec types and the distribution of the regions.The toxin gene was detected by PCR.Results Among 395 isolates of MRSA,SCCmec Ⅲ,untypeable type and type Ⅱ accounted for 61.5%(243/395),24.3%(96/395)and 14.2%(56/395)respectively.In Shenyang,60.7%(17/28)of the isolates were SCCmec Ⅱ,which was significantly higher than other areas. Twenty-four different types and 42 subtypes were found by PFGE typing.Clone A accounted for 50.1%, existing in 13 teaching hospitals in 12 cities and clone R accounted for 23.5%,existing in 9 teaching hospitals in 8 cities.Six sequence types(ST)were found in these isolates with ST239 and ST5 accounting for 75.5% and 17.0% among these isolates,respectively.The prevalence of pvl gene was 2.5% among 395 isolates of MRSA.Conclusions The most types of SCCmec in China were Ⅲ and Ⅱ,and distribution of SCCmec types differed among regions.MRSA outbreaks caused by epidemic multiple-drug resistant clones occurred in big teaching hospitals in China.Meanwhile,the same PFGE pattern may spread among areas. Several international epidemic MRSA clones may exist in China.

Idiopathic pulmonary fibrosis (IPF) is characterized by myofibroblast foci in lung parenchyma.Myofibroblasts are thought to originate from epithelial-to-mesenchymal transition (EMT).Wnt1 and lithium chloride (LiCl) induce EMT in alveolar epithelial cells (AECs),but the mechanisms are unclear.AECs were treated with Wnt1 and LiCl,respectively;morphological change and molecular changes of EMT,including E-cadherin,fibronectin,and vimentin,were observed.SB203580 was administrated to test the role of p38 MAPK signaling in EMT.Then AECs were treated with siRNAs targeting p38 MAPK to further test the effects of p38 MAPK,and the role was further confirmed by re-expression of p38 MAPK.At last β-catenin siRNA was used to test the role of β-catenin in the EMT process and relationship of β-catenin and p38 MAPK was concluded.Exposure of AECs to Wnt1 and LiCl resulted in upregulation of vimentin and fibronectin with subsequent downregulation of E-cadherin.Wnt1 and LiCl stimulated the p38 MAPK signaling pathways.Perturbing the p38 MAPK pathway either by SB203580 or through p38 MAPK siRNA blocked EMT and inhibited fibronetin synthesis,which were reversed by transfection of p38 MAPK expression plasmid.β-catenin siRNA attenuated the EMT process and decreased p38 MAPK phosphorylation,indicating that β-catenin is involved in the EMT-related changes through regulation of p38 MAPK phosphorylation.These findings suggest that p38 MAPK participates in the pathogenesis of EMT through Wnt pathway and that p38 MAPK may be a novel target for IPF therapy.

OBJECTIVE: To explore the effect of nitric oxide (NO) on the gene expression of hepatic TNF-α and IL-1β by crush injury of rat's soft tissues. METHODS: Rats were randomly divided into sham group, crush group, crush+aminoguanidine (AG) group, and crush+L-arginine (L-Arg) group. Activities of ALT and AST as well as NO level in serum were measured. Gene expressions of TNF-α and IL-1β were detected with RT-PCR. RESULTS: Obvious increase in TNF-α and IL-1β mRNA expression was detected in the crush group compared with the sham group (P<0.05). After pretreated L-Arg, expressions of TNF-α and IL-1β mRNA were markedly increased (P<0.05). After pretreated AG, those indices obviously decreased (P<0.05). Activities of ALT and AST enhanced and NO level increased in the crush group compared with the sham group (P<0.05). Pretreatment with L-Arg or AG led to substantial increased or reduced activities of ALT and AST as well as NO levels, respectively. CONCLUSION Endogenous NO mediated TNF-α, IL-1β mRNA up expression in liver induced by increased production of NO after crush injury of rat's soft tissues.
Animals ; Gene Expression ; Interleukin-1beta/metabolism* ; Liver ; Nitric Oxide/physiology* ; RNA, Messenger ; Rats ; Tumor Necrosis Factor-alpha/metabolism* ; Wounds and Injuries

BACKGROUNDCoronary artery lesions (CALs) are known to be the main complication in children with Kawasaki disease (KD). Instead of intravenous immunoglobulin (IVIG), corticosteroid therapy has been accepted to be used for children with KD who are unresponsive to IVIG. This study aimed to evaluate risk factors for CALs in children with KD.

METHODSWe retrospectively reviewed the clinical records of 2331 children with KD from January 2005 to December 2014. To identify the independent risk factors for CALs, multivariable logistic regression models were constructed using significant variables identified from univariate logistic regression analysis.

RESULTSThe incidence of CALs was 36.0% (840 of 2331), including 625 (26.8%) coronary artery dilations and 215 (9.2%) coronary artery aneurysms (CAAs). Multivariable logistic regression analysis identified that male, incomplete KD, longer fever duration, and C-reactive protein (CRP) >100 mg/L were independent risk factors for coronary artery dilatations. On the other hand, male, incomplete KD, longer fever duration, prolonged days of illness at the initial treatment, corticosteroid therapy, sodium ≤133 mmol/L, and albumin <35 g/L were the independent risk factors for CAAs. In addition, corticosteroid therapy, prolonged days of illness at the initial treatment, and albumin <35 g/L were the independent risk factors for giant CAAs.

CONCLUSIONSCALs might be associated with male sex, incomplete KD, longer fever duration, prolonged days of illness at the initial treatment, albumin <35 g/L, sodium ≤133 mmol/L, CRP >100 mg/L, and corticosteroid therapy. Corticosteroid therapy was an independent risk factor for CAAs and giant CAAs. Thus, corticosteroids should be used with caution in the treatment of KD with the risk for CALs.

Adolescent ; Adrenal Cortex Hormones ; adverse effects ; Child, Preschool ; Coronary Aneurysm ; chemically induced ; Female ; Humans ; Infant ; Logistic Models ; Male ; Mucocutaneous Lymph Node Syndrome ; drug therapy ; Retrospective Studies

T polymorphism,was selected as putative promoter.The recombinant constructions containing-1328-+812 of TGF-?_1 gene and CAT reporter gene(phTGF2.14T,phTGF2.14C)were constructed and transfected into HepG2 cells with liposomal trans- fection method,then the transfected HepG2 cells were treated with IL-10(4 ng/ml),HGF(10 ng/ml)or IFN-?(20 ng/ml). Reporter gene activity was analyzed by ELISA.Results:Reporter gene activity in cells transfected with phTGF2.14C was sig- nificantly higher than those transfected with phTGF2.14T(P

Objective To explore the effects of thyroid hormone on the expression of homeobox gene Nkx6.1 in offspring of hypothyroidism rats and the relationship between gene expression and hormone level by supplying their hypothyroidism pregnant mother with thyroid hormone. Method A total of 240 Wistar rats were half nude and half female. Female rats were randomly divided into eight groups: control, hypothyroidism group, hypothyroidism groups which were supplied with thyroid hormone in high, medium and low dosage in early stage(1- 17 d) and in late stage( 18 - 20 d). According to 100 grams of body weight, the concentration of thyroid hormone were 3.5,2.0,0.5 μg/d in high, medium and low dosage group. All the rats were fed with low-iodine food. The normal control group was given KIO_3 solution and the other groups were given deionized water. After three months female rats were mated with male rats. The content of Nkx6.1 mRNA in brain tissue of 17-day fetal rats, new-born and 20- day old offspring by real-time fluorescence quantitative PCR techniques. Results①A rat model of hypothyroidism was successfully established, there were statistical significance between 8 groups in TT_3,TT_4,FT_3,FT_4(F=4.08,31.99,5.79,26.34, all P < 0.01 ). ② The expression of Nkx6.1 mRNA had significant difference(F = 758.720, 1121.589,144.716, all P < 0.01 ) between groups in 17-day fetal rats, new-bern and 20-day old offsprings and intra- groups in different time (F=2898.863,325.605,716.285,56.329,236.727,196.678,7115.752,9152.306, all P < 0.01 ). ③The time factor and dosage factor had influence on Nkx6.1 mRNA expression(F = 1176.655,246.530, all P < 0.01 ). There were interaction between time and dosage factor(F = 1249.934, P < 0.01 ). ④Comparison of Nkx6.1 mRNA expression between hypothyroidism group and normal control group had significant difference in the above three time points(all P < 0.01 ). ⑤Comparisons of Nkx6.1 mRNA expression between 6 hypothyroidism groups which were supplied with thyroid hormone and hypothyroidism group had significant difference(all P < 0.01 ) in new-bern and 20-day old offspring; comparisons of Nkx6.1 mRNA expression between hypothyroidism groups which were supplied with high and medium thyroid hormone and hypothyroidism group had significant difference in 17-day fetal rats(all P < 0.01 ). ⑥Comparison of Nkx6.1 mRNA expression between hypothyroidism groups which were supplied with medium thyroid hormone in early stage and normal control group had no statistical significance (all P > 0.05), while between the other 5 groups which were supplied with thyroid hormone and normal control group had significant difference(all P < 0.01 ) in the above three time points.⑦Multiple comparison of early stage groups which were supplied with thyroid hormone showed that the expression of Nkx6.1 mRNA had significant difference(all P < 0.01) between high, low dosage groups and medium group in 17-day fetal rats, new-bern and 20-day offspring(all P< 0.01). ⑧Multiple comparison of late stage groups supplied with thyroid hormone showed that old offspring and between high dosage groups and low dosage groups in 17-day fetal rats and 20-day the expression of Nkx6.1 mRNA had significant difference(all P < 0.01 ) between three groups in new-bern and 20- day old offspring. Conclusion The expression of Nkx6.1 in rats offspring is highly related to the supply dosage and supply time of thyroid hormone in hypothyroidism pregnant rats.

Objective To study the single nucleotide polymorphisms (SNP) characteristics of Yersinia pestis strains from different natural foci in China.Methods Genome-wide comparison was done to find SNP sites by the Mummer program among 9 Yersinia pestis genome which was downloaded from NCBI.Then 13 genic fragments including 19 SNP sites were amplified by PCR and sequenced in 133 Yersinia pestis strains,and the results were cluster analyzed with the BioNumerics software.Results Three thousand seven hundred and eighty sequence variation sites were found by genome-wide comparison.Using the different combinations of SNP sites,UPGMA cluster analysis revealed obvious geographic regional and eco-aggregation characteristics of Yersinia pestis strains isolated from China.Conclusions As relatively stable genetic markers,SNP can better reflect the genome characteristics of Yersinia pestis in different plague natural foci of China.

PURPOSE: Intravenous gamma globulin(IVGG) treatment has reduced symptoms and complications in Kawasaki disease(KD). However, fever persisted in 20-30% of the patients, and there are no reliable data on the indication and dosage of IVGG re-treatment. Therefore, we tried to reveal the effectiveness of IVGG re-treatment and to find risk factors in predicting the re-treatment. METHODS: Among 57 patients with typical KD, 47(82.5%) patients were put into group A, which improved after the treatment with standard 2g/kg of IVIG. 10(17.5%) patients were put into group B, which retreated with 1-2g/kg of IVIG due to persistent fever for at least 3 days after standard IVIG therapy. We compared clinical symptoms, laboratory findings and echocardiograms between group A and B, retrospectively. RESULTS: All patients in group B responded IVGG re-treatment and no considerable side effects. The total duration of the fever was significantly longer(P<0.001) and the initial and peak levels of CRP and the peak levels of ESR were significantly higher(P<0.01) in group B compared to group A. Even though leukocytosis, cervical lymphadenitis and coronary artery aneurysm were more frequent, and the levels of serum lipids at admission were lower in group B, without significance. CONCLUSION: IVGG re-treatment appeared to be effective in the treatment of refractory KD, but could not reduce the incidence of coronary artery aneurysm. We concluded it was difficult to predict risk factors for IVGG re-treatment from these data. Further studies are needed to determine the indication and appropriate dosage of IVGG re-treatment.
Aneurysm ; Coronary Vessels ; Fever ; gamma-Globulins* ; Humans ; Immunoglobulins, Intravenous ; Incidence ; Leukocytosis ; Lymphadenitis ; Mucocutaneous Lymph Node Syndrome* ; Retrospective Studies ; Risk Factors

BACKGROUND: There have been few clinical studies of halo nevi. OBJECTIVE: The purpose of this study was aimed at evaluating the clinical and histopathologic features of halo nevi and correlation between halo nevi and vitiligo. METHODS: The medical records of 40 patients with halo nevi and biopsy specimens of 30 patients with halo nevi were reviewed. RESULTS AND CONCLUSION: 1.The ratio of male to female patients was 1:1.5. 2.The mean age of the onset was 20.1 years (male : 11.5, female : 25.9). 3.Multiple halo nevi were present in 32.5%(male : 25%, female : 37.5%). 4.The areas in which the lesions developed were head and neck(39.1%), back(32.8%), anterior chest(12.5%), abdomen(10.9%), groin(3.1%), and lower extremity(1.6%) in descending order. 5.The mean duration of each color in central nevus was as follows: black(2.7 years), pink(3.5 years), gray(4.9 years) 6.The most common pathology of the central nevus was intradermal(80%). 7.The halo nevi associated with vitiligo were 21 cases(52.5%) out of 40 patients. Among them, the cases with non-segmental vitiligo were 13(61.9%), the cases with segmental vitiligo were 8(38.1%). 8.Out of the 21 cases with halo nevi associated with vitiligo, the cases with halo nevi prior to vitiligo were 4(19.1%), concurrent onset 12(57.1%) and the cases with halo nevi after vitiligo were 5(23.8%).
Biopsy ; Female ; Head ; Humans ; Male ; Medical Records ; Nevus ; Nevus, Halo* ; Pathology ; Vitiligo