Handling Protocols Research Articles

A new β-cyclodextrin-based nickel as green and water-soluble supramolecular catalysts for aqueous Suzuki reaction

A water-soluble nickel complex based on amino-β-CD was developed using a facile method and exhibits excellent catalytic performance in the Suzuki reaction in water. This synthesized complex has been characterized using UV–Vis, AAS, TGA, and FT-IR techniques. The easily synthesized novel supramolecular catalysts have been applied as a green and eco-friendly catalyst in the Suzuki coupling for preparing diverse biaryls. This result indicates that using 2.5 mol% of nickel, K2CO3 as the best base, and water as the green solvent are the best reaction conditions. This new catalyst features easy handling, low-cost, mild, and simple protocol. The use of low-cost and accessibility of the reagents, modest conditions, and good yields of products are notable characteristics of this method. Using aqueous media with this catalyst as a proper catalyst makes the presented process a fascinating method compared to most reports. Under mild reaction conditions, this green Ni(II)-β-CD catalyst displayed recyclable behavior seven times with minor loss in its catalytic activity.

Development and implementation of plasma P‐tau immunoassays

AbstractBackgroundMultiple advances in plasma biomarkers have been reported in recent times, one of these being the development of plasma p‐tau assays.MethodsIn this invited presentation as part of the Perspectives Session on “Advances in plasma Alzheimer’s disease biomarkers,” I will provide an expert review of the current state of analytical development of plasma p‐tau biomarkers.ResultsSpecifically, I will discuss the biochemical and molecular bases of why specific p‐tau forms are accessible in blood whilst others are not, as well as why these blood p‐tau markers tend to associate predominantly with brain amyloid but not tau pathology. Furthermore, I will highlight new discoveries in the development of biofluid‐based tau markers (both phosphorylated and non‐phosphorylated) that associate with neurofibrillary tangle pathology. Other aspects of my talk will focus on associations of the novel biomarker plasma brain‐derived tau with AD‐signature neurodegeneration. Additionally, I will discuss how newly improved preanalytical handling protocols enable plasma biomarker measurements in remote and resource‐limited settings, towards democratizing access to diagnostics.ConclusionFinally, I will discuss and suggest solutions to challenges around cut‐off generation, inter‐laboratory and inter‐assay measurements.

The Hong Kong genome project: building genome sequencing capacity and capability for advancing genomic science in Hong Kong

Aim: The Hong Kong Genome Project (HKGP) is the first large-scale genome sequencing (GS) project in the Hong Kong Special Administrative Region. The Hong Kong Genome Institute (HKGI) is entrusted with the task of implementing the HKGP. With the aim to sequence 45,000-50,000 genomes in five years, it is the project’s goal to provide participants with more precise diagnosis and personalised treatment, and to drive the application and integration of genomic medicine into routine clinical care. Methods: The HKGI Laboratory’s hardware and software components were customised to tailor to the needs of the project. Sample handling and storage protocol, DNA extraction, and PCR-free GS workflow were developed and optimised. Quality control indicators and metrics for assessing the quality of samples, sequencing libraries and sequencing data were established. Results: The Laboratory is designed to facilitate a unidirectional GS workflow to minimise the risk of contamination. The Sample Manager system handles laboratory data generated from the HKGP samples and biobank. The Laboratory handles and analyses approximately 350-500 samples per week, the majority of which are whole blood. During the first 24 months since the launch of the HKGP, 12,937 participants and their family members (6,680 genomes) have been recruited and sequenced. The sequencing capacity of the Laboratory has been further enhanced to include the latest technologies, such as long-read sequencing and multi-omics in order to meet the target of the HKGP. Conclusion: HKGI Laboratory established a robust GS workflow for the HKGP. The clinical utility of GS will bring precision medicine into routine clinical practice.

Regional sex differences in neurochemical profiles of healthy mice measured by magnetic resonance spectroscopy at 9.4 tesla

ObjectiveTo determine sex differences in the neurochemical concentrations measured by in vivo proton magnetic resonance spectroscopy (1H MRS) of healthy mice on a genetic background commonly used for neurodegenerative disease models.Methods1H MRS data collected from wild type mice with C57BL/6 or related genetic backgrounds in seven prior studies were used in this retrospective analysis. To be included, data had to be collected at 9.4 tesla magnetic field using advanced 1H MRS protocols, with isoflurane anesthesia and similar animal handling protocols, and a similar number of datasets from male and female mice had to be available for the brain regions analyzed. Overall, 155 spectra from female mice and 166 spectra from male mice (321 in total), collected from six brain regions (brainstem, cerebellum, cortex, hippocampus, hypothalamus, and striatum) at various ages were included.ResultsConcentrations of taurine, total creatine (creatine + phosphocreatine), ascorbate, glucose and glutamate were consistently higher in male vs. female mice in most brain regions. Striatum was an exception with similar total creatine in male and female mice. The sex difference pattern in the hypothalamus was notably different from other regions. Interaction between sex and age was significant for total creatine and taurine in the cerebellum and hippocampus.ConclusionSex differences in regional neurochemical levels are small but significant and age-dependent, with consistent male–female differences across most brain regions. The neuroendocrine region hypothalamus displays a different pattern of sex differences in neurochemical levels. Differences in energy metabolism and cellular density may underlie the differences, with higher metabolic rates in females and higher osmoregulatory and antioxidant capacity in males.

Monitoring contamination of hazardous drug compounding surfaces at hospital pharmacy departments. A consensus Statement. Practice guidelines of the Spanish Society of Hospital Pharmacists (SEFH)

ObjectiveTo establish a series of recommendations based on available evidence for monitoring surface contamination in the areas devoted to compounding hazardous drugs in pharmacy departments. MethodBased on a literature search in the Medline and Embase databases (search period: January 2009 to July 2019), as well as on a review of standards and recommendations issued by different healthcare organines a committee of experts from the Spanish Society of Hospital Pharmacists defined a series of safe practices for handling hazardous drugs and monitoring compounding work surfaces. Recommendation decisions were adopted by consensus among the members of the expert group, considering the recommendations reviewed, the monitoring situation in Spanish hospital departments, and the associated costs. ResultsTen recommendations were formulated, structured into eight sections. They include aspects related to the drugs to be monitored; the areas to be monitored; when samples should be taken; risk determination and preparation of a sampling protocol; analytical techniques; contamination thresholds; and design of an action plan based on the sampling and decontamination results obtained. ConclusionsSurface monitoring allows hazardous drugs detection and evaluation of the effectiveness of current protocols for the safe handling of such drugs in hospital pharmacy departments. The evaluation should include an analysis of the efficacy of engineering controls, work practices and cleaning and decontamination processes.

Single-cell RNA sequencing of bronchoscopy specimens: development of a rapid minimal handling protocol.

Single-cell RNA sequencing (scRNA-seq) is an important tool for understanding disease pathophysiology, including airway diseases. Currently, the majority of scRNA-seq studies in airway diseases have used invasive methods (airway biopsy, surgical resection), which carry inherent risks and thus present a major limitation to scRNA-seq investigation of airway pathobiology. Bronchial brushing, where the airway mucosa is sampled using a cytological brush, is a viable, less invasive method of obtaining airway cells for scRNA-seq. Here we describe the development of a rapid and minimal handling protocol for preparing single-cell suspensions from bronchial brush specimens for scRNA-seq. Our optimized protocol maximizes cell recovery and cell quality and facilitates large-scale profiling of the airway transcriptome at single-cell resolution.

A-123 Increased Rate of Recurrent Hemolysis Is Associated With COVID-19 Infection and Propofol Administration in the Intensive Care Unit (ICU)

Abstract Background Hemolysis is a pre-analytic issue that challenges clinical laboratories. When a specimen with significant hemolysis is obtained in our laboratory, tests where the hemolysis threshold is exceeded are not reported and blood is recollected. Unfortunately, in a subset of patients, the recollected specimen is also hemolyzed (termed “recurrent hemolysis”). In these cases, the second recollection utilizes special collection and handling protocols (i.e., hand-carried to lab, plasma collection, etc.) to mitigate preventable causes of hemolysis. Lipemia is also a known contributor to specimen hemolysis. Mayo Clinic Rochester experienced an increased rate of hemolysis as well as recurrent hemolysis during the height of the COVID-19 pandemic (January 2022). The objective of this study was to investigate the increased rate of recurrent hemolysis and compare to a rate prior to the COVID-19 pandemic (January 2019). Methods Retrospective data analysis was performed for January 2019 (negative control) and January 2022. The number of Mayo Clinic Rochester hospital admissions, admitting diagnosis, medications administered, and number of test collections performed were obtained using reporting functionality in our Electronic Health Record (EHR), Epic. The Laboratory Information System, SoftLab, was utilized to obtain recollection rates due to hemolysis. Data were normalized to total patient admissions for the specified period to account for inter-year variability. R/R Studio was utilized to analyze and visualize the data. Pearson’s Chi-squared test for independence was performed on categorical variables (significance defined as P < 0.05). Results A 28% decrease in hospital admissions (n = 3958:Jan-22 vs n = 5517:Jan-19) and a 23% decrease in total specimens collected on hospitalized patients (n = 40 557:Jan-22 vs n = 52 373:Jan-19) was observed between Jan-19 to Jan-22. However, the laboratory observed a 6% increase in hemolyzed specimens (n = 256:Jan-22 vs n = 241:Jan-19). Recurrent hemolysis occurred in 24% (n = 62/256) of Jan-22 and 17% (n = 43/241) of Jan-19 hemolyzed specimens and required special handling protocols. COVID positive patients comprised 48% (n = 30/62) of those requiring special handling protocols in Jan-22. Further examination revealed 34% (n = 21/62) of Jan-22 and 12% (n = 5/43) of Jan-19 cases of recurrent hemolysis were due to lipemia (P < 0.01). Chart review highlighted a potential source of lipemia from the administration of propofol, a lipophilic sedative emulsion. Of those COVID positive cases with recurrent hemolysis due to lipemia (n = 15/30), 93% (n = 14/15) were receiving propofol at the time of collection (P < 0.001). Conclusion We observed increased rates of specimen hemolysis during the COVID-19 pandemic. While most hemolysis resolves upon specimen recollection, patients with recurrently hemolyzed specimens often require special specimen collection and handling protocols to mitigate hemolysis. Approximately half of the recurrent hemolysis cases in Jan 2022 were COVID-19 positive patients. Lipemia-induced hemolysis was also more frequent in this patient population presumably due to the administration of propofol, a lipophilic emulsion sedative commonly used for COVID positive patients requiring sedation and ICU-level care. It is critical to understand the variety of factors which may contribute to recurrent hemolysis, as intervening with specialized collection and handling protocols to reduce observed pre-analytical hemolysis is required to provide accurate and timely laboratory results.

Chlorine Gas Poisoning in an Aquatic Exercise Pool: A Case Report

Background and Purpose: This case report describes an incident of chlorine gas poisoning in an aquatic exercise class of senior adults in which 24 participants, facility employees, and first responders required emergency medical attention with some requiring hospitalization. The cause of the chlorine gas release, the health effects on participants, and the facility's subsequent response are described. The purpose of this case is to inform physical therapists and exercise professionals working in aquatic exercise pools about chlorine gas poisoning prevention and mitigation efforts. Case Description: The cause of this incident was a series of breakdowns in safety procedures leading to the release of chlorine gas while a class was in session. Because of limited exit points, physical debilitation of some participants, and a lack of proper safety equipment, the evacuation effort was delayed, overwhelming participants, facility employees, and first responders with toxic gas. Twenty-four people were transported to the emergency department with 7 hospitalized. At least 3 are known to suffer long-term health effects. Outcomes: The facility concluded that normal operating procedures and safety protocols were not followed. Following the review, the facility took measures to ensure adherence to proper procedures, revised its emergency action plan, and acquired the appropriate safety equipment. Discussion: The accidental release of chlorine gas in an exercise pool is a rare but potentially catastrophic event. Facilities operating exercise and aquatic therapy pools must follow strict protocols for the safe handling of pool chemicals and be prepared to respond immediately if a chlorine gas accident occurs. This case report serves as an example of risk management for the prevention of and response to a catastrophic release of chlorine gas during a group exercise or aquatic therapy session.

Proof of concept of using a membrane-sensing peptide for sEVs affinity-based isolation.

Introduction: One main limitation in biomarker studies using EVs is the lack of a suitable isolation method rendering high yield and purity samples in a quick and easily standardized procedure. Here we report an affinity isolation method with a membrane-sensing peptide (MSP) derived from bradykinin. Methods: We designed a protocol based on agarose beads carrying cation chelates to specifically bind to the 6His-tagged membrane-sensing peptide. This approach presents several advantages: 1) cation-carrying agaroses are widely used and standardized for His-tagged protein isolation, 2) the affinity protocol can be performed in small volumes, feasible and manageable for clinical routine and 3) elution with imidazole or EDTA allows a gentle and easy recovery without EV damage, facilitating subsequent characterization and functional analyses. Results: The optimized final procedure incubates 0.5mg of peptide for 10min with 10µL of Long-arm Cobalt agarose before an overnight incubation with concentrated cell conditioned medium. EV downstream analyses can be directly performed on the agarose beads adding lysis or nucleic-acid extraction buffers, or gently eluted with imidazole or EDTA, rendering a fully competent EV preparation. Discussion: This new isolation methodology is based on the recognition of general membrane characteristics independent of surface markers. It is thus unbiased and can be used in any species EV sample, even in samples from animal or plant species against which no suitable antibodies exist. Being an affinity method, the sample handling protocol is very simple, less time-consuming, does not require specialized equipment and can be easily introduced in a clinical automated routine. We demonstrated the high purity and yield of the method in comparison with other commercially available kits. This method can also be scale up or down, with the possibility of analyzing very low amounts of sample, and it is compatible with any downstream analyses thanks to the gentle elution procedure.

Revising the Taiwan Code of Ethics for Nurses

The 2005 revision of the code of ethics for nurses has been in effect in Taiwan for more than 17 years. Although this code has been smoothly implemented during this time, changing social expectations and ethical perspectives, advancements in science and technology, and the evolution of the nurse-patient relationship suggest this code should be once again be updated. This study was conducted to suggest revisions to the Taiwan code of ethics for nurses necessary to meet current needs and address social and medical care environment trends. A multivariate research approach was adopted. The classification of the code of ethics for nurses norms in six international nursing organizations and evidence-based ethical and philosophical thinking literature were referenced, with the main notification demands incorporated into the Ministry of Health and Welfare's Nursing Workplace Controversy Reporting Platform. After drafting the aspects and provisions of Taiwan code of ethics for nurses norms, expert review procedures such as focus groups, Delphi expert consensus, and public forums were conducted. After three expert focus group discussion rounds, a structured questionnaire was completed, and 50 Delphi experts in six fields completed the online questionnaire. After the second consensus round, the importance and clarity of the 47 ethical code provisions in the four aspects were determined with 100% and 99.8% agreement reached, indicating no significant difference in scores between the multidisciplinary expert group and the ethical code. The resulting revision proposed for the Taiwan code of ethics for nurses includes: nursing staff and care recipients (14 provisions), nursing staff and practice (13 provisions), nursing staff and the profession (10 provisions), and nursing staff and society (10 provisions). In terms of education, the revised Taiwan code of ethics for nurses should be integrated into the nursing education curriculum of nursing colleges and used as teaching material for the continuing education of nurses. In terms of policy, these norms should be included as evaluation benchmarks and inspection items for hospitals. In addition, the attention and support of senior managers in institutions must be secured and a handling protocol for moral dilemma cases and related consultation mechanisms must be established. Nursing supervisors should be guided to develop the ability to address these dilemmas to help foster a positive workplace and a respectful team atmosphere. All professional groups should participate actively in promoting issues related to nursing ethics, organize seminars and continuing education activities, and make the revised Taiwan code of ethics for nurses and learning cases available online for reference by nursing staff nationwide.

Attitude, Knowledge and Compliance of the Community in Implementing the Protocol for Handling of COVID-19 to Prevent the Transmission of COVID-19 Infection: A Literacy Study

Currently, the world is still struggling with Covid-19 infection, which has become a world health problem since WHO officially declared it a pandemic on March 11, 2022. Based on the high risk and incidence of Covid-19 infection, it is necessary to educate about preventive behavior and understand the protocol in handling Covid-19 patients because the current conditions are, of course, a concern for all people related to efforts to control and deal with the Covid-19 pandemic in Indonesia. The purpose of this study is to analyze attitudes related to knowledge and compliance in implementing the Covid-19 handling protocol in a literacy study. The technique of writing this article is based on a type of qualitative research using a literature review approach. A literature review is an activity of studying various forms of relevant sources. The conclusion is that an understanding of attitudes, knowledge and compliance in carrying out the Covid-19 handling protocol will support breaking the chain of transmission of Covid-19 infection so as to reduce the death rate of patients infected with Covid-19 so that in carrying out the Covid-19 handling protocol requires attitude, knowledge and good obedience. Providing specific, valid and targeted knowledge can increase positive behavior in efforts to prevent Covid-19 infection. Compliance is one of the main keys to success in preventing the transmission of Covid-19 infection because it is a positive attitude and behavior towards a suggestion, procedure, or regulation that must be carried out or obeyed in preventing the spread of Covid-19 infection.

Sex differences in hippocampal structural plasticity and glycosaminoglycan disaccharide levels after neonatal handling.

In this study we investigated the effects of a neonatal handling protocol that mimics the handling of sham control pups in protocols of neonatal exposure to brain insults on dendritic arborization and glycosaminoglycan (GAG) levels in the developing brain. GAGs are long, unbranched polysaccharides, consisting of repeating disaccharide units that can be modified by sulfation at specific sites and are involved in modulating neuronal plasticity during brain development. In this study, male and female Sprague-Dawley rats underwent neonatal handling daily between post-natal day (PD)4 and PD9, with brains analyzed on PD9. Neuronal morphology and morphometric analysis of the apical and basal dendritic trees of CA1 hippocampal pyramidal neurons were carried out by Golgi-Cox staining followed by neuron tracing and analysis with the software Neurolucida. Chondroitin sulfate (CS)-, Hyaluronic Acid (HA)-, and Heparan Sulfate (HS)-GAG disaccharide levels were quantified in the hippocampus by Liquid Chromatography/Mass Spectrometry analyses. We found sex by neonatal handling interactions on several parameters of CA1 pyramidal neuron morphology and in the levels of HS-GAGs, with females, but not males, showing an increase in both dendritic arborization and HS-GAG levels. We also observed increased expression of glucocorticoid receptor gene Nr3c1 in the hippocampus of both males and females following neonatal handling suggesting that both sexes experienced a similar stress during the handling procedure. This is the first study to show sex differences in two parameters of brain plasticity, CA1 neuron morphology and HS-GAG levels, following handling stress in neonatal rats.

Exploring the microstructure of hydrated collagen hydrogels under scanning electron microscopy.

Collagen hydrogels are a rapidly expanding platform in bioengineering and soft materials engineering for novel applications focused on medical therapeutics, medical devices and biosensors. Observations linking microstructure to material properties and function enables rational design strategies to control this space. Visualisation of the microscale organisation of these soft hydrated materials presents unique technical challenges due to the relationship between hydration and the molecular organisation of a collagen gel. Scanning electron microscopy is a robust tool widely employed to visualise and explore materials on the microscale. However, investigation of collagen gel microstructure is difficult without imparting structural changes during preparation and/or observation. Electrons are poorly propagated within liquid-phase materials, limiting the ability of electron microscopy to interrogate hydrated gels. Sample preparation techniques to remove water induce artefactual changes in material microstructure particularly in complex materials such as collagen, highlighting a critical need to develop robust material handling protocols for the imaging of collagen hydrogels. Here a collagen hydrogel is fabricated, and the gel state explored under high-vacuum (10-6 Pa) and low-vacuum (80-120Pa) conditions, and in an environmental SEM chamber. Visualisation of collagen fibres is found to be dependent on the degree of sample hydration, with higher imaging chamber pressures and humidity resulting in decreased feature fidelity. Reduction of imaging chamber pressure is used to induce evaporation of gel water content, revealing collagen fibres of significantly larger diameter than observed in samples dehydrated prior to imaging. Rapid freezing and cryogenic handling of the gel material is found to retain a porous 3D structure following sublimation of the gel water content. Comparative analysis of collagen hydrogel materials demonstrates the care needed when preparing hydrogel samples for electron microscopy.

Impact of an external ventricular shunt (EVD) handling protocol on secondary meningitis rates: a historical cohort study with propensity score matching

BackgroundExternal ventricular drainage (EVD) is frequently used in neurosurgical procedures for cerebrospinal fluid (CSF) drainage. It is, however, associated with high infection rates, namely secondary meningitis and ventriculitis. Based on a previous high prevalence of these infections among patients with EVDs, we have proposed and implemented a protocol in an effort to decrease the infection rate. The aim of this study was to measure the effect of hospital-wide implementation of the EVD handling protocol on secondary EVD infections.Patients and methodsWe included 409 consecutive patients who received a new EVD for other indications than infectious pathologies from January 2000 until June 2012. Patients above 18 years of age were divided into pre- (n = 228) and post-protocol (n = 181) groups. Patient and disease demographics, as well as EVD data together with confounders for secondary meningitis were recorded in a database. Propensity score matching was then performed to create groups matched for sex, age, reason for drainage, type of shunt, time in situ and duration of surgery to place the EVD. Binomial logistic regression for confounder adjustment and regression discontinuity analyses were then performed on the matched cohort.ResultsInfections occurred more frequently in the pre-protocol group (23% vs 9%, p < 0.001). The incidence of infection was 33/1000 drain-days pre-protocol and 9/1000 drain-days post-protocol. Regression analysis in a propensity score-matched cohort (n = 103 in the pre- and n = 178 in the post-protocol groups) showed that the pre-protocol period was independently associated with more infections (OR 2.69; 95%-CI 1.22–5.95, p = 0.01).ConclusionsThe incidence of secondary EVD infections can be reduced significantly by the implementation of a strict hospital-wide EVD handling protocol.

Impact of pre-analytical sample handling factors on plasma biomarkers of Alzheimer's disease.

An unmet need exists for reliable plasma biomarkers of amyloid pathology, in the clinical laboratory setting, to streamline diagnosis of Alzheimer's disease (AD). For routine clinical use, a biomarker must provide robust and reliable results under pre-analytical sample handling conditions. We investigated the impact of different pre-analytical sample handling procedures on the levels of seven plasma biomarkers in development for potential routine use in AD. Using (1) fresh (never frozen) and (2) previously frozen plasma, we evaluated the effects of (A) storage time and temperature, (B) freeze/thaw (F/T) cycles, (C) anticoagulants, (D) tube transfer, and (E) plastic tube types. Blood samples were prospectively collected from patients with cognitive impairment undergoing investigation in a memory clinic. β-amyloid 1-40 (Aβ40), β-amyloid 1-42 (Aβ42), apolipoprotein E4, glial fibrillary acidic protein, neurofilament light chain, phosphorylated-tau (phospho-tau) 181, and phospho-tau-217 were measured using Elecsys® plasma prototype immunoassays. Recovery signals for each plasma biomarker and sample handling parameter were calculated. For all plasma biomarkers measured, pre-analytical effects were comparable between fresh (never frozen) and previously frozen samples. All plasma biomarkers tested were stable for ≤24 h at 4°C when stored as whole blood and ethylenediaminetetraacetic acid (EDTA) plasma. Recovery signals were acceptable for up to five tube transfers, or two F/T cycles, and in both polypropylene and low-density polyethylene tubes. For all plasma biomarkers except Aβ42 and Aβ40, analyte levels were largely comparable between EDTA, lithium heparin, and sodium citrate tubes. Aβ42 and Aβ40 were most sensitive to pre-analytical handling, and the effects could only be partially compensated by the Aβ42/Aβ40 ratio. We provide recommendations for an optimal sample handling protocol for analysis of plasma biomarkers for amyloid pathology AD, to improve the reproducibility of future studies on plasma biomarkers assays and for potential use in routine clinical practice.

Effects of temperature on metabolic rate and lower dissolved oxygen tolerance of juvenile speckled peacock bass Cichla temensis.

The speckled peacock bass Cichla temensis is a popular sport and food fish that generates substantial angling tourism and utilitarian harvest within its range. Its popularity and value make this species important for management and a potential aquaculture candidate for both fisheries enhancement and food fish production. However, little is known of optimal physiochemical conditions in natural habitats, which also are important for the development of hatchery protocols for handling, spawning and grow-out. Speckled peacock bass have been documented to have high sensitivity to extreme temperatures, but the metabolic underpinnings have not been evaluated. In this study, the effects of temperature (25, 30 and 35°C) on the standard metabolic rate (SMR) and lower dissolved oxygen tolerance (LDOT) of juvenile speckled peacock bass (mean ± standard error total length 153 ± 2 mm and wet weight 39.09 ± 1.37 g) were evaluated using intermittent respirometers after an acclimation period of 2 weeks. Speckled peacock bass had the highest SMR at 35°C (345.56 ± 19.89 mgO2 kg-1 h-1 ), followed by 30°C (208.16 ± 12.45 mgO2 kg-1 h-1 ) and 25°C (144.09 ± 10.43 mgO2 kg-1 h-1 ). Correspondingly, the Q10 , or rate of increase in aerobic metabolic rate (MO2 ) relative to 10°C, for 30-35°C was also greater (2.76) than from 25 to 30°C (2.08). Similarly, speckled peacock bass were the most sensitive to hypoxia at the warmest temperature, with an LDOT at pO2 of 90 mmHg (4.13 mg l-1 ) at 35°C compared to pO2 values of 45 mmHg (2.22 mg l-1 ) and 30 mmHg (1.61 mg l-1 ) at 30 and 25°C, respectively. These results indicate that speckled peacock bass are sensitive to temperatures near 35°C, therefore we recommend managing and rearing this species at 25-30°C.

Getting a handle on rat familiarization: The impact of handling protocols on classic tests of stress in Rattus norvegicus.

Experimenter familiarization with laboratory rodents through handling prior to experimentation is an important practice in neurobehavioral research and is implicated in stress, study variability, and replicability. Unfortunately, different handling protocols have not been thoroughly examined. Determining optimal experimenter familiarization protocols is expected to reduce animal stress and thus improve welfare and data consistency. The impact of different handling protocols was determined through behavioral assessments (i.e. elevated plus maze, light/dark box, open field) as well as via analysis of fecal boli counts, ultrasonic vocalizations, and blood corticosterone. Male and female Sprague Dawley rats were distributed among three groups: never handled, picked-up, and handled for 5 min once daily over five days. Handled and picked-up rats spent more time in open arms and less time in closed arms of the elevated plus maze and more time in the center and less time at the perimeter of the open field compared to rats that were never handled, indicating that handled and picked-up rats were less anxious than those that were never handled. Male rats consistently defecated more frequently throughout the handling process and throughout behavioral testing, whereas females showed greater concentrations of blood corticosterone. Female rats were found to emit more 50-kHz calls and fewer 22-kHz calls compared to males. The results observed suggest that picking animals up may suffice as a handling method compared to time-intensive handling procedures, and that there are significant sex differences in response to handling.

Prognostic relevance of immune-related adverse events in lung cancer patients undergoing immune checkpoint inhibitor therapy: a systematic review and meta-analysis.

Immune checkpoint inhibitors (ICIs) work by activating the immune system, a mechanism that may also cause immune-related adverse events (irAEs). This study seeks to investigate on how different irAEs impact prognosis of advanced lung cancer (LC) patients and identify useful approaches to manage irAEs. A thorough literature search of PubMed, Embase, the Cochrane Library and manual searches up to January 2024 were undertaken. Treatment outcomes including progression-free survival (PFS), overall survival (OS), objective response rate (ORR), and disease control rate (DCR) were obtained. Meta-analysis was conducted using R software (version 4.3.1). There were 106 studies with 41,050 advanced or recurrent LC patients included. The occurrence of irAEs was correlated with better PFS [hazard ratio (HR) =0.54; 95% confidence interval (CI): 0.49-0.59], OS (HR =0.57; 0.51-0.63), ORR [risk ratio (RR) =2.03; 95% CI: 1.81-2.28] and DCR (RR =1.55; 95% CI: 1.40-1.72) and remained significant after adjusting programmed death-ligand 1 (PD-L1) level. IrAEs affecting skin (OS: HR =0.45; 95% CI: 0.38-0.53) and endocrine system (OS: HR =0.51; 95% CI: 0.41-0.62), of mild severity (OS: HR =0.52; 95% CI: 0.35-0.79), arising in multiple sites (OS: HR =0.47; 95% CI: 0.38-0.59), induced by monotherapy (OS: HR =0.58; 95% CI: 0.52-0.65), with a delayed onset (cutoff: 3 months; OS: HR =0.37; 95% CI: 0.19-0.71) were identified as positive prognostic markers. In contrast, though pulmonary irAEs were found to be corelated with enhanced treatment response (ORR: RR =1.75; 95% CI: 1.37-2.25), they may harm survival, especially those with grade ≥3 (OS: HR =2.40; 95% CI: 1.39-4.14). Treatment resumption tended to improve PFS but might not reduce the risk of death compared to permanent discontinuation. IrAEs suggest better treatment outcomes generally, yet severe pneumonia could increase mortality risk. Close supervision and appropriate handling protocols are warranted to weigh treatment benefit against risk.

Multimodal Techniques to Study Tumor Growth, Basement Membrane Breaching, and Invasion in 3D Matrices.

Cancer-derived organoids and three-dimensional (3D) extracellular matrix (ECM) are taking center stage as in vitro models to study neoplastic cell behavior, since they recapitulate the heterogeneous cellular composition of tumors and their extracellular environment. In combination with imaging and molecular/biochemical techniques, 3D organoid models have contributed substantially to our knowledge about the cellular and molecular mechanisms that regulate the growth of tumors and invasion into the surrounding tissue. We here outline a set of protocols that describe culturing of cancer-derived organoids in 3D matrices and various strategies that allow modeling of tumor growth, tumor cell penetration into basement membranes, and invasion into Collagen I-rich ECM. Furthermore, we specify protocols for subsequent handling of organoids culturedin 3D ECM for confocal microscopy and analysis of gene expression at the protein and mRNA level. Although we here use breast cancer-derived organoids, these protocols can be directly applied or adapted for organoids derived from other cancer types or healthy tissues. Thus, in addition to investigating cell behavior of multiple cancer types, the combination of protocols described here may be used to study processes such as cell differentiation and migration during homeostasis and normal development.

Current practice of saphenous vein graft harvesting in coronary artery bypass graft surgery: an Indian survey.

The treatment of occluded saphenous vein graft (SVG) is challenging, and thus preventing the graft occlusion is of utmost importance. However, despite its recognized importance, a paucity of data exists regarding howSVGsare handled and preserved. Hence, this survey was conducted to document the techniques ofveingraftpreservation and handling among cardiac surgeons in India. The survey had 26 questions regarding vein graft usage, harvesting, handling, and preservation techniques. Three hundred cardiac surgeons across India participated in this survey between March 2019 and July 2019. Responses were received from 215 (71.6%) surgeons across 13 states. Around 87% of respondents reported that ≥ 76% of veins were harvested by the open technique. Among the respondents, around 67% used one SVG in ≥ 96% of their patients, 45% used two SVGs in ≥ 86% of their patients, and 38% used ≥ 3 SVGs in < 25% of their patients, respectively. Around 54%, 27%, 9%, and 9% of respondents used autologous whole blood, saline solution, pH-buffered solution, and other solutions, respectively. In addition, 96.74% of respondents heparinized their vein graft preservation solutions, and 98.14% preserved the solution at room temperature. 34.88% and 83.26% of respondents used dual antiplatelet therapy (DAPT) before and after surgery in ≥ 76% of their patients, respectively. 63.26% of the surgeons reported that the patients received DAPT for > 12months. There is a significant variance in the process of vein harvesting, preservation, handling, and antiplatelet therapy protocols among various cardiac surgeons across India. Therefore, there is a need for standardization in the practice of vein harvesting.