The effect of calcium and cyclic AMP on amylase release in digitonin-permeabilized parotid gland cells

Rat parotid cells were permeabilized with digitonin to examine their secretory dynamics. Cells were isolated by a modification of the method previously described by Hootman [1985). J. Biol. Chem. 260, 4186-4194) in which alpha-chymotrypsin was included. The final preparation consisted of approx. 40-60% single cells. The cells were 85-90% viable by trypan blue exclusion and secreted amylase when stimulated with isoproterenol. Digitonin (2 or 5 microM) was sufficient for permeabilization while 2 microM digitonin was somewhat more effective in maintaining cell integrity as indicated by lactate dehydrogenase release. Digitonin had minimal effects on intracellular granules in the whole cell and was, thus, relatively selective. The response of digitonin-permeabilized cells to calcium (without secretagogues) in the incubation medium was monitored by amylase release. For a wide range of applied free calcium concentrations (1 X 10(-7) M to 10(-4) M) a statistically significant increase in amylase secretion was observed. Control cells did not release amylase to a similar extent without secretagogue. Cyclic AMP (50 microM) significantly enhanced amylase secretion from digitonin-treated cells at all concentrations of free calcium tested. Neither calcium nor cyclic AMP alone was sufficient to stimulate maximal amylase release. Our results provide direct evidence for a model in which calcium and cyclic AMP work on separate pathways as interacting regulators of exocytosis.     

Insulin Resistance,Hyperleptinemia, and Obstructive Sleep Apnea in Launois‐Bensaude Syndrome

Objective: Launois‐Bensaude Syndrome (LBS) is a very rare cause of obesity, characterized by a symmetrical accumulation of a very large number of lipomata in different regions of the body, excluding the face, the forearms, and the shanks. Obesity is known to be closely associated with insulin resistance, hyperleptinemia, and obstructive sleep apnea (OSA). We were interested in studying whether these conditions are also present in patients with obesity due to LBS with a similar frequency as in patients with “simple” truncal obesity. Research Methods and Procedures: We performed polysomnography and hyperinsulinemic euglycemic clamp studies and measured serum leptin in three patients with LBS and in six patients with “simple” truncal obesity, matched for sex and body mass index (LBS group, 36.39 kg/m²; controls, 35.82 kg/m²). Results: Polysomnography revealed severe OSA in one LBS patient with marked “horsecollar lipomata.” In the other LBS patients, no OSA could be demonstrated. The leptin levels of the two groups were comparable (LBS group, 36.39 μg/liter; controls, 37.18 μg/liter) and the insulin responsiveness index was also comparable in the two groups (LBS group, 3.47 μmol/kg · minute; controls, 3.79 μmol/kg · minute). Discussion: Patients with LBS demonstrated similar metabolic features in terms of insulin sensitivity and hyperleptinemia as patients with “simple” truncal obesity. LBS is not strictly associated with OSA.     

Metabolic profiling of a potential antifungal drug, 3-(4-bromophenyl)-5-acetoxymethyl-2,5-dihydrofuran-2-one, in mouse urine using high-performance liquid chromatography with UV photodiode-array and mass spectrometric detection

3-(4-bromophenyl)-5-acetyloxymethyl-2,5-dihydrofuran-2-one (LNO-18-22) is a representative member of a novel group of potential antifungal drugs, derived from a natural 3,5-disubstituted butenolide, (-)incrustoporine, as a lead structure. This lipophilic compound is characterized by high in vitro antifungal activity and low acute toxicity. For the purpose of in vivo studies, a new bioanalytical high-performance liquid chromatographic method with UV photodiode-array and mass spectrometric detection (HPLC-PDA-MS), involving a direct injection of diluted mouse urine was developed and used in the evaluation of the metabolic profiling of this drug candidate. The separation of LNO-18-22 and its phase I metabolites was performed in 37 min on a 125 mmx4 mm chromatographic column with Purospher RP-18e using an acetonitrile-water gradient elution. Scan mode of UV detection (195-380 nm) was employed for the identification of the parent compound and its biotransformation products in the biomatrix. Finally, the identity of LNO-18-22 and its metabolites was confirmed using HPLC-MS analyses of the eluate. These experiments demonstrated the power of a comprehensive analytical approach based on the combination of xenobiochemical methods and the results from tandem HPLC-PDA-MS (chromatographic behaviour, UV and MS spectra of native metabolites versus synthetic standards). The chemical structures of five phase I LNO-18-22 metabolites and one phase II metabolite were elucidated in the mouse urine, with two of these metabolites having very unexpected structures.     

Identification of the Pain Process by Cold Stimulation: Using Dynamic Causal Modeling of Effective Connectivity in Functional Near-Infrared Spectroscopy (fNIRS)

Background

Pain is an unpleasant sensory and emotional experience followed by anxiety, depression, and frustration. Functional Near-Infrared Spectroscopy (fNIRS) as an optical technique identifies the brain functional networks by investigating connectivity between functionally linked of different anatomical regions in response to pain stimulation.

Repetitive somatic embryogenesis in carnation on picloram supplemented media

An efficient method of repetitive somatic embryogenesis and plant regeneration of two carnation cultivars (Sagres and Impulse) was established using a two-step protocol. In the first step, embryogenic tissue were induced from petal explants on MS culture medium containing 9% sucrose (w/v), 9 μM 2,4-D and 0.8 μM BA. In the second step, embryogenic tissue transferred onto the MS medium containing 3% sucrose supplemented with different concentrations of picloram (0.8, 2, 4, 8 and 16 μM) to produce primary somatic embryos. Precotyledonary clumps and cotyledonary somatic embryos were then isolated and subcultured onto the same media as the second step where they formed secondary somatic embryos in repetitive cycles. Cotyledony somatic embryos were converted into plantlets when they transferred onto the growth regulator-free half-strength MS medium followed by being acclimated to the greenhouse conditions.     

Protection against discrimination in national dementia guideline recommendations: A systematic review

BackgroundNational dementia guidelines provide recommendations about the most effective approaches to diagnosis and interventions. Guidelines can improve care, but some groups such as people with minority characteristics may be disadvantaged if recommended approaches are the same for everyone. It is not known if dementia guidelines address specific needs related to patient characteristics. The objectives of this review are to identify which countries have national guidelines for dementia and synthesise recommendations relating to protected characteristics, as defined in the UK Equality Act 2010: age, disability, gender identity, marriage and civil partnership, pregnancy and maternity, race, religion or belief, sex, and sexual orientation.Methods and findingsWe searched CINAHL, PsycINFO, and Medline databases and the Guideline International Network library from inception to March 4, 2020, for dementia guidelines in any language. We also searched, between April and September 2020, Google and the national health websites of all 196 countries in English and in each country’s official languages. To be included, guidelines had to provide recommendations about dementia, which were expected to be followed by healthcare workers and be approved at a national policy level. We rated quality according to the iCAHE guideline quality checklist. We provide a narrative synthesis of recommendations identified for each protected characteristic, prioritising those from higher-quality guidelines. Forty-six guidelines from 44 countries met our criteria, of which 18 were rated as higher quality. Most guidelines (39/46; 85%) made at least one reference to protected characteristics, and we identified recommendations relating to age, disability, race (or culture, ethnicity, or language), religion, sex, and sexual orientation. Age was the most frequently referenced characteristic (31/46; 67%) followed by race (or culture, ethnicity, or language; 25/46; 54%). Recommendations included specialist investigation and support for younger people affected by dementia and consideration of culture when assessing whether someone had dementia and providing person-centred care. Guidelines recommended considering religion when providing person-centred and end-of-life care. For disability, it was recommended that healthcare workers consider intellectual disability and sensory impairment when assessing for dementia. Most recommendations related to sex recommended not using sex hormones to treat cognitive impairment in men and women. One guideline made one recommendation related to sexual orientation. The main limitation of this study is that we only included national guidelines applicable to a whole country meaning guidelines from countries with differing healthcare systems within the country may have been excluded.ConclusionsNational guidelines for dementia vary in their consideration of protected characteristics. We found that around a fifth of the world’s countries have guidelines for dementia. We have identified areas of good practice that can be considered for future guidelines and suggest that all guidelines provide specific evidence-based recommendations for minority groups with examples of how to implement them. This will promote equity in the care of people affected by dementia and help to ensure that people with protected characteristics also have high-quality clinical services.

Tiffeny James and team investigate which countries have national guidelines for dementia, and synthesise recommendations relating to protected characteristics as defined in the UK Equality Act 2010.     

Auto-CPAP therapy based on the forced oscillation technique.

Autoadjusting CPAP devices (APAP) are designed to continuously adjust the positive pressure to the required levels, and thus increase treatment quality and patient compliance. The results of APAP treatment strongly depend on the control mechanism of the respective APAP device. In agreement with other working groups, we have recently shown that the forced oscillation technique (FOT) is capable of detecting incipient upper airway obstruction prior to physiological reactions such as the onset of increasing esophageal pressure swings or microarousals. Therefore we studied efficacy and acceptance of a novel APAP device controlled exclusively by FOT. 100 consecutive patients with OSAS confirmed by polysomnography (mean AHI 47.9 +/- 22.6) and daytime sleepiness (Epworth sleepiness scale, ESS 12.6 +/- 3.9) were randomized to either APAP treatment (n = 50) or conventional CPAP treatment (n = 50). Polysomnographies were performed at the second treatment night and subjective sleepiness (modified ESS) was established in the morning. The respiratory disturbance was largely normalized in both treatment groups in the second treatment night (AHI 4.7 +/- 5.3 vs. 3.7 +/- 3.4; n.s.). Both groups showed largely improved sleep profiles and had markedly reduced ESS-scores (6.6 +/- 3.6 vs. 7.0 +/- 3.4; n.s.). The mean treatment pressure during APAP was significantly lower than during CPAP treatment (6.0 +/- 2.0 vs. 9.0 +/- 1.8 mbar; p < 0.001). There were no significant differences between APAP and CPAP treatment in any parameter of efficacy or acceptance. APAP treatment with this device controlled exclusively by FOT is well accepted by the patients and permits an adequate treatment of OSAS without the need for invidiual CPAP titration.     

Amperometric sulfide detection using Coprinus cinereus peroxidase immobilized on screen printed electrode in an enzyme inhibition based biosensor

In the present work, an amperometric inhibition biosensor for the determination of sulfide has been fabricated by immobilizing Coprinus cinereus peroxidase (CIP) on the surface of screen printed electrode (SPE). Chitosan/acrylamide was applied for immobilization of peroxidase on the working electrode. The amperometric measurement was performed at an applied potential of -150 mV versus Ag/AgCl with a scan rate of 100 mV in the presence of hydroquinone as electron mediator and 0.1M phosphate buffer solution of pH 6.5. The variables influencing the performance of sensor including the amount of substrate, mediator concentration and electrolyte pH were optimized. The determination of sulfide can be achieved in a linear range of 1.09-16.3 μM with a detection limit of 0.3 μM. Developed sensor showed quicker response to sulfide compared to the previous developed sulfide biosensors. Common anions and cations in environmental water did not interfere with sulfide detection by the developed biosensor. Cyanide interference on the enzyme inhibition caused 43.25% error in the calibration assay which is less than the amounts reported by previous studies. Because of high sensitivity and the low-cost of SPE, this inhibition biosensor can be successfully used for analysis of environmental water samples.