Corneal Collagen Cross-Linking for the Management of Mycotic Keratitis

Purpose

To evaluate the efficiency of corneal collagen cross-linking (CXL) in addition to topical voriconazole in cases with mycotic keratitis.

Design

Retrospective case series in a tertiary university hospital.

Participants

CXL was performed on 13 patients with mycotic keratitis who presented poor or no response to topical voriconazole treatment.

Methods

The clinical features, symptoms, treatment results and complications were recorded retrospectively. The corneal infection was graded according to the depth of infection into the stroma (from grade 1 to grade 3). The visual analogue scale was used to calculate the pain score before and 2 days after surgery.

Main Outcome Measures

Grade of the corneal infection.

Results

Mean age of 13 patients (6 female and 7 male) was 42.4 ± 17.7 years (20–74 years). Fungus was demonstrated in culture (eight patients) or cytological examination (five patients). Seven of the 13 patients (54%) were healed with topical voriconazole and CXL adjuvant treatment in 26 ± 10 days (15–40 days). The remaining six patients did not respond to CXL treatment; they initially presented with higher grade ulcers. Pre- and post-operative pain score values were 8 ± 0.8 and 3.5 ± 1, respectively (p < 0.05).

Conclusions

The current study suggests that adjunctive CXL treatment is effective in patients with small and superficial mycotic ulcers. These observations require further research by large randomized clinical trials.

     

Optimization of fecal sample preparation for untargeted LC-HRMS based metabolomics

Introduction

Collecting feces is easy. It offers direct outcome to endogenous and microbial metabolites.

Objectives

In a context of lack of consensus about fecal sample preparation, especially in animal species, we developed a robust protocol allowing untargeted LC-HRMS fingerprinting.

Methods

The conditions of extraction (quantity, preparation, solvents, dilutions) were investigated in bovine feces.

Results

A rapid and simple protocol involving feces extraction with methanol (1/3, M/V) followed by centrifugation and a step filtration (10 kDa) was developed.

Conclusion

The workflow generated repeatable and informative fingerprints for robust metabolome characterization.

     

Does centrifugation matter? Centrifugal force and spinning time alter the plasma metabolome

Background

Centrifugation is an indispensable procedure for plasma sample preparation, but applied conditions can vary between labs.

Aim

Determine whether routinely used plasma centrifugation protocols (1500×g 10 min; 3000×g 5 min) influence non-targeted metabolomic analyses.

Methods

Nuclear magnetic resonance spectroscopy (NMR) and High Resolution Mass Spectrometry (HRMS) data were evaluated with sparse partial least squares discriminant analyses and compared with cell count measurements.

Results

Besides significant differences in platelet count, we identified substantial alterations in NMR and HRMS data related to the different centrifugation protocols.

Conclusion

Already minor differences in plasma centrifugation can significantly influence metabolomic patterns and potentially bias metabolomics studies.

     

Minimal variation of the plasma lipidome after delayed processing of neonatal cord blood

Background

Cord blood lipids are potential disease biomarkers. We aimed to determine if their concentrations were affected by delayed blood processing.

Method

Refrigerated cord blood from six healthy newborns was centrifuged every 12 h for 4 days. Plasma lipids were analysed by liquid chromatography/mass spectroscopy.

Results

Of 262 lipids identified, only eight varied significantly over time. These comprised three dihexosylceramides, two phosphatidylserines and two phosphatidylethanolamines whose relative concentrations increased and one sphingomyelin that decreased.

Conclusion

Delay in separation of plasma from refrigerated cord blood has minimal effect overall on the plasma lipidome.

     

Reengineering substrate specificity of <Emphasis Type="Italic">E. coli</Emphasis> glutamate dehydrogenase using a position-based prediction method

Objective

To re-engineer the active site of proteins for non-natural substrates using a position-based prediction method (PBPM).

Results

The approach has been applied to re-engineer the E. coli glutamate dehydrogenase to alter its substrate from glutamate to homoserine for a de novo 1,3-propanediol biosynthetic pathway. After identification of key residues that determine the substrate specificity, residue K92 was selected as a candidate site for mutation. Among the three mutations (K92V, K92C, and K92M) suggested by PBPM, the specific activity of the best mutant (K92 V) was increased from 171 ± 35 to 1328 ± 71 μU mg^?1.

Conclusion

The PBPM approach has a high efficiency for re-engineering the substrate specificity of natural enzymes for new substrates.

     

R-wave synchronised atrial pacing in pediatric patients with postoperative junctional ectopic tachycardia: the atrioventricular interval investigated by computational analysis and clinical evaluation

Background

R-wave synchronised atrial pacing is an effective temporary pacing therapy in infants with postoperative junctional ectopic tachycardia. In the technique currently used, adverse short or long intervals between atrial pacing and ventricular sensing (AP–VS) may be observed during routine clinical practice.

Objectives

The aim of the study was to analyse outcomes of R-wave synchronised atrial pacing and the relationship between maximum tracking rates and AP–VS intervals.

Methods

Calculated AP–VS intervals were compared with those predicted by experienced pediatric cardiologist.

Results

A maximum tracking rate (MTR) set 10 bpm higher than the heart rate (HR) may result in undesirable short AP–VS intervals (minimum 83 ms). A MTR set 20 bpm above the HR is the hemodynamically better choice (minimum 96 ms). Effects of either setting on the AP–VS interval could not be predicted by experienced observers. In our newly proposed technique the AP–VS interval approaches 95 ms for HR > 210 bpm and 130 ms for HR < 130 bpm. The progression is linear and decreases strictly (? 0.4 ms/bpm) between the two extreme levels.

Conclusions

Adjusting the AP–VS interval in the currently used technique is complex and may imply unfavorable pacemaker settings. A new pacemaker design is advisable to allow direct control of the AP–VS interval.

     

Endoglucanase enzymatic modification of kraft pulp during recycling

Objective

To investigate the cellulose modification process on kraft pulp during recycling by mono-endoglucanase.

Results

Pichia pastoris expressing endoglucanase, EG1, was grown in a 10 l fermenter yielding a high carboxymethyl cellulase (CMCase) activity of 340 U mg^?1. EG1-mediated modification of kraft pulp resulted in a paper sheet with the tensile index and burst index increased by 10 and 6.5 %, respectively. The kink index (indicating abrupt bends in fibres) of the enzyme-treated group decreased sharply by 45 % after the first recycling, compared with a reduction of only 1 % in the control group. Furthermore, EG1 treatment decreased the growth of crystallinity from 73.5 to 73.2 % and crystal size from 7.45 to 7.21 nm, which alleviated paper aging.

Conclusion

Endoglucanase EG1 modifies the interfacial properties of fibers, which affects fibre morphology during the recycling process and improves the technical properties of the resulting pulp and paper.

     

Effect of food intake on myocardial performance index

Background

Myocardial performance index (MPI) has been investigated in a variety of populations, but the effect of food intake has not been evaluated. We assessed whether myocardial performance index is affected by food intake in healthy subjects.

Methods

Twenty-three healthy subjects aged 25.6?±?4.5 years were investigated. MPI was measured before, 30 min after, and 110 min after a standardized meal.

Results

MPI decreased significantly (P?<?0.05) from fasting values 30 min after the meal, and had almost returned to baseline after 110 min. MPI decreased from 0.28?±?0.06 (fasting) to 0.20?±?0.07 30 min after eating. At 110 min after eating the index value was almost back to the baseline value 0.26?±?0.06. (P?=?0.15).

Conclusions

This study shows that myocardial performance index is affected by food intake in healthy subjects.

     

Maternal-fetal hepatic and placental metabolome profiles are associated with reduced fetal growth in a rat model of maternal obesity

Introduction

Maternal obesity is associated with a range of pregnancy complications, including fetal growth restriction (FGR), whereby a fetus fails to reach its genetically determined growth. Placental insufficiency and reduced nutrient transport play a role in the onset of FGR.

Objectives

Metabolomic profiling was used to reveal altered maternal and fetal metabolic pathways in a model of diet induced obesity during pregnancy, leading to reduced fetal growth.

Methods

We examined the metabolome of maternal and fetal livers, and placenta following a high fat and salt intake. Sprague–Dawley rats were assigned to (a) control diet (CD; 1 % salt, 10 % kcal from fat), (b) high salt diet (SD; 4 % salt, 10 % kcal from fat), (c) high fat diet (HF; 1 % salt, 45 % kcal from fat) or (d) high-fat high-salt diet (HFSD; 4 % salt, 45 % kcal from fat) 21 days prior to pregnancy and during gestation. Metabolites from maternal and fetal livers, and placenta were identified using gas and liquid chromatography combined with mass spectrometry.

Results

Maternal HF intake resulted in reduced fetal weight. Altered metabolite profiles were observed in the HF maternal and fetal liver, and placenta. Polyunsaturated fatty acid metabolism was significantly altered in maternal and fetal liver by maternal fat intake.

Conclusion

Excess of linoleic and α-linoleic acid (essential fatty acids) may be detrimental during placentation and associated with a reduction in fetal weight. Additionally, maternal, placental and fetal response to increased fat consumption seems likely to involve palmitoleic acid utilization as an adaptive response during maternal obesity.

     

Application of residual polysaccharide-degrading enzymes in dried shiitake mushrooms as an enzyme preparation in food processing

Objective

To examine the activities of residual enzymes in dried shiitake mushrooms, which are a traditional foodstuff in Japanese cuisine, for possible applications in food processing.

Results

Polysaccharide-degrading enzymes remained intact in dried shiitake mushrooms and the activities of amylase, β-glucosidase and pectinase were high. A potato digestion was tested using dried shiitake powder. The enzymes reacted with potato tuber specimens to solubilize sugars even under a heterogeneous solid-state condition and that their reaction modes were different at 38 and 50 °C.

Conclusion

Dried shiitake mushrooms have a potential use in food processing as an enzyme preparation.

     

High resolution metabolomic analysis of ASD human brain uncovers novel biomarkers of disease

Introduction

Autism spectrum disorders (ASD) is a group of neurodevelopmental disorders believed to have a multifactorial basis. Presently, diagnosis is based on behavioral and developmental signs in children before the age of 3 and no reliable clinical biomarkers are available for early detection.

Objectives

This study aimed to biochemically profile the cerebellum from post-mortem human brain from ASD sufferers (n = 11) and compare their profiles to that of age-matched controls (n = 11) with no known brain disorder.

Methods

Using liquid chromatography combined with LTQ-Orbitrap mass spectrometry we detected 14,328 features in ESI+ mode in polar extracts of post-mortem brain.

Results

Of these only 37 were found to be statistically significantly different between ASD and controls (p < 0.05; fdr < 0.05). A panel of four features had a predictive power of 96.64 %, following statistical cross validation, for ASD detection. This model produced an AUC = 0.874 (CI 0.768–0.944) and a Fisher’s exact score of p = 4.50E?29.

Conclusion

Whilst at this time we were unable to chemically identify the four features of interest we believe that this study underscores the potential value of high resolution metabolomics for the study of ASD. Further characterization of the polar metabolome of post mortem ASD brains could lead to the identification of potential biomarkers and novel therapeutics for the disease. The development of accurate biomarkers could assist in the early detection of ASD and promote early intervention strategies to improve outcome.

     

Medical students and physical education students as CPR instructors: an appropriate solution to the CPR-instructor shortage in secondary schools?

Background

Integrating cardiopulmonary resuscitation (CPR) training in secondary schools will increase the number of potential CPR providers. However, currently too few certified instructors are available for this purpose. Training medical students and physical education student teachers to become CPR instructors could decrease this shortage.

Aim

Examine whether medical students and physical education student teachers can provide CPR training for secondary school pupils as well as (i.?e., non-inferior to) registered nurses.

Methods

A total of 144 secondary school pupils were randomly assigned to CPR training by a registered nurse (n = 12), a  medical student (n = 17) or a physical education student teacher (n = 15). CPR performance was assessed after training and after eight weeks in a simulated cardiac arrest scenario on a resuscitation manikin, using manikin software and video recordings.

Results

No significant differences were found between the groups on the overall Cardiff Test scores and the correctness of the CPR techniques during the post-training and retention test. All pupils showed sufficient CPR competence, even after eight weeks.

Conclusion

Training by medical students or physical education student teachers is non-inferior to training by a registered nurse, suggesting that school teachers, student teachers and medical students can be recruited for CPR training in secondary schools.

     

Pranlukast,a novel binding ligand of human Raf1 kinase inhibitory protein

Objectives

To study the binding of pranlukast to hRKIP and its regulatory role in the Raf1/MEK/ERK signal pathway.

Results

NMR and fluorescence experiments demonstrated hRKIP could bind pranlukast with a binding constant of 1016 mM^?1. Residues (Y81, S109 and Y181) on the conserved ligand-binding pocket of hRKIP played a crucial role in binding pranlukast, and their mutations reduced the binding affinity more than 85 %. Furthermore, 25 μM pranlukast could up-regulate the ERK phosphorylation by about 17 %.

Conclusion

Pranlukast may be used as a potential drug precursor for treating hRKIP involved diseases.

     

Novel microbial photobioelectrochemical cell using an invasive ultramicroelectrode array and a microfluidic chamber

Objective

To fabricate a novel microbial photobioelectrochemical cell using silicon microfabrication techniques.

Results

High-density photosynthetic cells were immobilized in a microfluidic chamber, and ultra-microelectrodes in a microtip array were inserted into the cytosolic space of the cells to directly harvest photosynthetic electrons. In this way, the microbial photobioelectrochemical cell operated without the aid of electron mediators. Both short circuit current and open circuit voltage of the microbial photobioelectrochemical cell responded to light stimuli, and recorded as high as 250 pA and 45 mV, respectively.

Conclusion

A microbial photobioelectrochemical cell was fabricated with potential use in next-generation photosynthesis-based solar cells and sensors.

     

Effecten van geriatrische COPD-revalidatie op ziekenhuisopnames en inspanningstolerantie: een retrospectief observationele studie

Introduction

Frail COPD patients are frequently not accepted for regular pulmonary rehabilitation programs due to low physical condition and functional limitations. Rehabilitation programs in nursing homes for geriatric patients with COPD have been developed. The effects of such programs are largely unknown.

Aims

To assess the course of COPD-related hospital admissions and exercise tolerance in a cohort of frail COPD patients participating in geriatric COPD rehabilitation.

Methods

Retrospective observational study with a follow up of 12 months after discharge from rehabilitation. COPD related hospital admission days were measured in the year before and after participating rehabilitation. Exercise tolerance was measured by the six minute walk test (6MWT) at admission and at discharge from rehabilitation.

Results

Fifty-eight participants accomplished the rehabilitation program. Twelve patients died in the first year after discharge. The median number of hospital admission days in the year before participating rehabilitation was 21 (IQR 10–33). The first year after discharge this was decreased to a median of 6 (IQR 0–12). The 6MWT increased from 194 (SD 85) meters at admission to 274 (SD 95) meters at discharge (mean difference 80 m, SD 72; p < 0.05).

Conclusions

Geriatric COPD rehabilitation in a nursing home setting seems to reduce hospital admissions in frail COPD patients and to increase exercise tolerance.

     

High-yield production of pure tagatose from fructose by a three-step enzymatic cascade reaction

Objective

To produce tagatose from fructose with a high conversion rate and to establish a high-yield purification method of tagatose from the reaction mixture.

Results

Fructose at 1 M (180 g l^?1) was converted to 0.8 M (144 g l^?1) tagatose by a three-step enzymatic cascade reaction, involving hexokinase, plus ATP, fructose-1,6-biphosphate aldolase, phytase, over 16 h with a productivity of 9 g l^?1 h^?1. No byproducts were detected. Tagatose was recrystallized from ethanol to a purity of 99.9% and a yield of 96.3%. Overall, tagatose at 99.9% purity was obtained from fructose with a yield of 77%.

Conclusion

This is the first biotechnological production of tagatose from fructose and the first application of solvent recrystallization for the purification of rare sugars.

     

Transient expression of recombinant tissue plasminogen activator (rt-PA) gene in cucurbit plants using viral vector

Objective

To use a transient expression system to express a truncated human tissue plasminogen activator (K2S) gene in cucurbit plants.

Results

The recombinant tissue plasminogen activator protein (K2S form) was expressed in active form in cucurbit plants. Its molecular weight was 43 kDa. The plant-derived rt-PA was determined using goat anti-rabbit antibody by western blotting. Among the infected lines, the highest expression of rt-PA was 62 ng/100 mg per leaf tissue as measured by ELISA. The enzymatic activity of the plant-derived rt-PA was 0.8 IU/ml.

Conclusions

The K25 form of rt-PA was expressed for the first time using the viral expression system. Plant-derived rt-PA showed similar potency to commercially-available PA.

     

Dexamethasone downregulates SIRT1 and IL6 and upregulates EDN1 genes in stem cells derived from gingivae via the AGE/RAGE pathway

Objectives

To evaluate the effects of dexamethasone on the aging of mesenchymal stem cells from human gingiva using next-generation sequencing.

Results

Four mRNAs were upregulated and 12 were downregulated when the results of dexamethasone at 24 h were compared with the control at 24 h. Expressions of SIRT1 and IL6 were decreased in dexamethasone at 24 h but expression of EDN1 was increased.

Conclusions

Application of dexamethasone reduced the expression of SIRT1 and IL6 but enhanced the expression of EDN1 of stem cells.

     

Impact of transduction towards the proliferation and migration as well as the transduction efficiency of human umbilical cord-derived late endothelial progenitor cells with nine recombinant adeno-associated virus serotypes

Objectives

To evaluate the transduction efficiency of human umbilical cord-derived, late endothelial progenitor cells late (HUCB-late EPCs) with nine recombinant adeno-associated virus (rAAV) serotypes and the ability of proliferation and migration of the cells after transduction.

Results

rAAV2 and rAAV6 showed a greater ability than other serotypes to transduce late EPCs (P < 0.05). After transduction, cell proliferation ability weakened (P < 0.05), but the ability of migration to stromal cell-derived factor (SDF-1) unchanged.

Conclusion

There is an advantage of choosing the optimal rAAV serotype as a gene vector to alter the biologic characteristics of late EPCs.

     

Stability of targeted metabolite profiles of urine samples under different storage conditions

Introduction

Few studies have investigated the influence of storage conditions on urine samples and none of them used targeted mass spectrometry (MS).

Objectives

We investigated the stability of metabolite profiles in urine samples under different storage conditions using targeted metabolomics.

Methods

Pooled, fasting urine samples were collected and stored at ?80 °C (biobank standard), ?20 °C (freezer), 4 °C (fridge), ~9 °C (cool pack), and ~20 °C (room temperature) for 0, 2, 8 and 24 h. Metabolite concentrations were quantified with MS using the AbsoluteIDQ? p150 assay. We used the Welch-Satterthwaite-test to compare the concentrations of each metabolite. Mixed effects linear regression was used to assess the influence of the interaction of storage time and temperature.

Results

The concentrations of 63 investigated metabolites were stable at ?20 and 4 °C for up to 24 h when compared to samples immediately stored at ?80 °C. When stored at ~9 °C for 24 h, few amino acids (Arg, Val and Leu/Ile) significantly decreased by 40% in concentration (P < 7.9E?04); for an additional three metabolites (Ser, Met, Hexose H1) when stored at ~20 °C reduced up to 60% in concentrations. The concentrations of four more metabolites (Glu, Phe, Pro, and Thr) were found to be significantly influenced when considering the interaction between exposure time and temperature.

Conclusion

Our findings indicate that 78% of quantified metabolites were stable for all examined storage conditions. Particularly, some amino acid concentrations were sensitive to changes after prolonged storage at room temperature. Shipping or storing urine samples on cool packs or at room temperature for more than 8 h and multiple numbers of freeze and thaw cycles should be avoided.