Size variations in synaptic terminals among different types of photoreceptors and across the zebrafish retina.

Photoreceptor synaptic terminals are responsible for transmitting visual information to downstream neurons. In vertebrate retinas, photoreceptor synaptic terminals are of different sizes and structures. The molecular mechanisms that underlie photoreceptor synaptic development are not clearly understood. Here, we have systematically examined the size variations in the synaptic terminals of cone and rod photoreceptors in the adult zebrafish retina. We reveal that the average cone pedicle sizes expand in the order of UV, blue, green, and red cones, echoing the increasing maximally sensitive wavelengths of the opsins expressed in the corresponding cone types. In addition, rod spherules are smaller than all cone pedicles. The terminals of each photoreceptor type also display distinct regional variations across the retina and between males and females. These findings establish the basis for using the zebrafish retina to study the molecular mechanisms that regulate the sizes and structures of photoreceptor terminals for proper visual functions.

Developmental and Neurotoxicity of Acrylamide to Zebrafish.

Acrylamide is a commonly used industrial chemical that is known to be neurotoxic to mammals. However, its developmental toxicity is rarely assessed in mammalian models because of the cost and complexity involved. We used zebrafish to assess the neurotoxicity, developmental and behavioral toxicity of acrylamide. At 6 h post fertilization, zebrafish embryos were exposed to four concentrations of acrylamide (10, 30, 100, or 300 mg/L) in a medium for 114 h. Acrylamide caused developmental toxicity characterized by yolk retention, scoliosis, swim bladder deficiency, and curvature of the body. Acrylamide also impaired locomotor activity, which was measured as swimming speed and distance traveled. In addition, treatment with 100 mg/L acrylamide shortened the width of the brain and spinal cord, indicating neuronal toxicity. In summary, acrylamide induces developmental toxicity and neurotoxicity in zebrafish. This can be used to study acrylamide neurotoxicity in a rapid and cost-efficient manner.

A Phenotypic and Genotypic Evaluation of Developmental Toxicity of Polyhexamethylene Guanidine Phosphate Using Zebrafish Embryo/Larvae.

Polyhexamethylene guanidine-phosphate (PHMG-P), a guanidine-based cationic antimicrobial polymer, is an effective antimicrobial biocide, potent even at low concentrations. Due to its resilient bactericidal properties, it has been used extensively in consumer products. It was safely used until its use in humidifiers led to a catastrophic event in South Korea. Epidemiological studies have linked the use of PHMG-P as a humidifier disinfectant to pulmonary fibrosis. However, little is known about its harmful impacts other than pulmonary fibrosis. Thus, we applied a zebrafish embryo/larvae model to evaluate developmental and cardiotoxic effects and transcriptome changes using RNA-sequencing. Zebrafish embryos were exposed to 0.1, 0.2, 0.3, 0.4, 0.5, 1, and 2 mg/L of PHMG-P from 3 h to 96 h post fertilization. 2 mg/L of PHMG-P resulted in total mortality and an LC50 value at 96 h was determined at 1.18 mg/L. Significant developmental changes were not observed but the heart rate of zebrafish larvae was significantly altered. In transcriptome analysis, immune and inflammatory responses were significantly affected similarly to those in epidemiological studies. Our qPCR analysis (Itgb1b, TNC, Arg1, Arg2, IL-1ß, Serpine-1, and Ptgs2b) also confirmed this following a 96 h exposure to 0.4 mg/L of PHMG-P. Based on our results, PHMG-P might induce lethal and cardiotoxic effects in zebrafish, and crucial transcriptome changes were linked to immune and inflammatory response.

A diaminomaleonitrile-appended BODIPY chemosensor for the selective detection of Cu2+ via oxidative cyclization and imaging in SiHa cells and zebrafish.

A specific Cu2+ sensor, 2-amino-3-(BODIPYmethyleneamino)maleonitrile (BDM), was established by a simple dehydration between BODIPY and diaminomaleonitrile. Cu2+ could be recognized by BDM over other competing metal ions in acetonitrile with distinct fluorescence emission signal response. Upon the addition of Cu2+ to BDM in acetonitrile, the maximum absorption at approximately 530 nm on the longer wavelength side was quenched, and the emission at 530 nm was ignited simultaneously. The fluorescence intensity enhancement could reach a maximum of 204 times the intensity of the BDM blank solution. The fluorescence "off-on" effect is established according to the Cu2+-induced fast intramolecular oxidative cyclization reaction, which could be deduced from the formation of an imidazole ring appended to the cyclization product (2-BODIPY-1H-imidazole-4,5-dicarbonitrile, BMC). Single-crystal structure analysis of the sensor BDM and cyclization product BMC further demonstrated this oxidative cyclization. Finally, the Cu2+ recognition property of BDM was validated in SiHa cells and living zebrafish. Additionally, the blood-brain barrier of the zebrafish can be penetrated by the BDM dye and the neuron cells in the brain were stained.

Development of an infant formula certified reference material for the analysis of organic nutrients.

Infant formula certified reference material (CRM, KRISS CRM 108-02-003) were developed for the analysis of organic nutrients. The CRM is a milk-based infant formula powder, packaged at 14 g per unit. Ten thousand units were prepared and stored at -70 °C. For the certification of each nutrient, ten units were analyzed for simultaneous value-assignment and homogeneity test. Analytical methods used were isotope dilution mass spectrometry (IDMS) based on liquid chromatography mass spectrometer (LC/MS) or gas chromatography mass spectrometer (GC/MS) as higher-order reference methods.13 vitamins, 3 fatty acids, and total cholesterol were certified. The between-unit relative standard deviation of measurement results for each nutrient ranged 0.2% to 2.5%, showing very good homogeneity. The expanded relative uncertainties of the certified values ranged from 1% to 8%, indicating that they have higher-order metrological quality. The values of proximates (proteins, lipids, carbohydrates, water, and ash) were assigned through inter-laboratory comparisons.

Nucleotide Analysis in Korean Dairy Products Using High- Performance Liquid Chromatography with Diode Array Detector.

Nucleotides play important roles in numerous intracellular biochemical processes and are used in infant formulas and other dairy products. However, domestic analytical methods for assessing nucleotide content in products have not yet been established, and therefore, methods for determining nucleotide content are urgently required. A rapid and simple analytical method for determining the content of five types of nucleotides in dairy products was improved using solid phase extraction clean-up and high-performance liquid chromatography with diode array detector. The extraction solvent used in the AOAC method was not well dissolved and was changed to hydrophilic EDTA-Na. In addition, the results obtained using the isocratic elution method and a single wavelength were similar to those obtained using the AOAC method, and the time taken for analysis was shortened from 40 min to 25 min. The process of method validation revealed the following parameters: accuracy (84.69%-102.72%), precision (1.51%-6.82%), linearity (0.999), and limit of detection (cytidine 5'-monophosphate, 0.09 mg/L; uridine 5'-monophosphate, 0.11 mg/L; adenosine 5'-monophosphate, 0.12 mg/L; guanosine 5'-monophosphate, 0.11 mg/L; and inosine 5'-monophosphate, 0.14 mg/L). The method was also used to determine the nucleotide concentration in 25 samples (infant formulas, 1.99-29.39 mg/100 g; and cow milk, 0.28-0.83 mg/100 g). The newly improved method was appropriate for analyzing nucleotides in infant formulas and other dairy products faster when compared to conventional methods.

Prognostic indicators of survival and survival prediction model following extracorporeal cardiopulmonary resuscitation in patients with sudden refractory cardiac arrest.

BACKGROUND: Extracorporeal cardiopulmonary resuscitation (ECPR) has been considered in selected candidates with potentially reversible causes during a limited period. Candidate selection and the identification of predictable conditions are important factors in determining outcomes during CPR in the emergency department (ED). The objective of this study was to determine the key indicators and develop a prediction model for survival to hospital discharge in patients with sudden cardiac arrest who received ECPR. METHODS: This retrospective analysis was based on a prospective cohort, which included data on CPR with ECPR-related variables. Patients with sudden cardiac arrest who received ECPR at the ED from May 2006 to June 2016 were included. The primary outcome was survival to discharge. Prognostic indicators and the prediction model were analyzed using logistic regression. RESULTS: Out of 111 ECPR patients, there were 18.9% survivors. Survivors showed younger age, shorter CPR duration (p < 0.05) and had tendencies of higher rate of initial shockable rhythm (p = 0.055) and higher rate of any ROSC event before ECPR (p = 0.066) than non-survivors. Eighty-one percent of survivors showed favorable neurologic outcome at discharge. In univariate analysis, the following factors were associated with survival: no preexisting comorbidities, initial serum hemoglobin level ≥14 g/dL, and mean arterial pressure ≥60 mmHg after ECPR. Based on multivariate logistic regression, predictors for survival in ECPR were as follows: age ≤56 years, no asystole as the initial arrest rhythm, CPR duration of ≤55 min, and any return of spontaneous circulation (ROSC) event before ECPR. The prediction scoring model for survival had a c-statistic of 0.875. CONCLUSIONS: With careful consideration of differences in the inclusion criteria, the prognostic indicators and prediction scoring model for survival in our study may be helpful in the rapid decision-making process for ECPR implementation during CPR in the ED.

Concealed resuscitation-related injuries as reversible cause of recurrent arrest following extracorporeal cardiopulmonary resuscitation.

A life-threatening cardiopulmonary resuscitation (CPR)-related injury can cause recurrent arrest after return of circulation. Such injuries are difficult to identify during resuscitation, and their contribution to failed resuscitation can be missed given the limitations of conventional CPR. Extracorporeal cardiopulmonary resuscitation (ECPR), increasingly being considered for selected patients with potentially reversible etiology of arrest, may identify previously occult CPR-related injuries by restoring arterial pressure and flow. Herein, we describe two cases of severe CPR-related injuries contributing to recurrent arrest. Each case had ECPR implemented within 60 minutes of the start of CPR. After the presumed cardiac etiology had been addressed with percutaneous coronary intervention, life-threatening cardiovascular injuries with recurrent arrest were noted, and resuscitative thoracotomy was performed under ECPR. One patient survived to hospital discharge. ECPR may provide an opportunity to identify and correct severe resuscitation-related injuries causing recurrent arrest. Chest compression depth >6 cm, especially in older women, may contribute to these injuries.

A system-wide approach from the community to the hospital for improving neurologic outcomes in out-of-hospital cardiac arrest patients.

OBJECTIVE: In the present study, we aimed to determine the effects of a system-wide approach from the community to the hospital in improving the neurologic outcomes in out-of-hospital cardiac arrest (OHCA) patients within Sungbuk in Korea. METHODS: This study used a before-after design. In 2011, compression-only cardiopulmonary resuscitation (CPR) for citizens, a state-wide standard dispatcher assisted-CPR protocol, medical control for regional emergency medical service (EMS), provision of high-quality advanced cardiac life support (ACLS) with capnography and extracorporeal CPR, and the standard postcardiac arrest care protocol were implemented in the system-wide CPR program. CPR provision and outcomes were compared between the 2009-2010 and the 2012-2013 periods. A multivariate logistic regression model for good outcome of OHCA was used to identify interventions with a significant impact. RESULTS: In total, 581 adult nontraumatic OHCA patients who received resuscitation attempts from 2009 to 2013 were selected for the analysis of CPR provision and outcomes. CPR provision improved significantly, as indicated by the following results from 2009-2010 to 2012-2013: from 15.9 to 50.4% for bystander CPR (P<0.001), 6.0 to 0% for the proportion of no documented arrest rhythm by EMS (P=0.004), 41.4 to 62.2% for ACLS with capnography (P=0.008), 1.4 to 10.5% for extracorporeal CPR (P=0.052), 3.7 to 34.4% for successful therapeutic hypothermia in coma patients (P<0.001), and 61.5 to 87.1% for immediate coronary angiography for presumed cardiac etiology (P=0.005). Moreover, the proportion of OHCA patients who received early EMS activation, bystander CPR, appropriate attempt of defibrillation at the prehospital level, high-quality ACLS, and standard postcardiac arrest care increased from 0.5% in 2009-2010 to 8.5% in 2012-2013 (P<0.001). The rates of discharge with a good neurologic outcome improved from 3.3% in 2009-2010 to 8.5% in 2012-2013 (P<0.001). CONCLUSION: The system-wide CPR program was associated with enhancements in CPR performance at both the prehospital and the hospital level, and yielded improved neurologic outcomes in OHCA patients in a small region.

Innate Color Preference of Zebrafish and Its Use in Behavioral Analyses.

Although innate color preference of motile organisms may provide clues to behavioral biases, it has remained a longstanding question. In this study, we investigated innate color preference of zebrafish larvae. A cross maze with different color sleeves around each arm was used for the color preference test (R; red, G; green, B; blue, Y; yellow). The findings showed that 5 dpf zebrafish larvae preferred blue over other colors (B > R > G > Y). To study innate color recognition further, tyrosinase mutants were generated using CRISPR/Cas9 system. As a model for oculocutaneous albinism (OCA) and color vision impairment, tyrosinase mutants demonstrated diminished color sensation, indicated mainly by hypopigmentation of the retinal pigment epithelium (RPE). Due to its relative simplicity and ease, color preference screening using zebrafish larvae is suitable for high-throughput screening applications. This system may potentially be applied to the analysis of drug effects on larval behavior or the detection of sensory deficits in neurological disorder models, such as autism-related disorders, using mutant larvae generated by the CRISPR/Cas9 technique.

Characterization of Total and Size-Fractionated Manganese Exposure by Work Area in a Shipbuilding Yard.

BACKGROUND: Shipbuilding involves intensive welding activities, and welders are exposed to a variety of metal fumes, including manganese, that may be associated with neurological impairments. This study aimed to characterize total and size-fractionated manganese exposure resulting from welding operations in shipbuilding work areas. METHODS: In this study, we characterized manganese-containing particulates with an emphasis on total mass (n = 86, closed-face 37-mm cassette samplers) and particle size-selective mass concentrations (n = 86, 8-stage cascade impactor samplers), particle size distributions, and a comparison of exposure levels determined using personal cassette and impactor samplers. RESULTS: Our results suggest that 67.4% of all samples were above the current American Conference of Governmental Industrial Hygienists manganese threshold limit value of 100 µg/m(3) as inhalable mass. Furthermore, most of the particles containing manganese in the welding process were of the size of respirable particulates, and 90.7% of all samples exceeded the American Conference of Governmental Industrial Hygienists threshold limit value of 20 µg/m(3) for respirable manganese. CONCLUSION: The concentrations measured with the two sampler types (cassette: total mass; impactor: inhalable mass) were significantly correlated (r = 0.964, p < 0.001), but the total concentration obtained using cassette samplers was lower than the inhalable concentration of impactor samplers.

Characterization of Lactobacillus fermentum PL9988 Isolated from Healthy Elderly Korean in a Longevity Village.

In this work, we wanted to develop a probiotic from famous longevity villages in Korea. We visited eight longevity villages in Korea to collect fecal samples from healthy adults who were aged above 80 years and had regular bowel movements, and isolated lactic-acid-producing bacteria from the samples. Isolated colonies that appeared on MRS agar containing bromophenol blue were identified by means of 16S rRNA sequencing, and 102 of the isolates were identified as lactic-acid-producing bacteria (18 species). Lactobacillus fermentum was the most frequently found species. Eight isolates were selected on the basis of their ability to inhibit the growth of six intestinal pathogens (Escherichia coli O157:H7, Salmonella enterica subsp. enterica Typhimurium, Salmonella enterica subsp. enterica Enteritidis, Enterococcus faecalis, Staphylococcus aureus, and Listeria monocytogenes) and their susceptibility to 15 antimicrobial agents. Among these eight isolates, four Lactobacillus fermentum isolates were found not to produce any harmful enzymes or metabolites. Among them, Lactobacillus fermentum isolate no. 24 showed the strongest binding to intestinal epithelial cells, the highest immune-enhancing activity, anti-inflammation activity, and anti-oxidation activity as well as the highest survival rates in the presence of artificial gastric juice and bile solution. This isolate, designated Lactobacillus fermentum PL9988, has all the characteristics for a good probiotic.

Initial Low Oxygen Extraction Ratio Is Related to Severe Organ Dysfunction and High In-Hospital Mortality in Severe Sepsis and Septic Shock Patients.

BACKGROUND: In severe sepsis and septic shock, global tissue hypoxia is a key development preceding multi-organ failure and death. OBJECTIVE: Our aim was to find whether the initial low oxygen extraction ratio (OER) is related to the severity of organ dysfunction and to predict the in-hospital mortality in severe sepsis or septic shock patients. METHODS: This was a secondary analysis of 169 patients with severe sepsis or septic shock in an emergency department. We calculated OER with 1- central venous oxygen saturation (ScvO2)/arterial oxygen saturation and compared the data according to the level of OER (high > 0.3, 0.2 ≤ normal ≤ 0.3, lower < 0.2). RESULTS: A total 133 patients were selected for analysis. OER was inversely proportional to ScvO2 (r(2) = 0.878; p < 0.001). The sepsis-related organ failure assessment score and in-hospital mortality of each group were 6.2 ± 3.7 and 37.0% for high OER, 5.7 ± 3.0 and 11.8% for normal OER, and 7.7 ± 3.9 and 41.7% for low OER, respectively (p = 0.034; p = 0.003). In patients with initial ScvO2 of >70%, in-hospital mortality of patients with low OER was significantly higher than patients with normal OER. CONCLUSIONS: Initial low OER was associated with severe organ dysfunction that resulted in high mortality with severe sepsis and septic shock. When patients had initial ScvO2 of > 70% but abnormally low OER, their in-hospital mortality was higher than in normal OER patients. Therefore, the OER should be considered when attempting to predict the outcome of septic patients using ScvO2 at an early stage of management for sepsis.

Various Enterotoxin and Other Virulence Factor Genes Widespread Among Bacillus cereus and Bacillus thuringiensis Strains.

Many strains of Bacillus cereus cause gastrointestinal diseases, and the closely related insect pathogen Bacillus thuringiensis has also been involved in outbreaks of diarrhea. The diarrheal diseases are attributed to enterotoxins. Sixteen reference strains of B. cereus and nine commercial and 12 reference strains of B. thuringiensis were screened by PCR for the presence of 10 enterotoxigenic genes (hblA, hblC, hblD, nheA, nheB, nheC, cytK, bceT, entFM, and entS), one emetogenic gene (ces), seven hemolytic genes (hlyA, hlyII, hlyIII, plcA, cerA, cerB, and cerO), and a pleiotropic transcriptional activator gene (plcR). These genes encode various enterotoxins and other virulence factors thought to play a role in infections of mammals. Amplicons were successfully generated from the strains of B. cereus and B. thuringiensis for each of these sequences, except the ces gene. Intriguingly, the majority of these B. cereus enterotoxin genes and other virulence factor genes appeared to be widespread among B. thuringiensis strains as well as B. cereus strains.

An optimal transition time to extracorporeal cardiopulmonary resuscitation for predicting good neurological outcome in patients with out-of-hospital cardiac arrest: a propensity-matched study.

INTRODUCTION: Prolonged conventional cardiopulmonary resuscitation (CCPR) is associated with a poor prognosis in out-of-hospital cardiac arrest (OHCA) patients. Alternative methods can be needed to improve the outcome in patients with prolonged CCPR and extracorporeal cardiopulmonary resuscitation (ECPR) can be considered as an alternative method. The objectives of this study were to estimate the optimal duration of CPR to consider ECPR as an alternative resuscitation method in patients with CCPR, and to find the indications for predicting good neurologic outcome in OHCA patients who received ECPR. METHODS: This study is a retrospective analysis based on a prospective cohort. We included patients ≥ 18 years of age without suspected or confirmed trauma and who experienced an OHCA from May 2006 to December 2013. First, we determined the appropriate cut-off duration for CPR based on the discrimination of good and poor neurological outcomes in the patients who received only CCPR, and then we compared the outcome between the CCPR group and ECPR group by using propensity score matching. Second, we compared CPR related data according to the neurologic outcome in matched ECPR group. RESULTS: Of 499 patients suitable for inclusion, 444 and 55 patients were enrolled in the CCPR and ECPR group, respectively. The predicted duration for a favorable neurologic outcome (CPC1, 2) is < 21 minutes of CPR in only CCPR patients. The matched ECPR group with ≥ 21 minutes of CPR duration had a more favorable neurological outcome than the matched CCPR group at 3 months post-arrest. In matched ECPR group, younger age, witnessed arrest without initial asystole rhythm, early achievement of mean arterial pressure ≥ 60 mmHg, low rate of ECPR-related complications, and therapeutic hypothermia were significant factors for expecting good neurologic outcome. CONCLUSIONS: ECPR should be considered as an alternative method for attaining good neurological outcomes in OHCA patients who required prolonged CPR, especially of ≥ 21 minutes. Younger or witnessed arrest patients without initial asystole were good candidates for ECPR. After implantation of ECPR, early hemodynamic stabilization, prevention of ECPR-related complications, and application of therapeutic hypothermia may improve the neurological outcome.

Rapid determination of vitamin D3 in milk-based infant formulas by liquid chromatography-tandem mass spectrometry.

A rapid and simple sample preparation method for vitamin D3 (cholecalciferol) was developed for emulsified dairy products such as milk-based infant formulas. A sample was mixed in a 50 mL centrifuge tube with the same amount of water and isopropyl alcohol to achieve chemical extraction. Ammonium sulfate was used to induce phase separation. No-heating saponification was performed in the sample tube by adding KOH, NaCl, and NH3. Vitamin D3 was then separated and quantified using liquid chromatography-tandem mass spectrometry. The results for added recovery tests were in the range 93.11-110.65%, with relative standard deviations between 2.66% and 2.93%. The results, compared to those obtained using a certified reference material (SRM 1849a), were within the range of the certificated values. This method could be implemented in many laboratories that require time and labour saving.

Aneurysms of distal posterior inferior cerebellar artery.

OBJECTIVE: Aneurysms are very rarely encountered in the distal posterior inferior cerebellar artery (PICA). The authors experienced 5 cases with a distal PICA aneurysm among 368 cases of intracranial aneurysms during the period from January 2003 to January 2008. Here, the authors describe their clinical and surgical experiences and include a review of the relevant literature. METHODS: Using radiologic findings and charts, we retrospectively reviewed the surgical results of 5 cases with a distal PICA aneurysm treated from January 2003 to January 2008. RESULTS: The current five cases were composed of four cases of 'Good' and one case of 'Fair'. No postoperative complications occurred other than a ventriculo-peritoneal shunt due to hydrocephalus in Case 2. In all five cases, treatment was successful without neurological deficit. CONCLUSION: Surgical outcome of PICA aneurysms have been reported to be excellent because the amount of intraparenchymal injury is limited. More clinical experience, microsurgical technique developments, and endovascular surgery advancements are certain to improve treatment outcomes.