Fibrolase is a non-hemorrhagic zinc metalloproteinase isolated from southern copperhead snake venom (Agkistrodon contortrix contortrix) and is capable of degrading fibrin clots resulting from purified fibrinogen or from blood plasma. Alfimeprase, a truncated form of fibrolase, as a clinical agent was successfully completed PhaseII clinical trials.The cDNA of alfimeprase was amplified by recursive PCR, digested with BamHI and HindII, and cloned into pET43.1a, pMALp2X and pMALc2X vectors to generate fusions with NusA, MBP and sMBP(with signal peptide), respectively. Nus/alfimeprase was expressed in soluble form by co-expressing with chaperone FkpA and inducing with1mmol/L IPTG. The fusion protein accounted for about 25 % of total protein following cell lysis. Alfimeprase was successfully purifiesd by Ni-NTA affinity chromatography and cleaved by enterokinase. The results demonstrate the fibrinolytic activity of recombinant alfimeprase using fibrin plate assays and fibrinogen hydrolysis.

Objective Toinvestigatetheprevalenceofurinaryincontinenceanditseffectonlifequalityamongfemale adultsinHangzhou,andtoprovideevidencetothecontroloffemaleurinaryincontinence.Methods Aquestionnaire survey was performed in Gongshu and Xiacheng districts in Hangzhou from October 2013 to June 2014,and 4 563 women aged over 20 years were interviewed through a questionnaire including International Consultation on Incontinence Questionnaire Lower Urinary Tract,IQ-FLUTS and demographic information.Single and multi factor logistic regression analysiswereusedtoanalyzetheriskfactorsofurinaryincontinence.Results Atotalof4785questionnairesweresent, and 4 563 effective questionnaires were recovered,with a recovery rate of 95.4%.The prevalence of urinary incontinence was 33.5%(1 530/4 563),and female with older age tended to have a higher prevalence of UI(P<0.01).Among which stress urinary incontinence (SUI ),urge incontinence (UUI )and mixed urinary incontinence (MUI ) were accounted for 20.2%(922/4 563),3.0%(135/4 563)and 10.3%(473/4 563)respectively.UUI and MUI had a greater effect on quality of life.According to the multi factor logistic regression analysis,gravidity,age,constipation,pelvic surgery and fat (BMI>24)wereriskfactorsforurinaryincontinence(P<0.05).Conclusion Becauseofthehighprevalenceofurinary incontinence among female adults,more attention should be paid to urinary incontinence suffers in order to improve the quality of life of female adults.

BACKGROUND: Yaotu Granules have been proved to protect human nucleus pulposus cells and delay their degeneration. Notably, Yaotu Granules for lumbar disc herniation has achieved good clinical results.OBJECTIVE: To investigate the effect of the herbal compound formula Yaotu Granules on the Fas/FasL expression in a rabbit model of lumbar disc degeneration, and further elucidate the underling mechanism of preventing and treating lumbar disc degeneration.METHODS: Twenty New Zealand white rabbits were enrolled and the models of lumbar disc degeneration were established by minimally invasive puncture and rotation cutting, followed by randomized into normal saline, low-, middle-,and high-dose groups (n=5 per group). 10 mL of normal saline, 10, 20, and 40 mL of water decoction of Yaotu Granules were administered intragastrically into the normal saline, low-, middle-, and high-dose drug groups for 21 days, twice daily, respectively. Subsequently, the expression level of Fas/FasL in the rabbit nucleus pulposus cells in each group was detected.RESULTS AND CONCLUSION: The signal intensity of the rabbit lumbar disc on MRI was decreased, and ruptured annulus and posterior herniated disc were visible at 12 weeks after modeling. Masson staining showed that the nucleus pulposus cells arranged in disorder, and even ruptured. Additionally, safranin O staining found that the number of nucleus pulposus cells was decreased obviously. The order of the relative expression levels of Fas and FasL mRNA in the nucleus pulposus cells was as follows: normal saline group > low-dose drug group > middle-dose drug group > high-dose drug group (P < 0.05). These results suggest that Yaotu Granules delay the rabbit lumbar disc degeneration by downregulating the expression level of Fas/FasL.

OBJECTIVETo investigate clinical features and treatments of cervical spinal injury in aged patients.

METHODSFrom July 2006 to October 2011, 27 patients with cervical spinal cord injury were divided into experiment group, including 20 males and 7 females with an average age of 69 years old (ranged 65 to 78 years old). The other 22 patients with the same degree of cervical spianl cord injury and the same period hospitalization were chosen to regard as control group, including 16 males and 6 females with an average age of 47 years old (ranged 38 to 65 years old). All of them had clear history of trauma, and were admitted to hospital within one week. Clinical data and result of follow up were evaluated between two group. The function of the spinal cord was measured by Frankel classification at admission and final follow-up.

RESULTSAll patients were followed up for 1 year except 6 died in aged group, among which 3 paients with complications were died during hospitalization and others were out of hospital. Twenty-three cases had accompanied diseases and 24 cases occurred complications in aged group. Contrast to control group, this difference was statistically significant. At the final follow-up, there were 2 cases at Frankel grade E in aged group and 6 cases in control group.

CONCLUSIONIn comparison with control group, there are more accompany diseases and higher complications and mortality rates in aged group, while poor improvement of spinal cord function. This patients were mainly treated with operation, but operation method should simplify and mainly focus on effective decompression and reconstuction stability and avoid expansion operation. It's nesscerry to pay more attention to prevent and treatment of respiratory complication.

Adult ; Aged ; Cervical Vertebrae ; injuries ; Female ; Humans ; Male ; Middle Aged ; Postoperative Complications ; epidemiology ; mortality ; therapy ; Spinal Injuries ; physiopathology ; surgery

Objective:To investigate clinical features and treatments of cervical spinal injury in aged patients. Methods:From July 2006 to October 2011,27 patients with cervical spinal cord injury were divided into experiment group,including 20 males and 7 females with an average age of 69 years old (ranged 65 to 78 years old). The other 22 patients with the same de-gree of cervical spianl cord injury and the same period hospitalization were chosen to regard as control group ,including 16 males and 6 females with an average age of 47 years old (ranged 38 to 65 years old). All of them had clear history of trauma, and were admitted to hospital within one week. Clinical data and result of follow up were evaluated between two group. The function of the spinal cord was measured by Frankel classification at admission and final follow up. Results:All patients were followed up for 1 year except 6 died in aged group ,among which 3 paients with complications were died during hospitalization and others were out of hospital. Twenty three cases had accompanied diseases and 24 cases occurred complications in aged group. Contrast to control group,this difference was statistically significant. At the final follow up,there were 2 cases at Frankel grade E in aged group and 6 cases in control group. Conclusion:In comparison with control group,there are more accompany diseases and higher complications and mortality rates in aged group ,while poor improvement of spinal cord function. This pa-tients were mainly treated with operation,but operation method should simplify and mainly focus on effective decompression and reconstuction stability and avoid expansion operation. It's nesscerry to pay more attention to prevent and treatment of res-piratory complication.

BACKGROUNDOur previous research has suggested that genes around D12S1056 in 12q13 may confer susceptibility to ventricular septal defect (VSD) in humans. The present study was to define the chromosome region assignment by transmission disequilibrium test (TDT), and to identify the important candidate gene by family-based association study and haplotype analysis.

METHODSSurrounding D12S1056, ten microsatellite markers including D12S329, D12S305, D12S1662, D12S1056, D12S1293, D12S334, D12S102, D12S83, D12S1655 and D12S1691 were chosen, and TDT was performed in 62 nuclear family trios each consisting of an affected child and two healty parents. Subsequently, the GLI gene, a positional candidate gene that maps to the target region, was selected for further analysis. Three single nucleotide polymorphisms (SNPs), G11888C, G11388A, and G11625T, were selected for family-based association study and haplotype analysis.

RESULTSVSD was significantly associated with all selected markers except D12S1691 [72.2 centi morgen (cM)] and D12S1700 (75.76 cM). VSD was also significantly associated with G11888C (chi(2) = 5.918, P = 0.015), G11388A (chi(2) = 8.067, P = 0.005), and G11625T (chi(2) = 11.842, P = 0.001). Haplotype analysis showed a strong linkage disequilibrium between G11888C and G11388A (D' = 0.999), but in significant (chi(2) = 1.035, df = 2, P > 0.05).

CONCLUSIONSThe susceptibility gene of VSD was mapped to 3.56 cM in 12q13 by TDT, and the GLI gene, an important candidate in the target region, was associated with VSD.

Child ; Child, Preschool ; Chromosome Mapping ; Chromosomes, Human, Pair 12 ; Female ; Genetic Predisposition to Disease ; Haplotypes ; Heart Septal Defects, Ventricular ; genetics ; Humans ; Linkage Disequilibrium ; Male ; Microsatellite Repeats ; Transcription Factors ; genetics ; Zinc Finger Protein GLI1

BACKGROUND: Methylene blue is used as a developer to identify intervertebral disc degeneration in the transforaminal endoscopic surgery. However, many scholars have indicated that methylene blue can accelerate the degeneration process, whilst foreign researches have reported that it may play therapeutic effect on degenerative intervertebral discs under acidic conditions due to its acidophily. Therefore, whether methylene blue holds toxic effect on the disc remains controversial. OBJECTIVE: To determine whether methylene blue exerts toxic effect on nucleus pulposus cells by cell counting-kit 8 (CCK-8) assay. METHODS: The discarded nucleus pulposus from two patients with intervertebral disc herniation were selected. After digestion, nucleus pulposus cells were extracted and cultured until proliferated to 80% of the medium. Then, the cells were digested to make cell suspensions, divided into six groups and inoculated into the 96-well plates: blank control (only the medium, CCK-8 solution), control group (only medium, cells and CCK-8 solution), and the other groups were cultured with 1%, 2%, 3%, and 4% methylene blue, respectively. The absorbance values were measured by CCK-8 assay at 24, 48, 72 and 96 hours after incubation to calculate the cell viability, and the color change was observed. RESULTS AND CONCLUSION: The color in the control group was the deepest, and the color became lighter with the concentration of methylene blue increasing. The cell viability was the highest in the control group, and it was decreased with the concentration of methylene blue increasing. Thus, methylene blue may exert toxic effect on human nucleus pulposus cells.

BACKGROUND: Our preliminary study has shown that methylene blue exerts toxic effect on human nucleus pulposus cells in a concentration-dependent manner, but the toxic range remains unclear. OBJECTIVE: To determine the critical range of the cytotoxicity of methylene blue to nucleus pulposus cells by cell counting-kit 8 (CCK-8) assay. METHODS: The nucleus pulposus was from a patient with intervertebral disc herniation. The nucleus pulposus cells were extracted and cultured. Passage 1 cells mere used to make cell suspensions. The cells were divided into nine groups for culture: blank control (only the medium, CCK-8 solution), control (only medium, cells and CCK-8 solution), and methylene blue groups (1%, 0.5%, 0.1%, 0.05%, 0.01% and 0.005% of methylene blue). The absorbance values were measured by CCK-8 assay at 24, 48, 72 and 96 hours after incubation. The cell viability was calculated, and the color was observed. RESULTS AND CONCLUSION: The color in the control group was the darkest, and the color in the methylene blue groups became lighter with the concentration of methylene blue increasing, and the 0.05%, 0.01%, 0.005% and 0.001% methylene blue groups showed darker color similar to the control group. The absorbance values in the 0.1% methylene blue group were significantly less than those in the control group (P＜ 0.05). There were no significant differences in the absorbance values and cell viability between 1% and 0.5% methylene blue groups (P ＞0.05). The absorbance values and cell viability in the 0.1% methylene blue group were significantly higher than those in the 0.5% methylene blue group, but were significantly less than those in the 0.05% methylene blue group (P ＜ 0.05). Thus, methylene blue exerts cytotoxicity to human nucleus pulposus cells, and the critical value of toxicity is between 0.1% and 0.05% and close to 0.05%. However, the exact value needs a further investigation.

OBJECTIVETo study the relationship between milk intake of 3 - 6 years old preschool children and their growth.

METHODSThe data bank of 2002 China National and Health Survey were used to analyze the relationship of milk consumption and the growth of children aged 3 - 6 in urban and rural areas.

RESULTSThe percentages of the diet with milk in urban and rural areas were 46.9% and 8.2% respectively, there was significant difference between the urban and rural areas. The averaged milk intake of preschool children were 23.8 g/d, the milk intake of urban children was significantly higher than that in the rural areas (P < 0.01). The height and weight of children with milk or its products in their diets in the group aged 4, 5 and 6 years were significantly higher than those of children without milk or its products in their diets (P < 0.05).

CONCLUSIONThere was close relationship between milk intake of children 3 - 6 years old and their growth, the volume of milk intake of preschool children should be improved in urban and rural areas.

Animals ; Body Height ; Body Weight ; Child ; Child, Preschool ; China ; Dairy Products ; Diet ; Health Surveys ; Humans ; Milk ; Rural Population ; statistics & numerical data ; Urban Population ; statistics & numerical data

OBJECTIVETo study the relationship between the hemoglobin level of children < or = 3 years and anemia of their mothers.

METHODSMulti-steps cluster sampling was adopted. In order to ensure sufficient number of babies and young children and postpartum in this survey, additional subjects were included in the sample. The subjects in study were collected in the same family.

RESULTSThe average anemia prevalence of children < or = 3 years was significant different between urban and rural (26.9%, 94/350 and 32.4%, 594/1835, P < 0.05) areas. The birth weight of anemia children < or = 3 years was significant lower than nonanemic children (P < 0.05); the incidence of low birth weight of anemia children < or = 3 years was significant higher than nonanemic children (P < 0.05); the percentage of Hb < 100 g/L and < 110 g/L were 7.6% (166/2185) and 24.9% (544/2185); the Hb levels of children fed by anemia mother and nonanemic mother were significantly different (115.6 +/- 17.4) g/L, (115.2 +/- 16.7) g/L and (123.3 +/- 14.9) g/L, (122.2 +/- 15.3) g/L, P < 0.001.

CONCLUSIONThere should be significant relation between the Hb level of children < or = 3 years and anemia of their mother.

Adult ; Anemia ; epidemiology ; Birth Weight ; Breast Feeding ; adverse effects ; statistics & numerical data ; Child, Preschool ; Female ; Hemoglobins ; metabolism ; Humans ; Infant ; Infant, Newborn ; Male ; Matched-Pair Analysis ; Mothers ; Prevalence ; Rural Population ; Sampling Studies ; Urban Population