Prediction Modelling for Gastroesophageal Variceal Bleeding in Patients With Chronic Hepatitis B Using Four-dimensional Flow MRI.

Background/Aims: In this study, we aim to develop a model for predicting gastroesophageal varices (GEV) bleeding in patients with chronic hepatitis B (CHB) by utilizing hemodynamic parameters obtained through four-dimensional flow MRI (4D flow MRI). Methods: This study conducted a prospective enrollment of CHB patients suspected of GEV from October 2021 to May 2022. The severity of varices and bleeding risk were evaluated using clinical findings and upper gastrointestinal endoscopy, and patients were classified into high-risk and non-high-risk groups. The study utilized serological examination, ultrasonographic examination, and 4D flow MRI. Relevant parameters were selected through univariate and multivariate analyses, and a prediction model was established using binary logistic regression analysis. The model was combined with the Baveno Ⅵ/Ⅶ and Expanded Baveno Ⅵ/Ⅶ criteria to evaluate diagnostic efficacy and the risk of avoiding endoscopic examination. Results: A total of 40 CHB patients were enrolled and categorized into the high-risk group (n = 15) and the non-high-risk group (n = 25). The spleen diameter and regurgitant fraction (R%) were independent predictors of variceal bleeding and a predictive model was established. The combination of this prediction model and the Baveno Ⅵ/Ⅶ criteria achieved high diagnostic efficiency, enabling 45.00% (18/40) of patients to be exempted from the unnecessary endoscopic procedure and the high-risk misclassification rate (0%) was less than 5%. Conclusion: The prediction model generated by 4D flow MRI has the potential to assess the likelihood of varices and can be supplemented by the Baveno VI/VII criteria to improve diagnostic accuracy in CHB patients.

Assessment of Viability of Listeria monocytogenes by Flow Cytometry.

In food industry, Listeria monocytogenes contamination can occur accidentally despite the quality control of raw materials and factory. Decontamination processes or inhibitory effects of ingredients/additives in food products are set up to ensure compliance with hygiene and microbiological criteria. These actions represent stresses for the pathogenic agent, causing fluctuations in its physiological states. Moreover, during these environmental stresses, Listeria monocytogenes can enter in a viable but nonculturable (VBNC) state which is not detected by plate counting but by flow cytometry. This technique coupled with cell staining by fluorescent dyes offers the possibility to assess different physiological states based on different cellular parameters: enzymatic activity, transmembrane integrity, membrane potential, and respiratory activity. In this chapter, we present a method to assess the viability of foodborne pathogens using a double-staining principle based on the assessment of membrane integrity and intracellular esterase activity.

Rapid visual detection of Helicobacter pylori and vacA subtypes by Dual-Target RAA-LFD assay.

BACKGROUND: Helicobacter pylori (H. pylori) infects over 50% of the global population and is a significant risk factor for gastric cancer. The pathogenicity of H. pylori is primarily attributed to virulence factors such as vacA. Timely and accurate identification, along with genotyping of H. pylori virulence genes, are essential for effective clinical management and controlling its prevalence. METHODS: In this study, we developed a dual-target RAA-LFD assay for the rapid, visual detection of H. pylori genes (16s rRNA, ureA, vacA m1/m2), using recombinase aided amplification (RAA) combined with lateral flow dipstick (LFD) methods. Both 16s rRNA and ureA were selected as identification genes to ensure reliable detection accuracy. RESULTS: A RAA-LFD assay was developed to achieve dual-target amplification at a stable 37 °C within 20 min, followed by visualization using the lateral flow dipstick (LFD). The whole process, from amplification to results, took less than 30 min. The 95 % limit of detection (LOD) for 16 s rRNA and ureA, vacA m1, vacA m2 were determined as 3.8 × 10-2 ng/µL, 5.8 × 10-2 ng/µL and 1.4 × 10-2 ng/µL, respectively. No cross-reaction was observed in the detection of common pathogens including Escherichia coli, Klebsiella pneumoniae, Enterococcus faecalis, Staphylococcus aureus, Pseudomonas aeruginosa, and Bacillus subtilis, showing the assay's high specificity. In the evaluation of the clinical performance of the RAA-LFD assay. A total of 44 gastric juice samples were analyzed, immunofluorescence staining (IFS) and quantitative polymerase chain reaction (qPCR) were used as reference methods. The RAA-LFD results for the 16s rRNA and ureA genes showed complete agreement with qPCR findings, accurately identifying H. pylori infection as confirmed by IFS in 10 out of the 44 patients. Furthermore, the assay successfully genotyped vacA m1/m2 among the positive samples, showing complete agreement with qPCR results and achieving a kappa (κ) value of 1.00. CONCLUSION: The dual-target RAA-LFD assay developed in this study provides a rapid and reliable method for detecting and genotyping H. pylori within 30 min, minimizing dependency on sophisticated laboratory equipment and specialized personnel. Clinical validation confirms its efficacy as a promising tool for effectively control of its prevalence and aiding in the precise treatment of H. pylori-associated diseases.

Flow cytometry in acute myeloid leukemia and detection of minimal residual disease.

Acute myeloid leukemia (AML) is a common type of acute leukemia (AL), belonging to malignant tumors of the hematopoietic system with the characteristics of rapid disease development, control with extreme difficulties, easy recurrence, poor prognosis, and incidence rate increasing with age. The traditionally diagnostic standard of French American British (FAB), being based on the morphological examination with high human subjectivity, can no longer meet the demand of clinical diagnosis and treatment of AML. Requirements of objective accuracy and low-dose sample, have become the indispensable method for AML diagnosis and monitoring prognosis. Flow cytometry is a modern technology that can quickly and accurately detect the series, antigen distribution, differentiation stage of AML cells, minimal residual lesions after AML therapy, so as to provide the great significance in guiding clinical diagnosis, hierarchical treatment, and prognosis judgement. This article will systematically elaborate on the application of flow cytometry in the diagnosis and classification of AML, and the detection of minimal residual lesions, thereby providing reference significance for dynamic monitoring and prognostic observation of AML with different immune subtypes of FAB.

Boundary-based registration improves sensitivity for detecting hypoperfusion in sporadic frontotemporal lobar degeneration.

Introduction: Frontotemporal lobar degeneration (FTLD) is associated with FTLD due to tau (FTLD-tau) or TDP (FTLD-TDP) inclusions found at autopsy. Arterial Spin Labeling (ASL) MRI is often acquired in the same session as a structural T1-weighted image (T1w), enabling detection of regional changes in cerebral blood flow (CBF). We hypothesize that ASL-T1w registration with more degrees of freedom using boundary-based registration (BBR) will better align ASL and T1w images and show increased sensitivity to regional hypoperfusion differences compared to manual registration in patient participants. We hypothesize that hypoperfusion will be associated with a clinical measure of disease severity, the FTLD-modified clinical dementia rating scale sum-of-boxes (FTLD-CDR). Materials and methods: Patients with sporadic likely FTLD-tau (sFTLD-tau; N = 21), with sporadic likely FTLD-TDP (sFTLD-TDP; N = 14), and controls (N = 50) were recruited from the Connectomic Imaging in Familial and Sporadic Frontotemporal Degeneration project (FTDHCP). Pearson's Correlation Coefficients (CC) were calculated on cortical vertex-wise CBF between each participant for each of 3 registration methods: (1) manual registration, (2) BBR initialized with manual registration (manual+BBR), (3) and BBR initialized using FLIRT (FLIRT+BBR). Mean CBF was calculated in the same regions of interest (ROIs) for each registration method after image alignment. Paired t-tests of CC values for each registration method were performed to compare alignment. Mean CBF in each ROI was compared between groups using t-tests. Differences were considered significant at p < 0.05 (Bonferroni-corrected). We performed linear regression to relate FTLD-CDR to mean CBF in patients with sFTLD-tau and sFTLD-TDP, separately (p < 0.05, uncorrected). Results: All registration methods demonstrated significant hypoperfusion in frontal and temporal regions in each patient group relative to controls. All registration methods detected hypoperfusion in the left insular cortex, middle temporal gyrus, and temporal pole in sFTLD-TDP relative to sFTLD-tau. FTLD-CDR had an inverse association with CBF in right temporal and orbitofrontal ROIs in sFTLD-TDP. Manual+BBR performed similarly to FLIRT+BBR. Discussion: ASL is sensitive to distinct regions of hypoperfusion in patient participants relative to controls, and in patients with sFTLD-TDP relative to sFTLD-tau, and decreasing perfusion is associated with increasing disease severity, at least in sFTLD-TDP. BBR can register ASL-T1w images adequately for controls and patients.

Case report: Clinical, genetic and immunological characterization of a novel XK variant in a patient with McLeod syndrome.

Introduction: Pathogenic variants in the XK gene are associated with dysfunction or loss of XK protein leading to McLeod syndrome (MLS), a rare X-linked neuroacanthocytosis syndrome with multisystemic manifestation. Here we present clinical, genetic and immunological data on a patient originally admitted to our clinic for presumed post-COVID-19 syndrome, where thorough clinical work-up revealed a novel frameshift deletion in XK causal for the underlying phenotype. We additionally review the clinicogenetic spectrum of reported McLeod cases in the literature. Methods: We performed in-depth clinical characterization and flow cytometry of cerebrospinal fluid (CSF) in a patient where multi-gene panel sequencing identified a novel hemizygous frameshift deletion in XK. Additionally, Kell (K) and Cellano (k) antigen expression was analysed by Fluorescence-activated Cell Sorting (FACS). KEL gene expression was examined by RNA sequencing. Results: A novel hemizygous frameshift deletion in the XK gene resulting in premature termination of the amino acid chain was identified in a 46-year old male presenting with decrease in physical performance and persisting fatigue after COVID-19 infection. Examinations showed raised creatine kinase (CK) levels, neuropathy and clinical features of myopathy. FACS revealed the K-k+ blood type and reduced Cellano density. CSF flow cytometry showed elevation of activated T Cells. Conclusion: In-depth clinical, genetic, immunological and ribonucleic acid (RNA) expression data revealed axonal neuropathy, myopathy and raised levels of activated CSF-T-lymphocytes in a patient with a previously unpublished frameshift deletion in the XK gene.

ERA-CRISPR/Cas12a system: a rapid, highly sensitive and specific assay for Mycobacterium tuberculosis.

Introduction: Mycobacterium tuberculosis, the causative agent of human tuberculosis, poses a significant threat to global public health and imposes a considerable burden on the economy. However, existing laboratory diagnostic methods for M. tuberculosis are time-consuming and have limited sensitivity levels. Methods: The CRISPR/Cas system, commonly known as the "gene scissors", demonstrates remarkable specificity and efficient signal amplification capabilities. Enzymatic recombinase amplification (ERA) was utilized to rapidly amplify trace DNA fragments at a consistent temperature without relying on thermal cyclers. By integrating of CRISPR/Cas12a with ERA, we successfully developed an ERA-CRISPR/Cas12a detection system that enables rapid identification of M. tuberculosis. Results: The sensitivity of the ERA-CRISPR/Cas12a fluorescence and lateral flow systems was 9 copies/µL and 90 copies/µL, respectively. Simultaneously, the detection system exhibited no cross-reactivity with various of respiratory pathogens and non-tuberculosis mycobacteria, demonstrating a specificity of 100%. The positive concordance rate between the ERA-CRISPR/Cas12a fluorescence system and commercial qPCR was 100% in 60 clinical samples. Meanwhile, the lateral flow system showed a positive concordance rate of 93.8% when compared to commercial qPCR. Both methods demonstrated a negative concordance rate of 100%, and the test results can be obtained in 50 min at the earliest. Discussion: The ERA-CRISPR/Cas12a system offers a rapid, sensitive, and specific method that presents a novel approach to laboratory diagnosis of M. tuberculosis.

Humoral and cellular immunity in response to an in silico-designed multi-epitope recombinant protein of Theileria annulata.

Tropical theileriosis is a lymphoproliferative disease caused by Theileria annulata and is transmitted by Ixodid ticks of the genus Hyalomma. It causes significant losses in livestock, especially in exotic cattle. The existing methods for controlling it, chemotherapeutic agents and a vaccine based on an attenuated schizont stage parasite, have several limitations. A promising solution to control this disease is the use of molecular vaccines based on potential immunogenic proteins of T. annulata. For this purpose, we selected five antigenic sequences of T. annulata, i.e. SPAG-1, Tams, TaSP, spm2, and Ta9. These were subjected to epitope prediction for cytotoxic T lymphocytes, B-cells, and helper T lymphocytes. CTL and B-cell epitopes with a higher score whereas those of HTL with a lower score, were selected for the construct. A single protein was constructed using specific linkers and evaluated for high antigenicity and low allergenicity. The construct was acidic, hydrophobic, and thermostable in nature. Secondary and tertiary structures of this construct were drawn using the PSIPRED and RaptorX servers, respectively. A Ramachandran plot showed a high percentage of residues in this construct in favorable, allowed, and general regions. Molecular docking studies suggested that the complex was stable and our construct could potentially be a good candidate for immunization trials. Furthermore, we successfully cloned it into the pET-28a plasmid and transformed it into the BL21 strain. A restriction analysis was performed to confirm the transformation of our plasmid. After expression and purification, recombinant protein of 49 kDa was confirmed by western blotting. An ELISA detected increased specific antibody levels in the sera of the immunized animals compared with the control group, and flow cytometric analysis showed a stronger cell-mediated immune response. We believe our multi-epitope recombinant protein has the potential for the large-scale application for disease prevention globally in the bovine population. This study will act as a model for similar parasitic challenges.

Exploring cell-derived extracellular vesicles in peripheral blood and bone marrow of B-cell acute lymphoblastic leukemia pediatric patients: proof-of-concept study.

Extracellular vesicles (EVs) are heterogeneous, phospholipid membrane enclosed particles that are secreted by healthy and cancerous cells. EVs are present in diverse biological fluids and have been associated with the severity of diseases, which indicates their potential as biomarkers for diagnosis, prognosis and as therapeutic targets. This study investigated the phenotypic characteristics of EVs derived from peripheral blood (PB) and bone marrow (BM) in pediatric patients with B-cell acute lymphoblastic leukemia (B-ALL) during different treatment stages. PB and BM plasma were collected from 20 B-ALL patients at three time points during induction therapy, referred to as: diagnosis baseline (D0), day 15 of induction therapy (D15) and the end of the induction therapy (D35). In addition, PB samples were collected from 10 healthy children at a single time point. The EVs were measured using CytoFLEX S flow cytometer. Calibration beads were employed to ensure accurate size analysis. The following, fluorescent-labeled specific cellular markers were used to label the EVs: Annexin V (phosphatidylserine), CD235a (erythrocyte), CD41a (platelet), CD51 (endothelial cell), CD45 (leukocyte), CD66b (neutrophil), CD14 (monocyte), CD3 (T lymphocyte), CD19, CD34 and CD10 (B lymphoblast/leukemic blast). Our results demonstrate that B-ALL patients had a marked production of EV-CD51/61+, EV-CD10+, EV-CD19+ and EV-CD10+CD19+ (double-positive) with a decrease in EV-CD41a+ on D0. However, the kinetics and signature of production during induction therapy revealed a clear decline in EV-CD10+ and EV-CD19+, with an increase of EV-CD41a+ on D35. Furthermore, B-ALL patients showed a complex biological network, exhibiting distinct profiles on D0 and D35. Interestingly, fold change and ROC curve analysis demonstrated that EV-CD10+CD19+ were associated with B-ALL patients, exhibited excellent clinical performance and standing out as a potential diagnostic biomarker. In conclusion, our data indicate that EVs represent a promising field of investigation in B-ALL, offering the possibility of identifying potential biomarkers and therapeutic targets.

Release of contaminants from polymer surfaces under condition of organized fluid flows.

The use of polymers for water storage or distribution is closely monitored, especially with regard to the possible contamination with substances coming from the material's surfaces. Different standards are practiced across countries according to type of applied materials and such test methods are prevalently based on constant temperature conditions. However, these polymers systems could be located in diverse environment which does not necessarily provide constant conditions. Experimental findings show that exposure of liquid inside polymeric materials to specific temperature gradients, and consequently to emerging organized flows, can result in an accelerated leaching of undesirable substances from the solid surface. In presented work model steady-state and organized flow conditions are used to compare release of contaminates from polyethylene by measuring of surface tension, UV-Vis spectroscopy, FTIR, scanning electron microscopy and elemental analysis of polymer surfaces and water leachates. The pilot study shows that convective flow generated via temperature gradient significantly affects contaminant release in comparison to a steady state and mixing flow conditions.

Exploring the impact of pre-anastomosis cerebral microcirculation on cerebral hyperperfusion syndrome in superficial temporal artery-middle cerebral artery bypass surgery of moyamoya disease.

Significance: Cerebral hyperperfusion syndrome (CHS), characterized by neurologic deficits due to postoperative high cerebral perfusion, is a serious complication of superficial temporal artery-middle cerebral artery (STA-MCA) surgery for moyamoya disease (MMD). Aim: We aim to clarify the importance of assessing pre-anastomosis cerebral microcirculation levels by linking the onset of CHS to pre- and post-anastomosis hemodynamics. Approach: Intraoperative laser speckle contrast imaging (LSCI) measured changes in regional cerebral blood flow (rCBF) and regional blood flow structuring (rBFS) within the cerebral cortical microcirculation of 48 adults with MMD. Results: Following anastomosis, all MMD patients exhibited a significant increase in rCBF ( 279.60 % ± 120.00 % , p < 0.001 ). Changes in rCBF and rBFS showed a negative correlation with their respective baseline levels (rCBF, p < 0.001 ; rBFS, p = 0.005 ). Baseline rCBF differed significantly between CHS and non-CHS groups ( p = 0.0049 ). The areas under the receiver operating characteristic (ROC) curve for baseline rCBF was 0.753. Hemorrhagic MMD patients showed higher baseline rCBF than ischemic patients ( p = 0.036 ), with a marked correlation between pre- and post-anastomosis rCBF in hemorrhagic cases ( p = 0.003 ), whereas ischemic MMD patients did not. Conclusion: Patients with low levels of pre-anastomosis baseline CBF induce a dramatic increase in post-anastomosis and show a high risk of postoperative CHS.

Coronary microvascular dysfunction in autoimmune rheumatic diseases: beyond coronary flow velocity reserve.

Autoimmune rheumatic diseases (ARDs) are a heterogeneous group of disorders characterized by an inappropriate immune reactivity against different body tissues. Patients affected by ARDs present increased cardiovascular morbidity and mortality, which significantly impacts long-term prognosis. Endothelial dysfunction, inflammation, oxidative stress, and autoimmunity are strictly involved in atherosclerosis progression and coronary microvascular dysfunction (CMD), both of which contribute to increased cardiovascular risk. CMD represents the inability of the coronary microvasculature to respond with vasodilation to increased cardiac metabolic demands and can be assessed by non-invasive and invasive imaging tests. Coronary flow velocity reserve assessed by echocardiography has been demonstrated to accurately identify ARDs patients with CMD. However, stress cardiac magnetic resonance (CMR) accurately assesses myocardial ischemia, perfusion, and viability in ARDs patients. The myocardial perfusion reserve index (MPRI) is a robust semiquantitative imaging marker that represents the vasodilatory capacity of the coronary microcirculation in response to a vasodilator stress. In the absence of significant coronary stenosis, ARDs patients revealed a reduced MPRI in comparison with the general population, regardless of the presence of myocardial fibrosis. Identification of CMD in asymptomatic patients could be crucial to precociously start targeted medical therapy, avoiding major adverse cardiac events in this clinical setting. This review aims to summarize the current evidence regarding CMD in ARDs patients, focusing on the role of stress CMR and the promising myocardial perfusion analysis.

Hypoperfusion of periaqueductal gray as an imaging biomarker in chronic migraine beyond diagnosis: A 3D pseudocontinuous arterial spin labeling MR imaging.

BACKGROUND: The periaqueductal gray (PAG) is at the center of a powerful descending antinociceptive neuronal network, and is a key node in the descending pain regulatory system of pain. However, less is known about the altered perfusion of PAG in chronic migraine (CM). AIM: To measure the perfusion of PAG matter, an important structure in pain modulation, in CM with magnetic resonance (MR) perfusion without contrast administration. METHODS: Three-dimensional pseudocontinuous arterial spin labeling (3D-PCASL) and brain structure imaging were performed in 13 patients with CM and 15 normal subjects. The inverse deformation field generated by brain structure image segmentation was applied to the midbrain PAG template to generate individualized PAG. Then the perfusion value of the PAG area of the midbrain was extracted based on the individual PAG mask. RESULTS: Cerebral blood flow (CBF) value of PAG in CM patients (47.98 ± 8.38 mL/100 mg min) was significantly lower than that of the control group (59.87 ± 14.24 mL/100 mg min). Receiver operating characteristic (ROC) curve analysis showed that the area under the curve was 0.77 (95% confidence interval [CI], 0.60, 0.94), and the cutoff value for the diagnosis of CM was 54.83 mL/100 mg min with a sensitivity 84.60% and a specificity 60%. CONCLUSION: Imaging evidence of the impaired pain conduction pathway in CM may be related with the decreased perfusion in the PAG, which could be considered as an imaging biomarker for the diagnosis and therapy evaluation.

Effects of intracranial artery stenosis of anterior circulation on cognition-A CT perfusion-based study.

BACKGROUND: Intracranial atherosclerotic stenosis (ICAS) is one of the most important independent risk factors for stroke that is closely related to the occurrence of cognitive impairment. The relationship between ICAS and vascular cognitive impairment (VCI) remains unclear. Cerebral hemodynamic changes are one of the main causes of cognitive impairment. Computed tomographic perfusion (CTP) imaging can quantitatively analyze cerebral blood perfusion and quantify cerebral hemodynamic changes. Previous research on the relationship between hypoperfusion induced by ICAS and cognitive impairment, as well as its underlying mechanisms, remains relatively insufficient. This study is dedicated to elucidating the characteristics and potential mechanisms of cognitive impairment in ICAS patients with abnormal perfusion, utilizing CTP imaging as our primary investigative tool. METHODS: This study recruited 82 patients who suffer from non-disabling ischemic stroke (IS group) and 28 healthy controls. All participants underwent comprehensive neuropsychological assessments both collectively and individually, in addition to CTP imaging. Within the patient group, we further categorized individuals into two subgroups: the ischemic penumbra group (IP, N = 28) and the benign oligemia group (BO, N = 54), based on CTP parameters-Tmax. The correlations between cognitive function and abnormal perfusion were explored. RESULTS: The cognitive function, including the overall cognitive, memory, attention, executive functions, and language, was significantly impaired in the IS group compared with that in the control group. Further, there are statistical differences in the stroop color-word test-dot (Stroop-D) and Montreal Cognitive Assessment (MoCA) sub-items (memory + language) between the BO and IP groups. In the BO group, the score of Stroop-D is lower, and the MoCA sub-items are higher than the IP group. There is no correlation between CTP parameters and cognitive function. CONCLUSION: Cognitive function is significantly impaired in patients with ICAS, which is related to cerebral perfusion. Executive, memory, and language function were better preserved in ICAS patients without IP. Hence, this study posits that managing hypoperfusion induced by ICAS may play a pivotal role in the development of VCI.

Modulation of cutaneous vasodilation by reactive oxygen species during local and whole-body heating in young and older adults.

OBJECTIVE: To evaluate reactive oxygen species (ROS) modulation of cutaneous vasodilation during local and whole-body passive heating in young and older adults. METHODS: Cutaneous vascular conductance normalized to maximum vasodilation (%CVCmax) was assessed in young and older adults (10 per group) using laser-Doppler flowmetry at 4 dorsal forearm sites treated with 1) Ringer's solution (control), 2) 100 µM apocynin (NADPH oxidase inhibitor), 3) 10 µM allopurinol (xanthine oxidase inhibitor), or 4) 10 µM tempol (superoxide dismutase mimetic), via intradermal microdialysis during local (protocol-1) and whole-body heating (protocol-2). Protocol-1: forearm skin sites were set at 33°C during baseline and then progressively increased to 39°C and 42°C (30 min each). Protocol-2: participants were immersed in warm water (35°C, mid-sternum) with the experimental forearm above water level and local skin sites maintained at 34°C. Bath temperature was increased (~40°C) to clamp core temperature at 38.5°C for 60 min. RESULTS: Protocol-1: there were significant treatment site by age interactions for the 39°C (P=0.015) and 42°C (P=0.004) plateaus. Although, no significant effects were observed after post-hoc adjustment. Protocol-2: there was a significant treatment site by age interaction (P<0.001) whereby %CVCmax in older adults was 11.0% [7.4,14.6] higher for apocynin (P<0.001), 8.9% [5.3,12.5] higher for allopurinol (P<0.001) and 4.8% [1.3,8.4] higher for tempol (P=0.016) sites relative to the control site. CONCLUSION: ROS derived from NADPH oxidase and xanthine oxidase attenuate cutaneous vasodilation in older adults during passive whole-body heating, but not during local skin heating, with negligible effects on their young counterparts for either heating modality.

Analysis of cell cycle stage, replicated DNA, and chromatin-associated proteins using high-throughput flow cytometry.

Flow cytometry is a versatile tool used for cell sorting, DNA content imaging, and determining various cellular characteristics. With the possibility of high-throughput analyses, it combines convenient labelling techniques to serve rapid, quantitative, and qualitative workflows. The ease of sample preparation and the broad range of applications render flow cytometry a preferred approach for many scientific questions. Yet, we lack practical adaptations to fully harness the quantitative and high-throughput capabilities of most cytometers for many organisms. Here, we present simple and advanced protocols for the analysis of total DNA content, de novo DNA synthesis, and protein association to chromatin in budding yeast and human cells. Upon optimization of experimental conditions and choice of fluorescent dyes, up to four parameters can be measured simultaneously and quantitatively for each cell of a population in a multi-well plate format. Reducing sample numbers, plastic waste, costs per well, and hands-on time without compromising signal quality or single-cell accuracy are the main advantages of the presented protocols. In proof-of-principle experiments, we show that DNA content increase in S-phase correlates with de novo DNA synthesis and can be predicted by the presence of the replicative helicase MCM2-7 on genomic DNA.

Hue-change coupled with CRISPR-Cas12a lateral flow assay for the semiquantitative detection of Salmo salar adulteration.

Salmo salar is one of the most popular salmon species due to its meaty texture and quality protein. Oncorhynchus mykiss, which has a muscle texture similar to that of Salmo salar and is less expensive, is often used as a substitute for Salmo salar. As Salmo salar and Oncorhynchus mykiss belong to the same subfamily of Salmonidae, traditional methods are ineffective in the specific detection of the two. In this study, we combined hue-change with CRISPR/Cas12a lateral flow assay to detect the Salmo salar adulteration. This method detected S. salar genomic DNA at a vLOD of 5 copies, and was able to accurately identify adulterated samples containing 5 % w/w Salmo salar within one hour. In addition, the detection of Salmo salar in processed food products was achieved with the naked-eye at a concentration range of 0 % âˆ¼ 70 % w/w, and the detection accuracy is between 93.3 % âˆ¼ 100 %.

Doing good well (Karma Yoga, the path of selfless action): Psychotherapeutic lessons from the East.

The tripartite classification of mental faculties into cognition, affect, and conation (motivation and action) continues to be the edifice on which the mind and the methods to address mental afflictions are studied. Eastern spiritual traditions offer insights into mental health as it pertains to each of these domains. Following up on our previous paper on the cognition path to psychotherapy (Knowing oneself, or Jnana Yoga), we herein focus on the path of selfless action (Karma Yoga). We review eastern concepts on the nature of karma and the approaches to optimal action (the will to do things, doing the right things, and doing them well). We then place these eastern insights in the context of emerging concepts in psychology on motivation and action. Current psychological concepts such as autonomy and intrinsic motivation, mastery, flow and growth mindset, higher purpose and value driven self-less action, equanimity and balance are convergent with ancient eastern concepts. We also review current neuroscientific underpinnings (such as neural circuitries, neurotransmitter systems and epigenetics and how these facilitate neural plasticity) relevant to karma, including free will, focused action, prosocial behaviors, extrinsic and intrinsic and motivation. These concepts have significant implications for psychotherapeutic models, especially in the areas of positive psychology and preventive psychiatry.

Orchestrating Blood Flow in the Retina: Interpericyte Tunnelling Nanotube Communication.

The retina transforms light into electrical signals, which are sent to the brain via the optic nerve to form our visual perception. This complex signal processing is performed by the retinal neuron and requires a significant amount of energy. Since neurons are unable to store energy, they must obtain glucose and oxygen from the bloodstream to produce energy to match metabolic needs. This process is called neurovascular coupling (NVC), and it is based on a precise mechanism that is not totally understood. The discovery of fine tubular processes termed tunnelling nanotubes (TNTs) set a new type of cell-to-cell communication. TNTs are extensions of the cellular membrane that allow the transfer of material between connected cells. Recently, they have been reported in the brain and retina of living mice, where they connect pericytes, which are vascular mural cells that regulate vessel diameter. Accordingly, these TNTs were termed interpericyte tunnelling nanotubes (IPTNTs), which showed a vital role in blood delivery and NVC. In this chapter, we review the involvement of TNTs in NVC and discuss their implications in retinal neurodegeneration.

Effects of hand-bathing on noise-induced vasoconstriction: A randomized controlled trial.

AIM: This study assessed the effects of hand-bathing on sympathetic nervous activity exacerbated by psychological stress. Participants immersed one hand in warm water for 2 min while exposed to noise, and changes in blood flow and skin temperature of the non-immersed hand were observed. METHODS: Twenty-nine healthy university students aged 20 years or older were randomly assigned to either the hand-bathing group (n = 14) or the control group (n = 15). After a brief rest in a quiet environment, participants were exposed to noise for 6 min. Those in the hand-bathing group submerged their left hand in a 40°C thermostatic bath for 2 min, starting 2 min into the noise exposure. The tympanic temperature, blood flow, and skin temperature of the non-immersed hand were continuously measured, along with blood pressure and subjective evaluations before and after the noise exposure. RESULTS: Both groups experienced a decrease in fingertip skin temperature at the start of the noise exposure, persisting longer in the control group. Conversely, the hand-bathing group showed increased fingertip skin temperature after 150 s, significantly higher after the noise exposure than the control group (p = .04). Participants in the hand-bathing group reported significantly increased overall body warmth, thermal comfort, and relaxation during hand-bathing (p = .007, p = .01, p < .001). CONCLUSIONS: The 2-min hand-bathing intervention reversed the pronounced vasoconstrictive response induced by noise exposure and elicited heightened sensations of overall body warmth, thermal comfort, and relaxation. Hand-bathing may mitigate heightened sympathetic nervous activity associated with psychological stress induced by noise exposure.