Palliative care in the emergency department as seen by providers and users: a qualitative study

Background Much effort has been made to explore how patients with advanced chronic illness and their families experience care when they attend the Emergency Department, and many studies have investigated how healthcare professionals perceive Palliative Care provision in the Emergency Department. Various models exist, but nonetheless incorporating palliative care into the Emergency Department remains challenging. Considering both healthcare professionals\u2019 and users\u2019 perspective on problems encountered in delivering and receiving appropriate palliative care within this context may provide important insight into meaningful targets for improvements in quality of care. Accordingly, this study aims at exploring issues in delivering palliative care in the Emergency Department from the perspective of both providers and users, as part of a larger project on the development and implementation of a quality improvement program in Italian Emergency Departments. Methods A qualitative study involving focus group interviews with Emergency Department professionals and semi-structured interviews with patients with palliative care needs in the Emergency Department and their relatives was conducted. Both datasets were analyzed using Thematic Analysis. Results Twenty-one healthcare professionals, 6 patients and 5 relatives participated in this study. Five themes were identified: 1) shared priorities in Emergency Department among healthcare professionals and patients, 2) the information provided by healthcare professionals and that desired by relatives, 3) perception of environment and time, 4) limitations and barriers to the continuity of care, and 5) the contrasting interpretations of giving and receiving palliative care. Conclusions This study provides insights into targets for changes in Italian Emergency Departments. Room for improvement relates to training for healthcare professionals on palliative care, the development of a shared care pathway for patients with palliative care needs, and the optimization of Emergency Department environment. These targets will be the basis for the development of a quality improvement program in Italian Emergency Departments

The formation of persister cells in stationary-phase cultures of Escherichia coli is associated with the aggregation of endogenous proteins.

Persister cells (persisters) are transiently tolerant to antibiotics and usually constitute a small part of bacterial populations. Persisters remain dormant but are able to re-grow after antibiotic treatment. In this study we found that the frequency of persisters correlated to the level of protein aggregates accumulated in E. coli stationary-phase cultures. When 3-(N-morpholino) propanesulfonic acid or an osmolyte (trehalose, betaine, glycerol or glucose) were added to the growth medium at low concentrations, proteins were prevented from aggregation and persister formation was inhibited. On the other hand, acetate or high concentrations of osmolytes enhanced protein aggregation and the generation of persisters. We demonstrated that in the E. coli stationary-phase cultures supplemented with MOPS or a selected osmolyte, the level of protein aggregates and persister frequency were not correlated with such physiological parameters as the extent of protein oxidation, culturability, ATP level or membrane integrity. The results described here may help to understand the mechanisms underlying persister formation

Genes encoding equine β-lactoglobulin (LGB1 and LGB2): Polymorphism, expression, and impact on milk composition.

β-lactoglobulin is one of the most abundant milk whey proteins in many mammal species, including the domestic horse. The aim of this study was to screen for polymorphism in the equine LGB1 and LGB2 gene sequences (all exons, introns, and 5'-flanking region) and to assess potential relationship of particular genotypes with gene expression levels (measured in milk somatic cells) and milk composition traits (protein, fat, lactose, and total β-lactoglobulin content). Direct DNA sequencing analysis was performed for twelve horse breeds: Polish Primitive Horse (PPH), Polish Coldblood Horse (PCH), Polish Warmblood Horse (PWH), Silesian, Hucul, Fjording, Haflinger, Shetland Pony, Welsh Pony, Arabian, Thoroughbred, and Percheron-and revealed the presence of 83 polymorphic sites (47 and 36 for LGB1 and LGB2 genes, respectively), including eight that were previously unknown. Association analysis of the selected polymorphisms, gene expression, and milk composition traits (conducted for the PPH, PCH, and PWH breeds) showed several statistically significant relationships; for example, the two linked LGB1 SNPs (rs1143515669 and rs1144647991) were associated with total milk protein content (p < 0.01). Our study also confirmed that horse breed had significant impact on both gene transcript levels (p < 0.01) and on milk LGB content (p < 0.05), whereas an influence of lactation period was seen only for gene relative mRNA abundances (p < 0.01)

The influence of acetate, MOPS and osmolytes on the levels of non-dividing and persister cells.

<p>Bacteria were grown aerobically for 24 h at 37°C in LB medium supplemented with 0.2% acetate, 40 mM MOPS pH 7.4, 0.2% glucose, 0.2% glycerol, 0.2% trehalose or 0.2% betaine. The control culture was grown in LB without supplements. (A) The total number of cells per ml was obtained using a hemocytometer. Dividing cell counts were estimated by plating serial dilutions on LA. The number of non-dividing cells was calculated by subtracting colony-forming units counts from the total number of cells. (B) <i>E. coli</i> stationary- phase cells grown in the presence of glucose are larger than control cells. 1000×magnification. (C) To isolate and estimate persisters the stationary cultures were diluted to an OD₅₉₅ = 0.1 and exposed to ampicillin (200 µg/ml) for 10 h at 37°C. Antibiotic- tolerant bacteria were plated for colony counting. 100% corresponds to the total number of cells (dividing and non-dividing) before antibiotic treatment. Means and standard deviation of three independent experiments are shown.</p

Aggregation of proteins, formation of persisters and oxidation of proteins in the presence of acetate, MOPS and osmolytes.

<p>Bacteria were grown aerobically for 24 h at 37°C in LB medium supplemented with 0.2% acetate, 40 mM MOPS pH 7.4, 0.2% glucose, 0.2% glycerol, 0.2% trehalose or 0.2% betaine, unless otherwise stated. The control culture was grown in LB without supplements. Protein aggregates were isolated from 24 h cultures, resolved by SDS-PAGE and visualized by Coomassie staining. Oxidized, DNPH-derivatized proteins were resolved by SDS-PAGE and immunodetected using anti-dinitrophenol antibodies (E). The amount of aggregated proteins in relation to total protein in cell extracts (A) and the relative level of oxidized proteins (C) were calculated on the basis of densitometry. (B) After 24 h the cultures were diluted to an OD₅₉₅ = 0.1 and exposed to ampicillin (200 µg/ml) for 10 h at 37°C. Antibiotic- tolerant bacteria were plated for colony counting at the times indicated in the graph. 100% corresponds to the number of colonies before antibiotic treatment. Means and standard deviation of three independent experiments are shown. (D) Representative growth curves of selected cultures are shown. (E) Samples corresponding to the same OD₅₉₅ were loaded on the gels. Lane 1- the control culture, lane 2- LB+acetate, lane 3 - LB +MOPS, lane 4 - LB+trehalose, lane 5- LB+glucose. The asterics indicates EF-Tu, the most abundant protein in the aggregates.</p

The level of ATP and membrane stability in <i>E. coli</i> stationary-phase cultures.

<p>The cultures were grown as described in the legend to Figue 4. (A) ATP level was determined using BacTiter Glo chemicals (Promega). (B) Membrane stability was assessed after staining cells with the LIVE/DEAD assay solution (Invitrogen). Error bars represent standard deviation of three independent experiments. RFU, relative fluorescence units.</p

Stationary-phase <i>E. coli</i> cultures that accumulate protein aggregates produce increased levels of persisters.

<p>(A) Growth curves of <i>E. coli</i> MC4100. The bacteria were grown aerobically (circles) or anaerobically (triangles) at 37°C in LB medium supplemented (black symbols) or not (white symbols) with 0.2% glucose. (B) Protein aggregates were isolated from the stationary-phase cultures as described in Materials and Methods. Equal volumes of insoluble fractions isolated in the sucrose gradient were resolved by SDS-PAGE and visualized by Coomassie staining. (C) After 24 h the cultures were diluted to an OD₅₉₅ = 0.1 and exposed to ampicillin (200 µg/ml) for 10 h at 37°C. Antibiotic- tolerant bacteria were plated for colony counting at the times indicated in the graph. The error bars indicate the standard deviations of three independent experiments.</p

Screening for the Most Suitable Reference Genes for Gene Expression Studies in Equine Milk Somatic Cells

<div><p>Apart from the well-known role of somatic cell count as a parameter reflecting the inflammatory status of the mammary gland, the composition of cells isolated from milk is considered as a valuable material for gene expression studies in mammals. Due to its unique composition, in recent years an increasing interest in mare's milk consumption has been observed. Thus, investigating the genetic background of horse’s milk variability presents and interesting study model. Relying on 39 milk samples collected from mares representing three breeds (Polish Primitive Horse, Polish Cold-blooded Horse, Polish Warmblood Horse) we aimed to investigate the utility of equine milk somatic cells as a source of mRNA and to screen the best reference genes for RT-qPCR using geNorm and NormFinder algorithms. The results showed that despite relatively low somatic cell counts in mare's milk, the amount and the quality of the extracted RNA are sufficient for gene expression studies. The analysis of the utility of 7 potential reference genes for RT-qPCR experiments for the normalization of equine milk somatic cells revealed some differences between the outcomes of the applied algorithms, although in both cases the <i>KRT8</i> and <i>TOP2B</i> genes were pointed as the most stable. Analysis by geNorm showed that the combination of 4 reference genes (<i>ACTB</i>, <i>GAPDH</i>, <i>TOP2B</i> and <i>KRT8</i>) is required for apropriate RT-qPCR experiments normalization, whereas NormFinder algorithm pointed the combination of <i>KRT8</i> and <i>RPS9</i> genes as the most suitable. The trial study of the relative transcript abundance of the beta-casein gene with the use of various types and numbers of internal control genes confirmed once again that the selection of proper reference gene combinations is crucial for the final results of each real-time PCR experiment.</p></div

The effect of selection of different types and numbers of reference genes on beta-casein <i>(CSN2)</i> gene relative transcription level in equine milk somatic cells.

<p>Reference gene combinations: A − <i>KRT8 + TOP2B + ACTB + GAPDH</i> (the best combination according to geNORM); B − <i>KRT8 + TOP2B +ACTB</i>; C − <i>KRT8 + TOP2B</i>, D − <i>TOP2B + RPS9</i> (the best combination according to NormFinder), E − <i>KRT8</i> (alone—the most stable reference gene according to both algorithms).</p