Objective The purpose of this study was to develop a high-throughput micro-plate radiobinding assay (RBA) of glutamic acid decarboxylase antibody (GAD-Ab) and to evaluate its clinical application. Methods 35labeled GAD65 antigen was incubated with sera for 24 h on a 96-well plate, and then transferred to the Millipore plate coated with protein A, which was washed with 4℃ PBS buffer, and then counted by a liquid scintillation counter. The GAD-Ab results were expressed by WHO standard unit (U/ml). A total of 224 healthy controls, 162 patients with type 1 diabetes mellitus(T1DM) and 210 patients with newly diagnosed type 2 diabetes (T2DM) were recruited. A total of 119 TI DM and healthy cases with gradually changing GAD-Ab levels were selected to compare the consistency of micro-plate RBA with conventional radioligand assay (RLA). Blood samples were obtained from the peripheral vein and finger tip in 32 healthy controls, 35 T1DM and 24 T2DM patients, and tested with micro-plate RBA and then compared with the conventional RLA to investigate the reliability of finger tip sampling. Linear correlation,student's t-test, variance analysis and receiver operating characteristic (ROC) curve were performed using SPSS 11.5. Results (1) The optimized conditions of micro-plate RBA included 2 μl serum incubated with3 ×104 counts/min 35S-GAD for 24 h under slow vibration, antigen-antibody compounds washed 10 times by 4℃ PBS buffer, and radioactivity counted with Optiphase Supermix scintillation liquid. (2)The intra-batch CV of the micro-plate RBA was 3.8%- 10.2%, and the inter-batch CV was 5.6%- 11.9%. The linearity analysis showed a good correlation when the GAD-Ab in serum samples ranged from 40.3 to 664 U/ml and the detection limit of measurement was 3.6 U/ml. The results from Diabetes Autoantibody Standardization Program (DASP) 2005 showed that the sensitivity and specificity for GAD-Ab were 78% (39 positive among 50 new-onset T1DM) and 98% (2 positive among 100 healthy controls). The results of GAD-Ab obtained with micro-plate RBA and RLA were closely correlated (r=0.915,P<0.001) with a high concordance level of 97.5% and a Kappa value of 0.95. (3)TI DM and T2DM patients showed higher positive rates for GAD-Ab than the healthy controls(46.9% and 5.2% vs 0.89% ,X2=123.5 and 10. 1 ,P <0.001 and <0.01, respectively). (4)The consistency of GAD-Ab measurement with RBA using finger tip blood and RLA measurement using venous blood was 96.7% (r =0.946,P <0.001, Kappa value: 0.905). Conclusions The micro-plate RBA of GAD-Ab has high sensitivity, specificity and reproducibility, and can be measured with finger tip blood sampling. It might be a better alternative for clinical practice.

AIMTo observe the effects of Ca2+ -activated K+ channel of primary cultured fetal SD rat cortex neurons in the veratridine triggered neuronal damage.

METHODSThe patch clamp technique of cell-attach and inside-out mode for these two kinds of single channel recordings were used.

RESULTSExtracellular veratridine activated the Kca. In Ca2+ bath solution of cell-attach mode, Vp + 30 mV, when the concentration (micromol/L) of veratridine were 15,25,50 and 75, the open probabilities of the channel were 0.014 +/- 0.003, 0.085 +/- 0.010, 0.132 +/- 0.016 and 0.059 +/- 0.006 (P < 0.01) respectively. It appeared concentration-dependent within 50 micromol/L veratridine. In Ca2+ free bath solution of cell-attach mode, Vp = +50 mV, when the concentration (micromol/L) of veratridine were 15, 40,60 and 100, the open probabilities of the channel were 0.014 +/- 0.010, 0.113 +/- 0.006, 0.141 +/- 0.004 and 0.295 +/- 0.009 (P < 0.05) respectively. In the 6 cases of inside-out mode patch clamp, Vp = +40 mV, when the concentration of veratridine were 0, 25 micromol/L and 50 micromol/L, the open probabilities of the channel were 0.011 +/- 0.008, 0.010 +/- 0.010 and 0.012 +/- 0.007 (P > 0.05) respectively. There were no significant difference on open probabilities, average open/close times and amplitudes at different intracellular veratridine concentration.

CONCLUSIONVeratridine can affect the activation of the Kca channel through regulating the concentration of cytoplasmic free Ca2+. The opening of Kca activated by increase of intracellular Ca2+ during the early stage of anoxia may be a protection reaction of ischemic neurons.

Animals ; Animals, Newborn ; Calcium ; metabolism ; Cells, Cultured ; Neurons ; cytology ; drug effects ; physiology ; Patch-Clamp Techniques ; Potassium Channels, Calcium-Activated ; metabolism ; Rats ; Rats, Sprague-Dawley ; Veratridine ; pharmacology

OBJECTIVETo conduct a meta-analysis to study the effect of antiviral therapy on the prognosis of patients with chronic hepatitis B (CHB)-related cirrhosis.

METHODSPubMed, EMBASE, the Cochrane Central Register of Controlled Trials, Chinese Biomedical Database, Chinese Journals Full-text Database, and Wan Fang Digital Journal Full-text Database were searched for studies on nucleoside analogues antiviral treatment outcome of patients with CHB-related cirrhosis (vs. controls without antiviral therapy) published between January 1998 and March 2012. Data extraction and quality assessment was performed by two independent investigators. Heterogeneity was assessed by the I2 index. In the case of homogeneity the random-effects model was applied, and in the case of heterogeneity the fixed-effects model was applied. The odds ratios (ORs) with 95% confidence intervals (CIs) were calculated.

RESULTSSeven studies were included in the meta-analysis: one high-quality randomized-controlled trial (RCT) study, four prospective cohort studies, and two case-control studies. Compared to the control group, the group treated with antiviral therapy showed a significantly lower incidence of hepatocellular carcinoma (11.2%, 76/680 vs. 6.7%, 75/1116; OR = 0.56, 95% CI: 0.40 to 0.79, P = 0.001) and lower mortality (23.6%, 78/331 vs. 10.8%, 43/398; OR = 0.36, 95% CI: 0.23 to 0.55, P = 0.000).

CONCLUSIONAntiviral therapy with nucleoside analogues significantly reduces the incidence of hepatocellular carcinoma and mortality in patients with CHB-related cirrhosis.

Antiviral Agents ; therapeutic use ; Hepatitis B, Chronic ; complications ; diagnosis ; drug therapy ; Humans ; Liver Cirrhosis ; diagnosis ; drug therapy ; etiology ; Nucleotides ; therapeutic use ; Prognosis

BACKGROUNDCytomegalovirus (CMV) remains a significant clinical problem among immunosuppressed renal transplant patients. Quantitative PCR assays have become the most common methods in the determination of CMV infections in transplant patients. This study was to determine the relationship between CMV infection and the acute rejection of the transplanted kidney.

METHODSPlasma samples from 77 renal transplant patients that were pre-transplant negative for CMV infection were tested using real-time quantitative PCR and CMV gene-specific primers. The detected viral loads were retrospectively compared with the acute rejection rate and the chronic or mild rejection rates of the renal transplant.

RESULTSCMV-DNA was detected in 29 of 77 recipients, yielding a positive rate of detection of 37.7% for this procedure. Twelve of the 21 recipients (57.1%) who suffered acute rejection had positive CMV-DNA. Among the 56 recipients suffered from chronic or mild rejection, 17 (30.4%) had positive CMV-DNA plasma. Moreover, of the 29 recipients who had detectable CMV-DNA after transplant, 12 (41.4%) suffered from acute rejection; of the 48 recipients with undetectable CMV-DNA, only nine (18.8%) developed acute rejection. Post-transplant patients with acute rejection had a higher rate (57.1% vs. 30.4%, P = 0.03) of post-transplant CMV infection than those with chronic or mild rejection.

CONCLUSIONCMV infection is a risk factor of acute renal transplant rejection and CMV infection should be prevented and treated in renal transplant recipients.

Adult ; Cytomegalovirus Infections ; diagnosis ; genetics ; DNA, Viral ; genetics ; Female ; Humans ; Kidney Transplantation ; adverse effects ; Male ; Middle Aged ; Real-Time Polymerase Chain Reaction ; methods ; Viral Load ; Young Adult

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

OBJECTIVEIn the candidate region 12q13 of simple congenital heart disease(CHD), four single nucleotide polymorphisms(SNPs) in HOXC4 gene were chosen in order to investigate the distribution of SNP and haplotypes in simple CHD patients and normal people.

METHODSThe genotype of 4 SNPs in 108 simple CHD patients and 200 normal people were analyzed by restriction fragment length polymorphism(RFLP) and denaturing high-performance liquid chromatography(DHPLC). The statistical contingency table method was used to analyze SNP genotype frequency and gene frequency in patients and control group; then, the haplotypes were established and their frequencies in the two groups were assessed by PHASE software.

RESULTSC16476T polymorphism was not detected; A17860G located in 3' flanking sequence of HOXC5 gene displayed significant difference between the two groups. The G allele frequency in simple CHD patients was higher than that in healthy controls(P < 0.05); the distribution of frequencies of 4 haplotypes showed significant difference(P < 0.01).

CONCLUSIONThe A17860G located in 3'flanking sequence of HOXC5 gene is associated with simple CHD; the risk of CHD in the persons with G17860 is higher than that in those with A17860. the haplotype of 3 SNPs may be linked with the susceptible gene of simple CHD.

Adolescent ; Adult ; Alleles ; Base Sequence ; Child ; Chromosomes, Human, Pair 12 ; genetics ; DNA Mutational Analysis ; Gene Frequency ; Genetic Predisposition to Disease ; genetics ; Genotype ; Haplotypes ; genetics ; Heart Defects, Congenital ; genetics ; Homeodomain Proteins ; genetics ; Humans ; Middle Aged ; Molecular Sequence Data ; Polymorphism, Single Nucleotide ; Young Adult

OBJECTIVE: To find the mutation of disease-causing genes of sudden unexplained death syndrome (SUDS) in the young by whole exome sequencing in one case. METHODS: One SUDS case was found no obvious fatal pathological changes after conventional autopsy and pathological examination. The whole exome sequencing was performed with the Ion Torrent PGM™ System with hg19 as reference sequence for sequencing data. The functions of mutations were analyzed by PhyloP, PolyPhen2 and SIFT. A three-step bioinformatics filtering procedure was carried out to identify possible significative single nucleotide variation (SNV), which was missense mutation with allele frequency < 1% of myocardial cell. RESULTS: Four rare suspicious pathogenic SNV were identified. Combined with the analysis of conventional autopsy and pathological examination, the mutation MYOM2 (8_2054058_G/A) was assessed as high-risk deleterious mutation by PolyPhen2 and SIFT, respectively. CONCLUSION Based on the second generation sequencing technology, analysis of whole exome sequencing can be a new method for the death cause investigation of SUDS. The gene MYOM2 is a new candidate SUDS pathogenic gene for mechanism research.
Autopsy ; Brugada Syndrome/genetics* ; Cause of Death ; DNA Mutational Analysis/methods* ; Death, Sudden/etiology* ; Exome ; Gene Frequency ; Genetic Testing/methods* ; High-Throughput Nucleotide Sequencing/methods* ; Humans ; Molecular Biology ; Molecular Diagnostic Techniques/methods* ; Molecular Sequence Data ; Mutation

Objective During June 2-8, 2009, 11 cases of the novel influenza A (H1N1)occurred in Sichuan Province, China. We investigated this outbreak to identify the source of infection,mode of transmission and risk factors for infection. Methods The primary case, a U.S. citizen,developed disease on June 2. From June 3 to 5, she joined Tour Group A for a trip to Jiuzhaigou. We telephoned passengers of the three flights on which the primary case had traveled in China, and members of Tour Group A. We asked whether they had any influenza-like symptoms during May 27 to June 12. Health authorities placed passengers whose seats were within three rows of the primary case on flights and members of Tour Group A on medical observation, and isolated individuals if they developed symptoms. We used real-time RT-PCR to test the throat swabs from symptomatic persons for the novel influenza virus and defined a confirmed case as one with influenza-like symptoms and laboratory confirmation. A retrospective cohort investigation to identify the risk factors for infection was conducted. We interviewed all members of Tour Group A about their detailed contact history with the primary case. Results During June 5 to 6, 9 (30%) of the primary case' s 30 fellow tour group members developed disease, compared with none of her 87 fellow passengers to Jiuzhaigou and 1 of her 87 fellow passengers on the returning trip (when several of the members of Tour Group A were symptomatic). 56% of the tourists who had talked with the primary case in close range ( ＜2 m) for ≥2 minutes developed disease, whereas none of the 14 other tour group members developed disease (RR= ∞; exact 95%CI: 2.0- ∞ ). Having conversed with the primary case for ≥ 10 minutes (vs. 2-9 minutes) increased the risk by almost five fold (RR=4.8, exact 95%CI: 1.3-180). Conversely, other kinds of contact, such as dining at the same table, receiving chewing gum from the primary case and sharing bus rides or planes with the primary case played no roles during this outbreak. Conclusion This novel influenza A (H1N1) outbreak was caused by an imported case, and transmitted mainly via droplet transmission when the primary case was talking with her fellow tourists during a tour. These findings highlight the importance of preventing droplet transmission during a pandemic.

INTRODUCTIONBased on the estimate results of the capacity and preparedness of Beijing hospitals to respond to pandemic influenza, using flu surge model to evaluate its applicable hypothesis and to provide government with sentient strategy in planning pandemic influenza. Through collection of medical resources information, we calculated the possible impaction on hospitals by Flu Surge model and explored the applicable hypothesis in model operation through a questionnaire, direct observation and group discussion in 3 hospitals in Beijing. Based on flu surge model estimation during a 6-week epidemic from a pandemic virus with 35% attack rate, Beijing would have had an estimation of 5 383 000 influenza illnesses, 2 691 500 influenza outpatients, 76 450 influenza hospitalizations and 14 508 excess deaths. For a 6-week period with 35% attack rate, there would be a peak demand for 8% of beds, 210% of ICU beds, and 128% of ventilators estimated. Outpatients in different level hospital were quite disproportionated with 1742/ hospital/day, 650/hospital/day, and 139/hospital/day respectively. The sampled health workers had a mastery of 63.4% of the total knowledge and skills of diagnosing and treating of influenza, 73.5% of them washed their hands and 63.5% used PPE correctly. The total beds capacity, medical beds capacity and respiratory medical beds capacity would increase 8%, 35% and 128% respectively.

CONCLUSIONThe estimation results could be referenced when planning the pandemic strategy, but the results should be treated objectively when considering the hypothesis and practical situation in this model being used.

Adolescent ; Adult ; Aged ; Child ; Child, Preschool ; Disease Outbreaks ; statistics & numerical data ; Female ; Hospital Bed Capacity ; Hospital Planning ; Hospitalization ; statistics & numerical data ; Humans ; Infant ; Infant, Newborn ; Influenza, Human ; epidemiology ; Male ; Middle Aged ; Models, Statistical ; Surge Capacity ; Young Adult

Objective To provide evidence-based recommendations to the government on strategies for reducing the impact of the imminent influenza pandemic, we conducted a survey on the personal protective and healthcare seeking behaviors of Beijing residents in the wake of the worldwide epidemic of the novel 2009 H1N1 influenza virus, and potential changes in these behaviors during a full-blown pandemic. Methods We used a two-stage Mitofsky-Waksberg telephone survey to collect information for Beijing residents ≥ 18 years of age and weighted the sample by the census estimate of Beijing population for 2008. A structured questionnaire was used to collect information about the respondents' knowledge regarding the novel influenza virus, current personal protective and healthcare seeking behaviors during a full-blown influenza pandemic. Results 286 Beijing urban residents were interviewed during May 15 to 18 (response rate: 62%). 77%(95%CI: 71%-82%) of the residents that they would cover their sneezes at home, and 93%(95%CI: 89%-96%) would do so in a public place. Of the residents would cover their sneezes at home, 66% (95%CI: 57%-74%) would covered their sneezes by hand, 33% (95%CI:25%-42%) by facial tissue, and only 0.4% (95%CI: 0.1%-2.9%) by sleeves. Similarly, of the residents would cover their sneezes at a public place, 64% (95%CI: 56%-71%) would cover their sneezes by hand, 33%(95%CI: 26%-41%) by facial tissue and 0% by sleeves. 46% (95% CI: 32%-59%) of the residents would wash their hands after covering their sneezes with hand in public places, and 77% (95%CI: 64%-89%) would do so at home. Higher percentages of residents would wear a mask (63%, 95% CI: 56%-70%) and get vaccinated (43%, 95% CI: 37%-50%) during a pandemic than under currently situation (3.5%, 95% CI: 1.7%-7.0% for wearing a mask; 13%, 95% CI: 9.8%-18% for getting vaccinated). 86% (95% CI: 81%-89%) of the residents would visit a doctor for an influenza-like illness during a pandemic, similar to the percentage currently seen (76%, 95%CI: 70%-81%). 71%(95%CI: 63%-78%) of the residents would visit a doctor for influenza-like illness and would choose a tertiary medical center; with percentage (74% , 95% CI: 66%-80%) similar to the one during a pandemic. Conclusion Beijing residents need to be aware of the proper ways to cover their noses when sneezing (especially using their sleeves more), and to wash their hands. An effective plan to triage patients should be immediately established to efficiently utilize the limited healthcare resources, which would likely be further strained during a pandemic.