Propagation and immunological characterization of coxsackievirus A10 in a serum-free HEK293A cell culture system.

Coxsackievirus A10 (CVA10) is one of enteroviral pathogens that cause the hand, foot, and mouth disease (HFMD). Since CVA10 was reported to be not easily propagated in the Vero cell culture, a feasible manufacture process for producing formalin-inactivated CVA10 vaccine is urgently needed. Several cell lines that commonly used for viral vaccine production was tested for CVA10 (M2014 strain) culture in this study, and our result showed that CVA10 could be easily propagated in the HEK293A cells. A serum-free HEK293A cell culture system was developed for CVA10 production and the yields have reached over 108 TCID50/mL. The biochemical and immunogenic properties of CVA10 particles obtained from this serum-free HEK293A culture were identical to our previous study. Two major particles of CVA10 were separated by ultracentrifugation, and only the infectious mature particles were capable of inducing CVA10 neutralizing antibody responses in the mouse immunogenicity studies. Additionally, we found that coxsackievirus A6 and enterovirus A71 could also be easily propagated using this serum-free HEK293A cell culture system. Our results provide a solution to overcome the obstacle in the propagation of CVA10 and facilitate the development of multivalent vaccines for prevention of HFMD.

Mechanistic analysis of carbon-carbon bond formation by deoxypodophyllotoxin synthase.

Deoxypodophyllotoxin contains a core of four fused rings (A to D) with three consecutive chiral centers, the last being created by the attachment of a peripheral trimethoxyphenyl ring (E) to ring C. Previous studies have suggested that the iron(II)- and 2-oxoglutarate-dependent (Fe/2OG) oxygenase, deoxypodophyllotoxin synthase (DPS), catalyzes the oxidative coupling of ring B and ring E to form ring C and complete the tetracyclic core. Despite recent efforts to deploy DPS in the preparation of deoxypodophyllotoxin analogs, the mechanism underlying the regio- and stereoselectivity of this cyclization event has not been elucidated. Herein, we report 1) two structures of DPS in complex with 2OG and (±)-yatein, 2) in vitro analysis of enzymatic reactivity with substrate analogs, and 3) model reactions addressing DPS's catalytic mechanism. The results disfavor a prior proposal of on-pathway benzylic hydroxylation. Rather, the DPS-catalyzed cyclization likely proceeds by hydrogen atom abstraction from C7', oxidation of the benzylic radical to a carbocation, Friedel-Crafts-like ring closure, and rearomatization of ring B by C6 deprotonation. This mechanism adds to the known pathways for transformation of the carbon-centered radical in Fe/2OG enzymes and suggests what types of substrate modification are likely tolerable in DPS-catalyzed production of deoxypodophyllotoxin analogs.

Association of Traumatic Injury With Adverse Pregnancy Outcomes in Taiwan, 2004 to 2014.

Importance: Literature has suggested that trauma among pregnant women is associated with an increased risk of adverse pregnancy outcomes. However, limited research has investigated the association of trauma with adverse pregnancy outcomes by using a national data set. Objective: To investigate the association between traumatic injury and adverse pregnancy outcomes. Design, Setting, and Participants: This population-based cohort study of pregnant women in Taiwan linked 3 data sets, the Taiwan Birth Registry, Household Registration Information, and National Health Insurance Research Database, from January 1, 2004, through December 31, 2014. Data, including the characteristics of pregnant women and infants, were extracted from the Taiwan Birth Registry data set; to obtain trauma data, this data set and the Household Registration Information data set were collectively linked to National Health Insurance Research Database data. The combined data set was analyzed from January to July 2019. Adverse pregnancy outcomes and premature delivery were defined using International Classification of Diseases, Ninth Revision, Clinical Modification codes. Exposures: The primary exposures of this study were 2 clinical variables related to injury during pregnancy: medical treatment in the emergency department (yes or no) and hospitalization (yes or no). Main Outcomes and Measures: The main outcome variable was adverse pregnancy outcomes, and the secondary outcome variable was premature delivery. Multivariate logistic regression models were used to investigate the association of injuries with adverse pregnancy outcomes after controlling for demographic characteristics and other pregnancy-related variables. Results: A total of 2 973 831 pregnant women (2 475 805 [83.3%] aged 20-34 years) were enrolled between 2004 and 2014, of whom 59 681 (2.0%) sought medical treatments due to injuries. Results of multivariate logistic regression models showed that women receiving emergency treatments more than once were 1.08 times as likely (adjusted odds ratio, 1.08; 95% CI, 1.05-1.10) to have adverse pregnancy outcomes than women who received no emergency treatment. Women with injury-related hospitalization were 1.53 times more likely (adjusted odds ratio, 1.53; 95% CI, 1.41-1.65) to have adverse pregnancy outcomes than women who did not sustain injuries. Furthermore, recurrent injuries were associated with a 572% increase in odds of premature delivery (adjusted odds ratio, 6.72; 95% CI, 2.86-15.80). Conclusions and Relevance: In this study, trauma among pregnant women was associated with an increased risk of adverse pregnancy outcomes, as were hospitalization and emergency department visits due to injury.

Development of a fall-risk assessment profile for community-dwelling older adults by using the National Health Interview Survey in Taiwan.

BACKGROUND: Falls represent a global health issue among older adults and cause a considerable burden on medical systems. In this study, a fall-risk assessment profile was developed for community-dwelling older adults. METHOD: The data of survey participants aged > 65 years were obtained from three rounds (2005, 2009, and 2013) of the National Health Interview Survey in Taiwan. In total, 8356 older participants were included in this study. Logistic regression analyses were used to determine potential predictors associated with falls. The regression coefficients of the predictors in the final model were translated into scores (by multiplying by 5) and then summed to obtain a total risk-score for falls. A receiver operating characteristic (ROC) curve was used to evaluate the discriminative performance of the risk assessment profile. RESULT: Self-reported falls within 1 year accounted for 19.1% of the total falls. The predictors that were included in the risk profile according to the logistic regression analysis results were as follows: female sex (adjusted odds ratio = 1.57; risk-score = 2), living alone (adjusted odds ratio = 1.56; risk-score = 2), urinary incontinence (adjusted odds ratio = 1.36; risk-score = 2), perceived unhealthiness (adjusted odds ratio = 1.32; risk-score = 1), perceived pain (adjusted odds ratio = 1.51; risk-score = 2), hospital admission in the past year (adjusted odds ratio = 2.42; risk-score = 4), low activity of daily living (ADL) scores (adjusted odds ratio = 1.29; risk-score = 1), and low mobility function scores (adjusted odds ratio = 1.68; risk-score = 3). At a total risk-score cutoff point of 6 (range 0-17), the model predicted falls with a sensitivity and specificity of 75.16 and 52.75%, respectively (area under the ROC curve = 0.70). CONCLUSION: The fall-risk assessment profile comprising eight predictors-female sex, living alone, incontinence, perceived unhealthiness, perceived pain, hospital admission in the past year, low ADL scores, and low mobility function scores-may serve as an assessment tool for identification of older adults with a high risk of falling, and assessment results can be used to facilitate community-based intervention.

Comparison of traffic-injury related hospitalisation between bicyclists and motorcyclists in Taiwan.

OBJECTIVES: Bicyclists and motorcyclists contribute substantially to the morbidity and mortality rates of road crash casualties. The objective of the study was to investigate the crash characteristics of bicyclist and motorcyclist casualties presented to hospitals in Taiwan resulting from crashes. METHODS: By using linked data from The National Traffic Crash Dataset and the National Health Insurance Database between 2003 and 2012, logistic regression models were used to examine the determinants of hospitalisation among motorcyclist and bicyclist casualties. The examined variables include demographic characteristics, road and weather conditions, and vehicle characteristics. RESULTS: A total of 1,998,606 two-wheelers were enrolled in the study, of whom 216,600 were hospitalised: 203,623 were motorcyclists and 12,964 were bicyclists. Bicyclists were more likely to be hospitalised than motorcyclists were (14.0% vs. 10.7%). The pooled logistic regression model shows that bicyclists had higher odds of hospitalisation than motorcyclists (adjusted odds ratio [AOR] = 1.11, 95% confident interval [CI] = 1.08-1.14). In the motorcyclist and bicyclist models, helmet non-use appears to be a determinant of hospitalisation for motorcyclists (AOR = 1.14, CI = 1.12-1.16), although insignificant for cyclists (AOR = 1.03, CI = 0.94-1.12). Other important determinants of hospitalisation for motorcyclists and cyclists include female riders, elderly riders, rural roadways, unlicensed riding (for motorcyclists only), curved roadways, defective roadways, alcohol consumption (only for motorcyclists), and single-vehicle crashes (for motorcyclists only). CONCLUSIONS: The result that bicyclists had an increased probability of being hospitalised than motorcyclists is particularly noteworthy, because there have recently been much more users of bikesharing systems in metropolitan cities where cycle helmets are not provided. We further found that helmet non-use was also a risk factor for motorcyclists, but insignificant for cyclists, possibly due to lower helmet utilization rates among bicyclists. Our findings regarding the increased hospitalisation percentage emphasize the importance of helmet use.

Enhancing enterovirus A71 vaccine production yield by microcarrier profusion bioreactor culture.

Hand, foot and mouth diseases (HFMD) are mainly caused by Enterovirus A71 (EV-A71) infections. Clinical trials in Asia conducted with formalin-inactivated EV-A71 vaccine candidates produced from serum-free Vero cell culture using either roller bottle or cell factory technology, are found to be safe and highly efficacious. To increase vaccine yields and reduce the production costs, the bioprocess improvement for EV-A71 vaccine manufacturing is currently being investigated. The parameters that could affect and enhance the production yields of EV-A71 virus growth in the microcarrier bioreactor were investigated. The medium replacement culture strategy included a multi-harvested semi-batch process and perfusion technology and was found to increase the production yields more than 7-14 folds. Based on the western blot and cryo-EM analyses of the EV-A71 virus particles produced from either the multi-harvested semi-batch (MHSBC) or perfusion cultures were found to be similar to those virus particles obtained from the single batch culture. Mouse immunogenicity studies indicate that the EV-A71 vaccine candidates produced from the perfusion culture have similar potency to those obtained from single batch bioprocess. The physical structures of the EV-A71 particles revealed by the cryo-EM analysis were found to be spherical capsid particles. These results provide feasible technical bioprocesses for increasing virus yields and the scale up of EV-A71 vaccine manufacturing using the bioreactor cell culture methods.

A population-based case-control study of hospitalisation due to head injuries among bicyclists and motorcyclists in Taiwan.

INTRODUCTION: According to official statistics in Taiwan, the main body region of injury causing bicyclist deaths is the head, and bicyclists are 2.6 times more likely to be fatally injured than motorcyclists. There is currently a national helmet law for motorcyclists but not for bicyclists. OBJECTIVES: The primary aim of this study was to determine whether bicyclist casualties have higher odds of head-related hospitalisation than motorcyclists. This study also aims to investigate the determinants of head injury-related hospitalisation among bicyclists and motorcyclists. METHODS: Using linked data from the National Traffic Accident Dataset and the National Health Insurance Research Database for the period 2003-2012, this study investigates the crash characteristics of bicyclist and motorcyclist casualties presenting to hospitals due to motor vehicle crashes. Head injury-related hospitalisation was used as the study outcome for both road users to evaluate whether various factors (eg, human attributes, road and weather conditions, vehicle characteristics) are related to hospital admission of those who sustained serious injuries. RESULTS: Among 1 239 474 bicyclist and motorcyclist casualties, the proportion of bicyclists hospitalised for head injuries was higher than that of motorcyclists (10.0% vs 6.5%). However, the multiple logistic regression model shows that, after adjustment of this result for other factors such as helmet use, bicyclists were 18% significantly less likely to be hospitalised for head injuries than motorcyclists (AOR 0.82, 95% CI 0.79 to 0.85). Other important determinants of head injury-related hospitalisation for bicyclists and motorcyclists include female riders, elderly riders, crashes occurring in rural areas, moped riders, riding unhelmeted, intoxicated bicyclists and motorcyclists, unlicensed motorcyclists, dusk and dawn conditions and single-vehicle crashes. CONCLUSIONS: Our finding underscores the importance of helmet use in reducing hospitalisation due to head injuries among bicyclists while current helmet use is relatively low.

Immunological and biochemical characterizations of coxsackievirus A6 and A10 viral particles.

Childhood exanthema caused by different serotypes of coxsackievirus (CV-A) and enterovirus A71 (EV-A71) has become a serious global health problem; it is commonly known as hand, foot, and mouth disease (HFMD). Current EV-A71 vaccine clinical trials have demonstrated that human antibody responses generated by EV-A71 vaccinations do not cross-neutralize coxsackievirus A16 (CV-A16). An effective multivalent HFMD vaccine is urgently needed. From molecular epidemiological studies in Southeast Asia, CV-A6 and CV-A10 are commonly found in HFMD outbreaks. In this study, CV-A6 and CV-A10 were individually cultured in rhabdomyosarcoma (RD) cells grown in medium containing serum, harvested and concentrated. In viral downstream purification, two viral fractions were separated by sucrose gradient zonal ultracentrifugation and detected using a SDS-PAGE analysis and a virus infectivity assay. These two viral fractions were formalin-inactivated, and only the infectious particle fraction was found to be capable of inducing CV-A serotype-specific neutralizing antibody responses in animal immunogenicity studies. These mouse and rabbit antisera also failed to cross-neutralize EV-A71 and CV-A16 infections. Only a combination of formalin-inactivated EV-A71, CV-A6, CV-A10 and CV-A16 multivalent vaccine candidates elicited cross-neutralizing antibody responses in both mouse and rabbit immunogenicity studies. The current results certainly provide important information for multivalent HFMD vaccine development.

First Fall-Related Injuries Requiring Hospitalization Increase the Risk of Recurrent Injurious Falls: A Nationwide Cohort Study in Taiwan.

OBJECTIVES: Recurrent falls not only have risk factors different from those of single falls but also have less favorable outcomes. The aim of our study was to determine the association between the injury characteristics of a first fall and the likelihood of recurrent injurious falls in a cohort of hospitalized patients. METHODS: We designed a nationwide retrospective cohort study and selected hospitalized patients who had injurious falls between 2001 and 2010. Cox proportional hazards models were used to estimate the hazard ratios (HRs) of recurrent injurious falls requiring hospitalization in the following year on the basis of the patients' demographic characteristics, comorbidities, and the characteristics of injuries from the first injurious fall requiring hospitalization. RESULTS: Among the 504 512 patients hospitalized for injurious falls, 19 442 experienced recurrent injurious falls requiring hospitalization. The 1-year incidence of recurrent injurious falls requiring hospitalization was 3.85%. The incidence density was the highest within the 3-month period after the first injurious fall. The risk of recurrent injurious falls among patients aged 40 to 64, 65 to 74, and ≥ 75 years increased progressively (HR: 2.11, 95% confidence interval [CI]: 1.90-2.34; HR: 2.80, 95% CI: 2.51-3.11; and HR: 3.80, 95% CI: 3.42-4.23, respectively). The length of hospitalization (LOH) ≥ 15 or ≥ 31 days (HR: 1.39, 95% CI: 1.30-1.48; and HR: 1.59, 95% CI: 1.43-1.77, respectively) and injury to the head (HR: 1.59, 95% CI: 1.53-1.65) or spine (HR: 1.66, 95% CI: 1.59-1.74) were also found to be major risk factors. CONCLUSIONS: Our findings show that the LOH and head and spine injuries are associated with an increased risk of recurrent injurious falls leading to hospitalization. The risk of recurrent injurious falls requiring hospitalization increased significantly among adults older than 40 years. We suggest further research on the effects of injury characteristics associated with the first injurious fall requiring hospitalization and resultant anatomical damages on the risk of recurrent injurious falls requiring hospitalization. High-risk patients should receive tailored rehabilitation addressing their respective injuries within 3 months after hospital discharge.

Recombinant adeno-vaccine expressing enterovirus 71-like particles against hand, foot, and mouth disease.

Enterovirus 71 (EV71) and coxsackieviruses (CV) are the major causative agents of hand, foot and mouth disease (HFMD). There is not currently a vaccine available against HFMD, even though a newly developed formalin-inactivated EV71 (FI-EV71) vaccine has been tested in clinical trial and has shown efficacy against EV71. We have designed and genetically engineered a recombinant adenovirus Ad-EVVLP with the EV71 P1 and 3CD genes inserted into the E1/E3-deleted adenoviral genome. Ad-EVVLP were produced in HEK-293A cells. In addition to Ad-EVVLP particles, virus-like particles (VLPs) formed from the physical association of EV71 capsid proteins, VP0, VP1, and VP3 expressed from P1 gene products. They were digested by 3CD protease and confirmed to be produced by Ad-EVVLP-producing cells, as determined using transmission electron microscopy and western blotting. Mouse immunogenicity studies showed that Ad-EVVLP-immunized antisera neutralized the EV71 B4 and C2 genotypes. Activation of VLP-specific CD4+ and CD8+/IFN-γ T cells associated with Th1/Th2-balanced IFN-ɣ, IL-17, IL-4, and IL-13 was induced; in contrast, FI-EV71 induced only Th2-mediated neutralizing antibody against EV71 and low VLP-specific CD4+ and CD8+ T cell responses. The antiviral immunity against EV71 was clearly demonstrated in mice vaccinated with Ad-EVVLP in a hSCARB2 transgenic (hSCARB2-Tg) mouse challenge model. Ad-EVVLP-vaccinated mice were 100% protected and demonstrated reduced viral load in both the CNS and muscle tissues. Ad-EVVLP successfully induced anti-CVA16 immunities. Although antisera had no neutralizing activity against CVA16, the 3C-specific CD4+ and CD8+/IFN-γ T cells were identified, which could mediate protection against CVA16 challenge. FI-EV71 did not induce 3C-mediated immunity and had no efficacy against the CVA16 challenge. These results suggest that Ad-EVVLP can enhance neutralizing antibody and protective cellular immune responses to prevent EV71 infection and cellular immune responses against CV infection.

Effect of weather and time on trauma events determined using emergency medical service registry data.

INTRODUCTION: Trauma admissions are associated with weather and temporal factors; however, previous study results regarding these factors are contradictory. We hypothesised that weather and temporal factors have different effects on specific trauma events in an emergency medical service (EMS) system. METHODS: EMS data from January 1, 2009, to December 31, 2010, were obtained from the fire department of Taipei City and associated with the local weather data. EMS trauma events were categorised into total trauma, traffic accidents (TAs), motorbike accidents (MBAs), and falls. Hourly data on trauma patients were analysed using the zero-inflated Poisson model. RESULTS: The hourly incidence of total trauma increased with the magnitude of precipitation (incidence rate ratio [IRR]=1.06, 1.09, and 1.11 in light, moderate, and heavy rain, respectively), and this effect was more prominent in fall patients than in patients with other injuries (IRR=1.07, 1.21, and 1.32). However, the hourly incidence of TAs and MBAs was associated only with light rain (IRR=1.11 and 1.06, respectively). An hour of sunshine exposure was associated with an increase in the hourly incidence of all groups, and higher temperatures were associated with an increased hourly incidence of total trauma, TAs, and MBAs, but not falls. The hourly incidence of falls increased only in late fall and winter. Compared with the hourly incidence between 3 am and 7 am, the hourly incidence of all groups plateaued between 7 am and 11 pm and declined from 11 pm to 3 am. During the plateau period, 2 peaks in the incidence of TAs (IRR=5.03 and 5.07, respectively) and MBAs (IRR=5.81 and 5.51, respectively) were observed during 7-11 am and 3-7 pm. The hourly incidence of total trauma, TAs, and MBAs plateaued during workdays, peaked on Fridays, declined on Saturdays, and troughed on Sundays. The incidence of falls increased only on Mondays (IRR=1.09). CONCLUSIONS: Weather and temporal factors had different impacts on the incidence of traffic-related accidents and falls. Therefore, EMS data may have implications in preventing injuries and planning resource use for prehospital trauma rescue.

Atom-to-atom interactions for atomic layer deposition of trimethylaluminum on Ga-rich GaAs(001)-4 × 6 and As-rich GaAs(001)-2 × 4 surfaces: a synchrotron radiation photoemission study.

High-resolution synchrotron radiation photoemission was employed to study the effects of atomic-layer-deposited trimethylaluminum (TMA) and water on Ga-rich GaAs(001)-4 × 6 and As-rich GaAs(001)-2 × 4 surfaces. No high charge states were found in either As 3d or Ga 3d core-level spectra before and after the deposition of the precursors. TMA adsorption does not disrupt the GaAs surface structure. For the (4 × 6) surface, the TMA precursor existed in both chemisorbed and physisorbed forms. In the former, TMA has lost a methyl group and is bonded to the As of the As-Ga dimer. Upon water purge, the dimethylaluminum-As group was etched off, allowing the now exposed Ga to bond with oxygen. Water also changed the physisorbed TMA into the As-O-Al(CH3)2 configuration. This configuration was also found in 1 cycle of TMA and water exposure of the (2 × 4) surface, but with a greater strength, accounting for the high interface defect density in the mid-gap region.

A population-based study on the association between gastric ulcers and erectile dysfunction in Taiwan.

INTRODUCTION: While erectile dysfunction (ED) and cardiovascular disease have long been known to share endothelial dysfunction as a common contributory underlying mechanism, little research has been conducted taking endothelial dysfunction as common ground to investigate the potential association between ED and gastric ulcers (GUs). AIM: This population-based case-control study aimed to investigate the association of ED with GU. METHODS: This study used data from the Longitudinal Health Insurance Database 2000 in Taiwan. The study group comprised 6,906 patients who visited ambulatory care centers or were hospitalized with a diagnosis of ED. The comparison group was 20,718 randomly selected enrollees. Conditional logistic regression was used to examine associations between ED and prior GU. MAIN OUTCOME MEASURE: The prevalence and risk between cases and controls were calculated of having been previously diagnosed with GU. RESULTS: Of the sampled subjects, 3,861 (14%) were diagnosed before the index date, 1,358 (19.7%) were cases, and 2,503 (12.1%) were controls (P<0.001). After adjusting for hypertension, diabetes, hyperlipidemia, renal disease, coronary heart disease, obesity, alcohol abuse/alcohol dependence syndrome, and socioeconomic status (SES), conditional logistic regression analysis revealed that cases were more likely to have been diagnosed with GU than controls (odds ratio [OR]=1.65, 95% confidence interval [CI]=1.53-1.77). Stratification by age revealed that the youngest group (18-29) of ED patients had the most increased likelihood of having been previously diagnosed with GU when compared with matched controls (OR=4.12, 95% CI=2.41-7.03). The likelihood decreased with age, with the oldest group of ED patients having the least increased likelihood of prior GU when compared with matched controls (OR=1.44, 95%CI=1.23-1.68). CONCLUSION: Our findings suggest a positive association between prior GU and a subsequent diagnosis with ED.

Identification and characterization of a cross-neutralization epitope of Enterovirus 71.

Enterovirus 71 (EV71) infections in children manifest as exanthema and are most commonly known as hand-foot-and-mouth disease (HFMD). Because it can cause severe neurological complications like poliomyelitis, EV71 has now emerged as an important neurotropic virus in Asia. EV71 virus has been shown to consist of 3 (A, B and C) genotypes and many subgenotypes. Although EV71 vaccine development has recently yielded promising preclinical results, yet the correlation between the content of antigen(s) in vaccine candidates and the level of protective antibody responses is not established. The neutralization epitope(s) of EV71 antigens could be used as the surrogate biomarker of vaccine potency. Using peptide ELISA, antisera generated from animals immunized with formalin-inactivated EV71 virion vaccine formulated in alum, EV71-specific neutralizing monoclonal antibody (nMAb) and a panel of 153 overlapping synthetic peptides covering the entire sequences of VP1, VP2 and VP3 of EV71, we screened for immunodominant linear neutralization epitope(s). Synthetic peptide VP2-28, corresponding to residues 136-150 of VP2, was found to bind to and inhibit the binding to EV71 of nMAb MAB979 that was found to have cross-neutralizing activity against different genotypes of EV71 virus. In addition, VP2-28 was found to be recognized only by neutralizing antisera generated from rabbits immunized with the formalin-inactivated whole EV71 virion vaccine but not by antisera from immunized mice and rats. During the epitope mapping, a murine EV71 genotype- and strain-specific linear neutralization epitope VP1-43 was identified within residues 211-220 of VP1. Furthermore, based on sequence alignment and structure prediction analysis using poliovirus as the template for molecular modeling, the VP1-43 and VP2-28 epitopes were shown to run in parallel within 0.1 nm and form a rim of the canyon at the junction site of VP1 and VP2 in the viral capsid. In mouse, rat and rabbit immunogenicity studies, a dose-dependent relationship between the number of VP2-28 epitope units measured by a quantitative assay in vaccine preparations and the magnitude of neutralizing titers was demonstrated. VP2-28 has amino acid sequences that are highly conserved among EV71 genotypes, is not affected by formalin-treatment and long-term storage. Thus, VP2-28 could be used as the surrogate biomarker in the potency testing of candidate EV71 vaccines.

The effect of ankle position on pelvic floor muscle contraction activity in women.

PURPOSE: Stress urinary incontinence is extremely common in women. Conservative management includes pelvic floor muscle exercises to increase strength and muscular contraction forces to improve incontinence. We examined the effects of passive and active ankle flexion on pelvic floor muscle activity during exercises. MATERIALS AND METHODS: A total of 31 married women performed random pelvic floor muscle contractions while standing, and in 8 passive and active positions while pelvic floor muscle activity was simultaneously measured by electromyography. RESULTS: All ankle positions resulted in greater pelvic floor muscle activity than the horizontal foot position. Significantly greater muscle activity was seen with ankles in the plantar position with raised arms (p = 0.0051). CONCLUSIONS: Pelvic floor muscle exercises performed with active ankle positions may increase the effectiveness of these exercises. Further studies are required to elucidate mechanisms for this finding.

Gastrointestinal stromal tumor of the prostate: a case report and literature review.

We report a case of gastrointestinal stromal tumor (GIST) of the prostate in a 75-year-old man with dysuria and urinary retention. Digital rectal examination revealed a markedly enlarged prostate. The serum level of prostate-specific antigen was 0.2 ng/dL. Imaging studies showed an expanded prostate, measuring 6.7 x 5.6 x 5.5 cm, with heterogeneity in contrast enhancement. No metastatic disease was found. The pathologic diagnosis of prostatic GIST was made based on characteristic morphological features, immunoprofiles, and molecular analysis. The possibility of secondary involvement by a rectal GIST was excluded by radiological and intraoperative findings. To our knowledge, this is the second case of a primary GIST of the prostate reported in the literature.

Continuous monitoring of enzyme reactions on a microchip: application to catalytic RNA self-cleavage.

Kinetic analysis of RNA enzymes, or ribozymes, typically involves the tedious process of collecting and quenching reaction time points and then fractionating by polyacrylamide gel electrophoresis (PAGE). As a way to automate and simplify this process, continuous analysis of a ribozyme reaction is demonstrated here using completely automated capillary sample introduction onto a microfabricated device with laser-induced fluorescence detection. The method of injection is extremely reproducible thereby standardizing data analysis. A 30-nucleotide ribozyme model, the self-cleaving lead-dependent ribozyme, or "leadzyme", which cleaves into a 24-mer and a 6-mer in the presence of Pb(2+), was end-labeled with fluorescein (FAM) and used to demonstrate the potential of this technique. After manually initiating the cleavage reaction by Pb(2+) addition, reaction samples were automatically injected directly into the parallel separation lanes of the chip via a capillary at predetermined time intervals, thus eliminating the need for additional sample-handling steps. The FAM-labeled leadzyme starting material and products were monitored for 60 min in order to ascertain kinetic information. The effect of lead acetate concentration on cleavage rates was also studied, and the results are in agreement with rates determined by conventional hand-mixing/PAGE analysis. This work demonstrates, through the use of a simple ribozyme model, the potential of this method to provide valuable kinetic information for other, more complex, biologically relevant RNA and protein enzymes.