Objective To study the effect of different concentrations of fluoride on cultured human umbilical vein vascular endothelial cells(HUVEC). Methods Different doses of sodium fluoride (NaF) were added to HUVEC culture medium, fluoride concentrations were 0(control), 100,400,700,1000,2000 μmol/L, respectively,6 re-set hole in each group. After continuous culture for 48 h, cells and culture medium were collected. Cell morphology was studied by Wright-Giemsa staining; cells apoptosis was determined by acridine orange fluorescence staining; cell activity was measured by methyl thiazolyl tetrazolium (MTT) assay; superoxide dismutase (SOD),glutathione peroxidase(GSH-Px) activity, malonaldehyde(MDA) content, induced nitricoxide synthase(iNOS), and endothelia nitricoxide synthase(eNOS) activity in cell culture medium were determined by spectrophotometry; cell iNOS mRNA and eNOS mRNA expression were detected by RT-PCR; intercellular adhesion molecule-1 (ICAM-1)and vascular cell adhesion molecule-1 (VCAM-1) levels were detected by double antibody sandwich ELISA method.Results With increased dose of fluoride, HUVEC cells decreased, the structure changed. In 400 - 2000 μmol/L group, the SOD activity[(6.627 ± 0.213), (6.668 ± 0.152), (5.935 ± 0.122), (4.755 ± 0.182)kU/L] was lower than those of the control group[(7.457 ± 0.398)kU/L, P ＜ 0.05 or ＜ 0.01], GSH-Px activity[(481.284 ± 43.785),(492.223 ± 16.474), (382.762 ± 25.167), (293.687 ± 24.881 )kU/L] was also lower than those of the control group [(585.078 ± 47.323)kU/L, P ＜ 0.05 or ＜ 0.01], MDA level[(0.609 ± 0.011 ), (0.646 ± 0.016), (0.852 ± 0.013),(1.188 ± 0.045)nmol/L] was higher than those of the control group[(0.512 ± 0.027)nmol/L, P ＜ 0.05 or ＜ 0.01];iNOS activity[(3.604 ± 0.115), (3.615 ± 0.075), (3.848 ± 0.103), (4.275 ± 0.079)kU/L] also was higher than those of the control group[(2.798 ± 0. 136)kU/L, all P ＜ 0.01], iNOS mRNA expression increased, eNOS activity [(5.539 ± 0.079), (5.503 ± 0.064), (5.226 ± 0.142), (4.809 ± 0. 107)kU/L] decreased compared to those of control group[(5.996 ± 0.155)kU/L, P ＜ 0.05 or ＜ 0.01], eNOS mRNA expression decreased; ICAM-1 levels [(0.852 ± 0. 102), (0.886 ± 0.061 ), (0.961 ± 0.158), (1.418 ± 0. 167)μg/L] increased compared to those of the control group[(0.687 ± 0.046)μg/L, P ＜ 0.05 or ＜ 0.01], VCAM-1 levels[(2.719 ± 0.197), (2.946 ± 0.167),(3.173 ± 0.225 ), (3.613 ± 0. 153 ) μg/L] was higher than those of the control group [(2.375 ± 0.067 ) μg/L, all P ＜0.01]. Conclusions High concentrations of fluoride reduce the activity of antioxidant enzymes, which leads to metabolic disorders of nitric oxide and abnormal cytokines expression, thereby inhibiting vascular endothelial cell growth, structural change and induced apoptosis. This is an important factor in high fluoride-induced vascular endothelial injury.

OBJECTIVETo explore the relation of angiotensin-converting enzyme (ACE) gene polymorphism, angiotensin II type I receptor (ATIR) gene polymorphism and other factors on cerebral infarction.

METHODSOne thousand three hundred fifty-one subjects from Tangshan coalmine were enrolled with study method of cluster sampling. Face to face interviews were conducted to fill in questionnaires by trained interviewers. ACE gene, ATIR gene and inflammation factors including tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), IL-8, IL-10, C reactive protein (CRP), fibrinogen (Fg), fibrin monome polymerized velocity (FMPV), absorbance maximum (A(max)), FMPV/A(max), were measured.

RESULTSNo different prevalence rates of ACE genotype were found on cerebral infarction. The distributions of AA genotype of ATIR gene in the cerebral infarction was higher than that of the controls. The prevalence of AA genotype was higher than other groups, but the prevalence of combined genotype did not show much difference. Under the existence of factors that related to cerebral infarction, AA genotype frequencies were higher than those of non-smoking and with hypertension. IL-6, ATIR gene polymorphism, sex, FMPV/A(max) were strongly related to cerebral infarction. The level of IL-6 was higher than the normal ones.

CONCLUSIONSThe prevalence of cerebral infarction obviously increased in the hypertensive groups having AA genotype of ATIR gene. In the cerebral infarction groups, the level of IL-6 was higher than that in the normal population, indicating that these can be resulted from local inflammation and immunity reactivity. Environmental and genetic factors in the pathogenesis of cerebral infarction might have coordinating functions.

Aged ; Cerebral Infarction ; genetics ; Cross-Sectional Studies ; Female ; Genotype ; Humans ; Logistic Models ; Male ; Middle Aged ; Peptidyl-Dipeptidase A ; genetics ; Polymorphism, Genetic ; Receptor, Angiotensin, Type 1 ; genetics

Objective To study the decomposition kinetics of omethoate in blood.Methods The acetonitrile precipitated protein was added into the blood, with the chromatographic column of a Waters BEH C18column (2.1 mm×50 mm, 1.7μm), the mobile phase of 5 mmol/L ammonium acetate aqueous solution-methanol, and the gradient elution with a flow rate of 0.3 mL/min and injection volume of 2μL.With electrospray ionization (ESI) source and positive ion detection, qualitative and quantitative analyses were taken using multi-reaction monitoring mode.Omethoate standard was added into blank human blood to the mass concentrations of 0.78, 1.40, 2.30, 4.50, and 7.20μg/mL, and each mass concentration was preserved at 3 temperatures of-20℃, 4℃, and 20℃, respectively.The content of omethoate was detected at different time points (0, 1, 3, 4, 7, 11, 15, 24, 32, 40, 48, 64, 80, 96, and 120 d).Results Different concentrations of omethoate all showed a descended trend in human blood under different temperature conditions.The decomposition in storage environment of-20℃, 4℃, and 20℃was fit to a one-compartment open model with a first-order kinetic process, which could be expressed as Ct=Coe-αt, with the calculated theoretical values of omethoate concentration close to the measured values.Conclusion All concentrations of omethoate are decomposed in the blood, which vary a lot in different preservation conditions.It is suggested that blood samples should be frozen and detected timely in suspected omethoate poisoning cases.

Objective To assess the interference by calcium dobesilatein 7 peroxidase-baseduric acid assays and to determine its clinical significance.Methods In the in vitro experiments, uric acid in pooled serum with final concentrations of calcium dobesilate additions (0, 2, 4, 8, 16, 32, and 64μg/ml) were measured by 7 peroxidase-based assays.Percent Bias (%) was calculated relative to the drug-free specimen.In the in vivo experiments, changes in serum uric acid and calcium dobesilate concentrations were observed before and after calcium dobesilate administration ( baseline, 0 h, 1 h, 2 h, 3 h, 4 h, 6 h ) involunteers.The interference in different assays was assessed compared with LC-IDMS/MS method. Calcium dobesilate levels in 40 specimens from those taking calcium dobesilate were measured by HPLC method.Of the 40 specimens, 10 were selected to analyse the levels of uric acid by both peroxidase and UV measurement method to assess the impact in clinical status.Results In the in vitro study, concentrations of uric acid measured by 7 peroxidase-based assays were reduced by -6.3%to -21.2%compared with drug-free serum, when theconcentration of calcium dobesilate was16μg/ml.In the in vivo study, comparedto UA levels at 0 h, the biasesof serum uric acid determined by peroxidase method after calcium dobesilate administration(1 h, 2 h, 3 h, 4 h, 6 h) were of -3.33%, -6.79%, -7.49%, -6.07%, -4.09%, respectively.The observed uric acid concentrations for 8 participants measured by enzymatic assays were inhibited by -3.75% to -6.89% at 0 hour and by -16.9% to-22.22% at 2 hours relative to the concentrations measured by the LC-IDMS/MS method. Conclusions Calcium dobesilate produced a clinically significant negative interference with uric acid in all peroxidase-based uric acid assays,which may result in false evaluation of uric acid level in clinical status.Significant differences in the degree of interference were observed among the assays.

OBJECTIVETo observe the distribution character of angiotensin converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism in Chinese population and its relationship with essential hypertension.

METHODSPolymerase chain reaction (PCR) technique was used to detect the angiotensin converting enzyme gene I/D polymorphism in 2966 subjects of Kailuan Coal Mine, and further restriction analysis was performed.

RESULTSThe frequencies of ACE II, ID, DD genotypes in total study population were 41.5%, 38.4%, 20.1%, respectively. The DD genotypes in hypertensive group and that in control group were 18.9% and 21.0%, respectively. There was no significant difference between hypertensive group and control group (P>0.05). The same result was seen after stratification by age and gender in each group, respectively(P>0.05). The DD genotype and D allele showed a tendency to decrease with the increase of age (P<0.001).

CONCLUSIONThe above results suggested that essential hypertension was not associated with ACE I/D polymorphism. The distributions of ACE genotype and allele varied with age, and the subjects with the character of DD genotype were at higher risk of early death.

Adult ; Age Factors ; Aged ; Aged, 80 and over ; Female ; Gene Frequency ; Genotype ; Humans ; Hypertension ; enzymology ; genetics ; Male ; Middle Aged ; Peptidyl-Dipeptidase A ; genetics ; Polymorphism, Genetic ; Sex Factors

The E2 envelope glycoprotein of virulent Shimen strain and avirulent C-strain of Classical swine fever virus (CSFV) has 5 and 6 potential glycosylation sites, respectively, and the potential glycosylation site 986N is unique to C-strain. To study the differences in glycosylation between the virus pair, the E2 genes (removing signal sequence and transmembrane anchor regions) of the two strains fused with the melittin signal sequence were expressed in the Sf9 insect cells. The recombinant E2 proteins were secreted into the medium of Sf9 cells in dimer form with different molecular weight (MW). Deglycosylation of the recombinant E2 proteins by endo H and PNGase F showed that the deglycosalated Shimen-E2 and HCLV-E2 have the same MW, indicating that the different MW between Shimen-E2 and HCLV-E2 proteins came from different glycosylation. Site-directed mutagenesis in the potential glycosylation site at 986N demonstrated that the mutated Shimen-E2 protein had the same MW as the wild-type HCLV-E2 protein, while the mutated HCLV-E2 had the same MW as the wild-type Shimen-E2 protein. We suggest that the different MW between Shimen-E2 and HCLV-E2 is resulted from the different glycosylation on 986 N glycosylation site.
Baculoviridae ; genetics ; Blotting, Western ; Classical swine fever virus ; chemistry ; classification ; Glycosylation ; Molecular Weight ; Mutation ; Recombinant Proteins ; chemistry ; Viral Envelope Proteins ; chemistry ; Virulence

OBJECTIVETo develop a primer-extension in combination with denaturing high-performance liquid chromatography (PE-DHPLC)-based assay for prenatal diagnosis of the five most common beta-thalassemia mutations in Chinese.

METHODSThe human beta-globin gene fragment was amplified by PCR, followed by a multiple PE reaction specific for each five mutations. Then the PE product mixtures were separated for genotyping of beta-globin gene mutations using fully-denaturing DHPLC analysis.

RESULTSIn a blind study, prenatal diagnosis was performed on thirty-six at-risk families for beta-thalassemia major. Reverse dot blot (RDB) analysis was used to validate each result, showing an accuracy rate of 100% for PE-DHPLC in a total of 108 samples tested. Overall, by PE-DHPLC analysis, the authors could identify the genotypes involving the five mutations and normal alleles corresponding to 94.4% (102/108) and actually make final decision for prenatal diagnosis covering 97.2% (35/36).

CONCLUSIONThe PE-DHPLC protocol can be a simple, rapid, and highly accurate assay in the prenatal detection of common beta-thalassemia mutations.

Base Sequence ; Chromatography, High Pressure Liquid ; methods ; DNA Mutational Analysis ; methods ; DNA Primers ; Female ; Fetal Diseases ; diagnosis ; genetics ; Genotype ; Globins ; genetics ; Humans ; Molecular Sequence Data ; Point Mutation ; Pregnancy ; Prenatal Diagnosis ; beta-Thalassemia ; diagnosis ; genetics

This study was aimed to explore the prognostic significance of telomere length in patients with chronic lymphocytic leukemia (CLL) and to analyze relation of telomere length with Binet stage, IgVH mutation status, CD38, ZAP-70 expression as well as other clinical features. 35 CLL patients who contained 80% or more tumor cells in the peripheral blood or bone marrow samples were selected as objects studied, while 13 healthy donors were served as normal controls. The telomere relative length was detected by using a real-time fluorescent quantitative polymerase chain reaction method (qPCR); the expression of CD38 and ZAP-70 protein were detected by flow cytometry, the IgVH mutation was detected by multiplex PCR. The results showed that the mean telomere relative length in CLL patients and normal controls were 0.384 and 0.443 respectively, but the difference between them was not significant (p > 0.05). The telomere length was significantly correlated with Binet stages and IgVH mutation status. Patients in Binet stage B and C showed significantly shorter telomeres than those in Binet stage A (p = 0.001). Mean telomere relative lengths in patients without IgVH mutation were shorter than those in patients with IgVH mutation (p = 0.015). No relation of telomere length with sex, age, ZAP-70 protein and CD38 were found (p > 0.05). It is concluded that telomere length may have a prognostic significance for CLL patients. Combining telomere length and IgVH mutation status may achieve a better prognostic subclassification for CLL patients.
ADP-ribosyl Cyclase 1 ; metabolism ; Adult ; Aged ; Aged, 80 and over ; Case-Control Studies ; Female ; Humans ; Leukemia, Lymphocytic, Chronic, B-Cell ; genetics ; metabolism ; Male ; Middle Aged ; Mutation ; Prognosis ; Telomere ; chemistry ; genetics ; metabolism ; ZAP-70 Protein-Tyrosine Kinase ; metabolism

Objective:By exploring a construction and improvement system for total lab automation in emergency lab, this work aims to improve the success rates of project implementation and the effective utilization of medical resources.Methods:Firstly, clarify the short board of current solution and determine the configuration of the TLA solution which include the number of pre-analytic, post-analytic modules and analyzers by workflow and data analysis which based on the data from 16th to 22nd September 2019. Secondly, define the basic principles and design the optional solutions. Thirdly, determine the final solution based on the simulation results. Fourthly, evaluate the effectiveness of the solution by comparing the turnaroud time (TAT) before and after implementation.Result:We upgrade the system to improve the automation level of the track by add 1 online centrifuge module and upgrade the capacity of online storage module from 9 000 samples to 15 000 samples, and increase the throughput of the clinical chemistry analyzers form 2 500 tests/h to 5 400 tests/h and immunoassay analyzers from 668 tests/h to 1 320 tests/h with no more space. The 95th percentile TAT (from sample check in to validation) have 33% reduction on the premise of 9% increase on the workload of clinical chemistry and immunoassay, and it is highly consistent with the results obtained by data simulation whose 95th percentile TAT is 67 min.Conclusion:Good uasge of workflow and data analysis in the stage of solution design can help to ensure that the result can meet the expectations of the clinicians and effective utilization of medical resources.

Aim To investigate the protective effect of Qi ZhiXiaoke granules ( QZXK ) on nerve injury using zebrafish and nerve cell injury models. Methods The nerve injury model was established using wild zebrafish AB line, 72 hours after fertilization treated with 1-methyl-4-phenyl-1 , 2 , 3 , 6-four pyridine ( MPTP ) .Then QZXK of different doses were administered for three days,and the trajectory of the zebrafish behavior was recorded and analyzed. Neuroblastoma PC12 cells were incubated with different concentrations of QZXK and MPTP,and the cell viability of PC12 cells was de-tected by MTT. The mitochondrial membrane potential and expression of apoptosis related protein Caspase3 were measured by kits. Results Compared with con-trol group,MPTP reduced the movement distance of ze-brafish,and with the increase of concentration, QZXK promoted the movement distance and reversed the swimming behavior abnormality of zebrafish. Compared with control group, QZXK could inhibit the apoptosis induced by MPTP and promote the cell viability of PC12 cells with MPTP. QZXK improved the membranepotential and decreased the expression of Caspase3 . Conclusions QZXK exerts neuroprotective effect in the process of nerve injury induced by MPTP. The mechanism may be related with inhibiting apoptosis of neural cells. These experiment provides experimental and theoretical foundation for QZXK promoting cogni-tive function.