Phase Marker Research Articles

Ultrafast spectroscopy of coherent phonons across the pressure driven insulator to metal phase transition in V2O3

Nowadays, materials science is moving towards the understanding and control of materials in nonequilibrium states by making use of perturbative techniques to investigate their dynamical responses. From this perspective, the use of ultrashort light pulses seems to be a relevant approach as it can selectively address different degrees of freedom in solid-state systems and more particularly electrons. Such a method can help to decipher the physical phenomena arising from electronic correlations and complements a more conventional methodology where the phase diagrams of materials are investigated at thermodynamical equilibrium. Here, we combine femtosecond optical spectroscopy and a high-pressure setup to monitor the ultrafast out-of-equilibrium photo response of a V2O3 thin film across the pressure driven insulator-to-metal transition. The experimental results demonstrate the possibility to use the spectroscopy of coherent phonons as a thermodynamical phase marker in V2O3 thin films. In addition, the frequency behavior of the ultrafast coherent phonon mode (A1g character) seems to reflect the manifestation of a strong coupling between the lattice and electronic degrees of freedom near first-order transition lines with a pronounced drop in frequency around the critical pressure. Published by the American Physical Society 2024

Glutathione accelerates the cell cycle and cellular reprogramming in plant regeneration.

The plasticity of plant cells underlies their wide capacity to regenerate, with increasing evidence in plants and animals implicating cell cycle dynamics in cellular reprogramming. To investigate the cell cycle during cellular reprogramming, we developed a comprehensive set of cell cycle phase markers in the Arabidopsis root. Using single-cell RNA-seq profiles and live imaging during regeneration, we found that a subset of cells near an ablation injury dramatically increases division rate by truncating G1. Cells in G1 undergo a transient nuclear peak of glutathione (GSH) prior to coordinated entry into S phase followed by rapid divisions and cellular reprogramming. A symplastic block of the ground tissue impairs regeneration, which is rescued by exogenous GSH. We propose a model in which GSH from the outer tissues is released upon injury licensing an exit from G1 near the wound to induce rapid cell division and reprogramming.

Evaluation of Inflammation and Platelet Apoptosis Parameters in Obese Patients in Various Types of Anticoagulant Prophylaxis of Venous Thromboembolic Complications in Context of COVID-19

INTRODUCTION: The physical inactivity, hypoventilation, as well as chronic inflammation in obese patients aggravates their condition in various diseases. These features have become important with the advent of the COVID-19 pandemic, in which inflammation and platelet-activated coagulopathy are closely linked. AIM: To study laboratory parameters of inflammation and platelet apoptosis in obese patients using various types of anticoagulant prophylaxis of venous thromboembolic complications with the underlying COVID-19. MATERIALS AND METHODS: The study included 370 patients with COVID-19. Depending on the presence or absence of obesity and the type of parenteral anticoagulant, patients in our study were divided into groups: group 1 — non-obese + low molecular weight heparin (LMWH) (n = 114), group 2 — non-obese + unfractionated heparin (UFH) (n = 58), group 3 — obesity + LMWH (n = 76), group 4 — obesity + UFH (n = 66). The incidence of venous thromboembolic complications (VTEC), bleeding, general markers of the acute phase of inflammation, and specific markers of platelet apoptosis (phosphatidylserine and calreticulin) have been analyzed. RESULTS: At the end of hospital treatment, a decrease in ferritin levels was noted in patients both with and without obesity receiving LMWH. The concentration of calreticulin was higher in patients taking LMWH (groups 1 and 3). Phosphatidylserine levels were high in patients receiving LMWH only if they were obese. In patients taking UFH compared to LMWH, a high incidence of pulmonary embolism (PE) without a source (13.6% of cases versus 2.6%, respectively, p = 0.029) and PE with a source in the lower extremities (9.1% of cases versus 0%, respectively, p = 0.018) was found. When using LMWH, a lower incidence of bleeding was observed compared to using UFH (5.3% of cases versus 16.7%, respectively, p = 0.056). CONCLUSION: The levels of phosphatidylserine and calreticulin are higher in obese patients receiving LMWH. At the same time, patients in this group have a low incidence of VTEC and hemorrhagic complications compared to the group of patients taking UFH.

Liver disease in primary antibody deficiencies.

Liver disease has emerged as a major risk factor for increased mortality in patients with common variable immunodeficiency (CVID). This is mostly due to presinusoidal portal hypertension (PHTN) frequently secondary to nodular regenerative hyperplasia (NRH). Its pathogenesis is still poorly understood and treatment strategies for its various stages are often guided by trial and error. This review summarizes the most recent findings in the light of previous literature. In the last 2 years, different groups have addressed pathology, diagnostics, treatment, and liver transplantation. Histological examinations seem to support the pathogenetic sequence of T-cell mediated infiltration and damage of the sinusoidal space with secondary development of NRH, pericellular fibrosis, and the manifestation of PHTN. While markers of the early phase - beyond slight elevation of cholestatic enzymes - are still missing, elevated liver stiffness and splenomegaly above 16 cm longitudinal diameter have been suggested as warning signs for PHTN in CVID patients. Data on immunosuppressive treatment of this manifestation is still very heterogeneous, but a recent report on liver transplantation was encouraging for end stage liver disease. Liver disease deserves higher attention in the management of CVID. More studies are needed to understand its pathogenesis and optimal treatment.

Bleeding us dry: The financial impact of full blood examinations in the immediate postoperative period.

Full blood examinations, often referred to as FBE, are commonly ordered postoperatively, despite limited utility in many of its markers in the acute phase. It is estimated that in the 2022-2023 financial year, the Australian healthcare system billed over $13 million for full blood examinations (FBEs) to Medicare. This study aims to assess the cost of using FBE following surgery. We explore potential cost savings by using a haemoglobin examination (HE) in replace of FBE, with both tests run on identical machines, producing the same result, but at a fraction of the cost. A retrospective analysis was conducted at a single institution, including all patients who underwent minimally invasive laparoscopic pelvic surgeries between 1/7/2017 and 30/6/2019. Patient records were examined to identify patient demographics, postoperative pathology tests and interventions. Patients who received FBE in the first 24 h following surgery were identified and included in the study. Using national surgery and admission statistics, a potential cost-saving analysis will be performed. Among 519 men who underwent robotic-assisted pelvic surgery, 325 patients had routine postoperative investigations, with 323 receiving FBE and 2 receiving HE. Abnormal results were found in the majority of patients that underwent FBE. Eight patients received packed red blood cell transfusion, none of these meeting the hospital-specific criteria for transfusion protocol. Twelve patients received antibiotics, none were in response to abnormal FBE, with all patients experiencing a fever, given prophylactically or in the days following the surgery. FBE and HE are both listed on the Medicare Benefits Scheme at $16.95 and $7.85, respectively, the difference being $9.10. Extrapolating the existing data, within the first 24 h following surgery, the estimated savings were $8818, with savings increasing accumulatively with longer observation intervals following surgery. When similar savings are applied to national figures, the potential savings to the Australian Public Healthcare system likely exceeds millions. Our study revealed that over half of the patients who underwent a RARP received FBE within the first 24 h postoperatively, the vast majority of which exhibited abnormal results that were not acted upon. These findings substantiate the limited utility of FBE in the postoperative setting. Cell markers observed in FBE are predominantly subjective, but consensus exists regarding the importance of evaluating haemoglobin levels postoperatively. Considering that one in four hospital admissions involves surgical procedures, and a $9.10 price differential between FBE and HE, the potential annual economic impact of utilising routine FBEs for assessing haemoglobin levels immediately after surgery is likely to reach millions. Although having obvious flaws, these results underscore the potential accumulative cost arising from everyday clinical judgement and the importance of thoughtful consideration when ordering pathology. The routine ordering of FBE postoperatively, without properly considering its indication, incurs significant costs. This study highlights the potential cost savings by HE instead, emphasising the need for revaluation and appropriate utilisation of pathology tests in the immediate postoperative period given the physiological acute phase response expected postoperatively.

A novel mouse model of hepatocyte-specific apoptosis-induced myeloid cell-dominant sterile liver injury and repair response.

Apoptosis, inflammation, and wound healing are critical pathophysiological events associated with various liver diseases. Currently, there is a lack of in vivo approaches to study hepatocyte apoptosis-induced liver injury and repair. To address this critical knowledge gap, we developed a unique genetically modified mouse model, namely, 3-Transgene (Tg) with inducible Hepatocyte-Specific Apoptosis Phenotype (3xTg-iHAP) in this study. The 3xTg-iHAP mice possess three transgenes including Alb-Cre, Rosa26-rtTA, and tetO-Fasl on a B6 background. These mice are phenotypically normal, viable, and fertile. After subcutaneous administration of a single dose of doxycycline (5 mg/kg, Dox) to 3xTg-iHAP mice, we observed a complete histological spectrum of sterile liver wound-healing responses: asymptomatic hepatocyte apoptosis at 8 h, necrotic liver injury and sterile inflammation at 48 h, followed by hepatocyte mitosis and regeneration within 7 days. During the injury phase, the mice exhibited an increase in the biomarkers of alanine aminotransferase (ALT), chemokine (C-X-C motif) ligand 1 (CXCL1), and IL-6 in peripheral blood, as well as α-smooth muscle actin (α-SMA) protein in liver tissues. Conversely, the mice displayed a decrease in these markers in the recovery phase. Remarkably, this model shows that the sterile liver injury following elevated hepatocyte apoptosis is associated with an increase in myeloid cells in the liver. Within 7 days post-Dox administration, the liver of Dox-treated 3xTg-iHAP mice displays a normal histological structure, indicating the completion of wound healing. Together, we established a novel mouse model of injury and regeneration induced by hepatocyte apoptosis. This tool provides a robust in vivo platform for studying the pathophysiology of sterile liver inflammation, regeneration, and new therapeutic interventions for liver diseases.NEW & NOTEWORTHY Bu et al. present a triple-transgenic mouse model, namely, 3xTg-iHAP mice that are engineered to explore hepatocyte apoptosis-triggered sterile liver injury and regeneration. This model demonstrates a full spectrum of liver wound-healing responses from asymptomatic apoptosis to injury, myeloid cell-dominant sterile inflammation, and repair after induction of hepatocyte-specific apoptosis. The robust nature of this model makes it an invaluable in vivo tool for studying sterile liver inflammation, regeneration, and new therapeutic strategies.

A Protocol to Determine Circadian Phase by At-Home Salivary Dim Light Melatonin Onset Assessment.

Internal circadian phase assessment is increasingly acknowledged as a critical clinical tool for the diagnosis, monitoring, and treatment of circadian rhythm sleep-wake disorders and for investigating circadian timing in other medical disorders. The widespread use of in-laboratory circadian phase assessments in routine practice has been limited, most likely because circadian phase assessment is not required by formal diagnostic nosologies, and is not generally covered by insurance. At-home assessment of salivary dim light melatonin onset (DLMO, a validated circadian phase marker) is an increasingly accepted approach to assess circadian phase. This approach may help meet the increased demand for assessments and has the advantages of lower cost and greater patient convenience. We reviewed the literature describing at-home salivary DLMO assessment methods and identified factors deemed to be important to successful implementation. Here, we provide specific protocol recommendations for conducting at-home salivary DLMO assessments to facilitate a standardized approach for clinical and research purposes. Key factors include control of lighting, sampling rate, and timing, and measures of patient compliance. We include findings from implementation of an optimization algorithm to determine the most efficient number and timing of samples in patients with Delayed Sleep-Wake Phase Disorder. We also provide recommendations for assay methods and interpretation. Providing definitive criteria for each factor, along with detailed instructions for protocol implementation, will enable more widespread adoption of at-home circadian phase assessments as a standardized clinical diagnostic, monitoring, and treatment tool.

Dysregulation of circadian clock gene expression patterns in a treatment-resistant animal model of depression.

Circadian rhythm (CR) disturbances are among the most commonly observed symptoms during major depressive disorder, mostly in the form of disrupted sleeping patterns. However, several other measurable parameters, such as plasma hormone rhythms and differential expression of circadian clock genes (ccgs), are also present, often referred to as circadian phase markers. In the recent years, CR disturbances have been recognized as an essential aspect of depression; however, most of the known animal models of depression have yet to be evaluated for their eligibility to model CR disturbances. In this study, we investigate the potential of adrenocorticotropic hormone (ACTH)-treated animals as a disease model for research in CR disturbances in treatment-resistant depression. For this purpose, we evaluate the changes in several circadian phase markers, including plasma concentrations of corticosterone, ACTH, and melatonin, as well as gene expression patterns of 13 selected ccgs at 3 different time points, in both peripheral and central tissues. We observed no impact on plasma corticosterone and melatonin concentrations in the ACTH rats compared to vehicle. However, the expression pattern of several ccgs was affected in the ACTH rats compared to vehicle. In the hippocampus, 10 ccgs were affected by ACTH treatment, whereas in the adrenal glands, 5 ccgs were affected and in the prefrontal cortex, hypothalamus and liver 4 ccgs were regulated. In the blood, only 1 gene was affected. Individual tissues showed changes in different ccgs, but the expression of Bmal1, Per1, and Per2 were most generally affected. Collectively, the results presented here indicate that the ACTH animal model displays dysregulation of a number of phase markers suggesting the model may be appropriate for future studies into CR disturbances.

Simultaneous plasma and interstitial profiles of hormones and metabolites using URHYTHM: a novel ambulatory collection device

Increasing the sampling resolution when examining plasma hormone and metabolite profiles will more accurately describe diurnal/circadian rhythms and expose previously undiscovered ultradian rhythms that underpin metabolic physiology(1). In human research studies this must be tempered by the practical, ethical and safety concerns of repeated cannulation/blood draws. Here we examine the correlation between hormone and metabolite profiles present in interstitial fluid microdialysate(2), and plasma to help solve this dilemma.Hormonal markers of circadian phase (melatonin, cortisol) and metabolites will show similar profiles in plasma and interstitial fluid.Compare time-series hormone and metabolite profiles collected in interstitial fluid using a novel ambulatory microdialysis collection device (U-RHYTHM) with simultaneously drawn plasma samples.All study protocols were reviewed by Health Sciences Faculty Research Ethics Committee, University of Bristol. Fasted healthy male volunteers aged 18-35 (n=3) were fed a standardised breakfast (08:00), lunch (13:00), dinner (19:00) and snack (22:00) (2225kCal [83g protein, 273g carbohydrate, 83g fat, 27g fibre]). Participants remained on the study bed, lights off/sleep occurred between 23:00-07:00 (<4 lux).A 20kDa cutoff 30 mm linear microdialysis membrane was placed in periumbilical subcutaneous tissue, perpendicular to the midline. The membrane was perfused at 1µl/min and the microdialysate collected every 20 minutes for 25 hours into discrete samples using a portable fraction collector (URHYTHM) worn around the waist. Time-matched blood samples were obtained from an antecubital fossa canula.All samples were stored at -80°C prior to analysis. Hormones and metabolites were measured via targeted UPLC-MS/MS metabolomics, for the metabolites the AbsoluteIDQ® p180 kit (Biocrates Life Sciences AG)) was used with 10µL of plasma(3) and 15µL of microdialysate.Simultaneous time-courses of n = 24 metabolites were detected in plasma and interstitial microdialysate: alanine, arginine, asparagine, citrulline, glutamine, glutamate, glycine, histidine, isoleucine, leucine, lysine, methionine, ornithine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, valine, trans-4-Hydroxyproline, taurine, carnitine and acetylcarnitine.Spearman rank cross correlation identified the time shift that was required to reach highest correlation between the plasma and microdialysate profiles. Excluding taurine, microdialysate profiles were delayed by 32.3 minutes (range 0-130) and 12.9 minutes (range 0-50) for participant 1 and 2, respectively (p <0.05), only 3/20 metabolite profiles were correlated for participant 3. Trans4-hydroxyproline had the strongest correlation (rs 0.890, 0.814 and 0.808), with only participant 3 displaying a time shift (50 minutes). Taurine had the weakest correlation in all 3 participants. (rs 0.096, 0.182, 0.325).These data demonstrate that our novel approach can be used to comprehensively detect hormone and metabolite profiles in interstitial fluid. Thus, the U-RHYTHM would serve as a useful tool, not only to increase sampling resolution in controlled laboratory studies but would prove particularly useful for examining circadian/ultradian profiles for chronobiology and nutrition studies in free-living individuals.

Utility of methylated DNA markers for the diagnosis of malignant biliary strictures.

Early identification of malignant biliary strictures (MBS) is challenging, with up to 20% classified as indeterminants after preliminary testing and tissue sampling with endoscopic retrograde cholangiopancreatography (ERCP). We aimed to evaluate the use of methylated DNA markers (MDM) from biliary brushings to enhance MBS detection in a prospective cohort. Candidate MDMs were evaluated for their utility in MBS diagnosis through a series of discovery and validation phases. DNA was extracted from biliary brushing samples, quantified, bisulfite-converted, and then subjected to methylation-specific Droplet Digital Polymerase Chain Reaction (ddPCR). Patients were considered to have no malignancy if the sampling was negative and there was no evidence of malignancy after 1 year or definitive negative surgical histopathology. Fourteen candidate MDMs were evaluated in the discovery phase, with top-performing and new markers evaluated in the technical validation phase. The top four MDMs were TWIST1, HOXA1, VSTM2B, and CLEC11A, which individually achieved AUC values of 0.82, 0.81, 0.83, and 0.78, respectively, with sensitivities of 59.4%, 53.1%, 62.5%, and 50.0%, respectively, at high specificities for malignancy of 95.2-95.3% for the final biologic validation phase. When combined as a panel, the AUC was 0.86, achieving 73.4% sensitivity and 92.9% specificity, which outperformed cytology and fluorescent in situ hybridization (FISH). The selected methylated DNA markers demonstrated improved performance characteristics for the detection of MBS compared to cytology and FISH. ​ Therefore, MDMs should be considered viable candidates for inclusion in diagnostic testing algorithms.​.

Saliva stress biomarkers in ERCP trainees before and after familiarisation with ERCP on a virtual simulator.

Stress during the early ERCP learning curve may interfere with acquisition of skills during training. The purpose of this study was to compare stress biomarkers in the saliva of trainees before and after familiarisation with ERCP exercises on a virtual simulator. Altogether 26 endoscopists under training, 14 women and 12 men, completed the three phases of this study: Phase 1. Three different ERCP procedures were performed on the simulator. Saliva for α-amylase (sAA), Chromogranin A (sCgA), and Cortisol (sC) were collected before (baseline), halfway through the exercise (ex.), and 10 min after completion of the exercise (comp.); Phase 2. A three-week familiarisation period where at least 30 different cases were performed on the virtual ERCP simulator; and Phase 3. Identical to Phase 1 where saliva samples were once again collected at baseline, during, and after the exercise. Percentage differences in biomarker levels between baseline and exercise (Diffex) and between baseline and completion (Diffcomp) during Phase 1 and Phase 3 were calculated for each stress marker. Mean % changes, Diffex and Diffcomp, were significantly positive (p < 0.05) for all markers in both Phase 1 and Phase 3. Diffex in Phase 1 was significantly greater than Diffex in Phase 3 (p < 0.05) for sAA and sCgA. Diffcomp for sAA in Phase 1 was significantly greater than Diffcomp in Phase 3 (p < 0.05). No significant differences were found in sC concentration between Phases 1 and 3. This study shows that familiarisation with the ERCP simulator greatly reduced stress as measured by the three saliva stress biomarkers used with sAA being the best. It also suggests that familiarisation with an ERCP simulator might reduce stress in the clinical setting.

High Resolution Reconstruction of the Proximal Tubule Apical Endocytic Pathway

Kidney proximal tubule (PT) cells are specialized for effcient apical uptake of albumin and other proteins that escape the glomerular filtration barrier. The apical endocytic pathway is uniquely organized to maintain highly dynamic retrieval of these proteins via binding to the multiligand receptors megalin and cubilin. Ultrastructural studies following the fate of internalized markers have provided a temporal model for the delivery of receptor-bound ligands and fluid phase markers to lysosomes. In this model, internalization from the apical membrane occurs via budding of irregular clathrin-coated invaginations that form at the base of microvilli. After uncoating, budded vesicles fuse with apical early endosomes (AEEs) where acidification triggers dissociation of ligands from their receptors. AEEs further mature and expand into apical vacuoles (AVs) that ultimately deliver ligands and fluid to lysosomes. Receptor recycling to the apical membrane is thought to occur via dense apical tubules (DATs) that have a uniform diameter and membrane thickness and which appear to bud from both AEEs and AVs. We used Zeiss Arivis Pro technology to annotate and reconstruct apical membrane invaginations and endocytic compartments in a 3D volume of mouse kidney with isotropic voxels of 4 nm, acquired using focused ion-beam scanning electron microscopy (FIB-SEM). Strikingly, 3D reconstruction of DATs revealed a large and highly interconnected network of tubules in the subapical region of PT cells. Connections were observed between the DAT network and endosomal compartments, as well as between DATs and apical membrane invaginations. Our data suggest the possibility that DATs exist as a stable compartment that interacts transiently with the apical membrane to enable receptor recycling. Current studies are underway to better understand how PT cells modulate endocytic entry and recycling via the same portals. National Institute of Health: 5T32GM133353-03, R01 DK118726, R01 DK125049, U54 DK137329. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

RAD51 as an immunohistochemistry-based marker of poly(ADP-ribose) polymerase inhibitor resistance in ovarian cancer.

Effective functional biomarkers that can be readily used in clinical practice to predict poly(ADP-ribose) polymerase inhibitor (PARPi) sensitivity are lacking. With the widespread adoption of PARPi maintenance therapy in ovarian cancer, particularly in patients with BRCA mutation or HR deficiencies, accurately identifying de novo or acquired resistance to PARPi has become critical in clinical practice. We investigated RAD51 immunohistochemistry (IHC) as a functional biomarker for predicting PARPi sensitivity in ovarian cancer. Ovarian cancer patients who had received PARPi and had archival tissue samples prior to PARPi exposure ("pre-PARPi") and/or after progression on PARPi ("post-PARPi") were selected. RAD51 IHC expression was semi-quantitatively evaluated using the H-score in geminin (a G2/S phase marker)- and γH2AX (a DNA damage marker)-positive tissues. A RAD51 H-score of 20 was used as the cutoff value. In total, 72 samples from 56 patients were analyzed. The median RAD51 H-score was 20 (range: 0-90) overall, 10 (0-190) in pre-PARPi samples (n = 34), and 25 (1-170) in post-PARPi samples (n = 19). Among patients with BRCA mutations, RAD51-low patients had better progression-free survival (PFS) after PARPi treatment than RAD51-high patients (P = 0.029). No difference was found in PFS with respect to the genomic scar score (P = 0.930). Analysis of matched pre- and post-PARPi samples collected from 15 patients indicated an increase in the RAD51 H-score upon progression on PARPi, particularly among pre-PARPi low-RAD51-expressing patients. RAD51 is a potential functional IHC biomarker of de novo and acquired PARPi resistance in BRCA-mutated ovarian cancer and can be used to fine-tune ovarian cancer treatment.

P143 High throughput multiplex assay identifies clinically meaningful clusters in systemic sclerosis patients with low cardiovascular risk and no history of heart disease

Abstract Background/Aims People with systemic sclerosis (SSc) often show subclinical evidence of primary heart involvement (pHI), which when clinically evident, is a prognostic factor of poor outcome. We aimed to profile serum proteins in patients with SSc and no history of pHI and identify clinically meaningful clusters to aid early identification and stratification for pHI. Methods Serum from 78 patients from CONVAS (‘CONnective Tissue Disease and VASculitis Cohort’) and ELCASA (‘ELectrophysiology and CArdiac imaging in SclerodermA’) cohorts [no history of pHI, pulmonary hypertension, diabetes or &amp;gt; 2 traditional cardiovascular (CV) risk factors] was used to measure 355 proteins across inflammation, cardiometabolic and cardiovascular II/III Olink panels. Unsupervised hierarchical clustering using scaled normalised protein expression (NPX) was used to define clusters. Demographic and clinical characteristics were compared between clusters and significant differences reported using Kruskal-Wallis rank sum test for continuous or Pearson’s chi-squared test for categorical variables, at the 5% significance threshold. Results Clinical characteristics of the patient cohorts have been previously published (also summarised in table 1,column 2). Unsupervised hierarchical clustering revealed three patient clusters - C1 [27/78 (35%)], C2 [33/78 (42%)] and C3 [18/78 (23%)] with intermediate, low and high protein expression respectively. C2 patients were younger (median age 51 years) compared to C1 (61 years) and C3 (58 years) (p &amp;lt; 0.001). High expression cluster C3 had a significantly higher proportion of males [7/18 (39%)](p = 0.009), shorter disease duration [median 1.15 years, IQR (0.66 to 7.23)], higher modified Rodnan skin scores (MRSS) [median 5.50, IQR (3.25 to 11)] (p = 0.002) and current disease modifying anti-rheumatic drugs (DMARD) use [16/18 (89%)](p = 0.005). NTproBNP and CRP were elevated in C1 and C3, and high-sensitivity troponin I (hsTnI) was highest in C3[median 6.15 IQR (5.08 to 89.58)](p = 0.002); n = 3 within C1 showed especially high NTproBNP (median 227.50pg/ml) and CRP (median 13mg/dl). No differences in antibody profiles were noted across the three clusters (table 1). Conclusion Differences in serum protein expression identifies clinically meaningful clusters of SSc patients with elevated acute phase and serum cardiac markers despite low CV risk profile. Future work should assess their utility as prognostic biomarkers of SSc-pHI. Disclosure R. Shukla: None. R. Dumitru: Grants/research support; ACORN charity and Charitable Foundation Fellowship, Leeds Teaching Hospital. B. Erhayiem: None. G. Fent: None. F. Del Galdo: Consultancies; Abbvie, AstraZeneca, Boehringer-Ingelheim, Capella, Chemomab, GSK, Janssen, Mitsubishi-Tanabe. S. Plein: Other; funded by a British Heart Foundation Personal Chair. D. Plant: None. M.H. Buch: Consultancies; AbbVie, Arxx Therapeutics, Boehringer Ingelheim, Galapagos, Gilead, and Pfizer (all paid to host institution).

Astaxanthin suppresses the malignant behaviors of nasopharyngeal carcinoma cells by blocking PI3K/AKT and NF-κB pathways via miR-29a-3p

BackgroundAs a natural extraction, astaxanthin is gaining increasing attention because of its safety and anti-tumor properties. It has been reported to participate in the progression of various types of cancer such as gastric cancer and ovarian cancer. Nevertheless, the role of astaxanthin in nasopharyngeal carcinoma (NPC) has not been investigated.ObjectThe study aimed to explore the anticancer mechanism of astaxanthin in regulating NPC cell proliferation, cell cycle progression, apoptosis, migration, and invasion.MethodsHuman NPC cells (C666-1) were treated with different concentrations of astaxanthin (0, 1, 10, 20 mg/mL) followed by detection of cell viability. Then, C666-1 cell proliferation, apoptosis, cell cycle progression, invasion, and migration in response to 10 mg/mL astaxanthin, LY294002 (PI3K/AKT inhibitor) or parthenolide (PTL; NF-κB inhibitor) treatment were measured using cell counting kit-8 assay, colony forming assay, flow cytometry analyses, Transwell assay, and wound healing assay, respectively. Western blotting was performed to quantify protein levels of factors involved in PI3K/AKT and NF-κB signaling pathways, cell cycle phase markers (Cyclin D1, p21) and apoptotic markers (Bcl-2 and Bax).ResultsC666-1 cell proliferation, invasion, and migration were significantly suppressed by astaxanthin while cell apoptosis and cell cycle arrest at G1 phase were effectively enhanced in the context of 10 mg/mL astaxanthin. Protein levels of p-AKT, p-P65 and p-IκB levels were suppressed by astaxanthin treatment. After LY294002 or PTL treatment, the suppressive impact of astaxanthin on C666-1 cell process was strengthened, accompanied by the more obvious decrease in cell activity and cell colony number, more enhanced cell apoptosis and G1 phase arrest, and further inhibited cell migration and invasion. Moreover, the inhibitory effect of astaxanthin on Cyclin D1 and Bcl-2 protein levels as well as the promoting impact of astaxanthin on p21 and Bax were also amplified in combination with LY294002 or PTL treatment.ConclusionsAstaxanthin significantly suppresses NPC cell proliferation, cell cycle arrest, migration, invasion while promoting cell apoptosis by inactivating PI3K/AKT and NF-κB pathways. The study first reveals the anticancer role of astaxanthin in NPC, providing a potential candidate for NPC treatment.

Validation of dot blot immunoassay for measurement of complement opsonization of nanoparticles

Complement plays a critical role in the immune response toward nanomaterials. The complement attack on a foreign surface results in the deposition of C3, assembly of C3 convertases, the release of anaphylatoxins C3a and C5a, and finally, the formation of membrane attack complex C5b-9. Various technologies can measure complement activation markers in the fluid phase, but measurements of surface C3 deposition are less common. Previously, we developed an ultracentrifugation-based dot blot immunoassay (DBI) to measure the deposition of C3 and other protein corona components on nanoparticles. Here, we validate the repeatability of the DBI and its correlation with pathway-specific and common fluid phase markers. Moreover, we discuss the advantages of DBI, such as cost-effectiveness and versatility, while addressing potential limitations. This study provides insights into complement activation at the nanosurface level, offering a valuable tool for nanomedicine researchers in the field.

Proteomic characteristics of the treatment trajectory of patients with COVID-19.

The ongoing COVID-19 pandemic caused by SARS-CoV-2 has prompted global concern due to its profound impact on public health and the economy. Effective treatment of COVID-19 patients in the acute phase or of those with long COVID is a major challenge. Using data-independent acquisition (DIA) technology, we performed proteomic profiling on plasma samples from 22 COVID-19 patients and six healthy controls at Dazhou Central Hospital. Randomforest and least absolute shrinkage and selection operator algorithms were used for analysis at various COVID-19 treatment stages. We identified 79 proteins that were differentially expressed between COVID-19 patients and healthy controls, mainly involving pathways associated with cell processes and binding. Across different treatment stages of COVID-19, five proteins-PI16, GPLD1, IGFBP3, KRT19, and VCAM1-were identified as potential molecular markers for dynamic disease monitoring. Furthermore, the proteins BTD, APOM, IGKV2-28, VWF, C4BPA, and C7 were identified as candidate biomarkers for distinguishing between SARS-CoV-2 positivity and negativity. Analysis of protein change profiles between the follow-up and healthy control groups highlighted cardiovascular changes as a concern for patients recovering from COVID-19. Our study revealed the infection profiles of SARS-CoV-2 at the protein expression level comparing different phases of COVID-19. DIA mass spectrometry analysis of plasma samples from COVID-19 patients undergoing treatment identified key proteins involved in signaling pathways that might be used as markers of the recovery phase. These findings provide insight for the development of therapy options and suggest potential blood biomarkers for COVID-19.

In-situ investigation of supercooling behaviour during high-pressure shift freezing of pure water and sucrose solution

Supercooling is a main controllable factor for the fundamental understanding the high-pressure shift freezing (HPSF). In the study, a self-developed device based on the diamond anvil cell (DAC) and confocal Raman microscopy was utilized to realize an in-situ investigation of supercooling behaviour during HPSF of the pure water and sucrose solution. The spectra were used to determine the freezing point which is shown as a spectral phase marker (SD). The hydrogen bond strengths of water and sucrose solution under supercooling states were estimated by peak position and peak area ratio of sub-peaks. The results showed that the OH stretching bands had redshift under supercooling states. Moreover, the addition of sucrose molecules could strengthen the hydrogen bonding strength of water molecules under supercooling states. Thus, the DAC combined with Raman spectroscopy could be considered a novel strategy for a deep understanding of the supercooling behaviour during HPSF.

Assessment of Factor VIII Activity and D-Dimer Levels in the Post-COVID Period.

Changes in the hemostatic system during COVID infection lead to hypercoagulability. Numerous studies have evaluated hemostatic abnormalities in COVID patients during acute infection, in the period of hospitalization. However, the hemostatic status following hospital discharge has not been sufficiently assessed. Considering the importance of FVIII and D-dimer levels as markers for the assessment of thrombosis, our study aimed to evaluate changes in these markers, as well as the influence of patient's age and clinical presentation of COVID infection on those hemostatic markers in the post-COVID phase. This prospective study (July 2020 to December 2022) included 115 COVID patients, 68 (59%) with asymptomatic/mild and 47 (41%) with moderate/severe clinical presentation. Patient follow-up included laboratory evaluation of FVIII and D-dimer levels at 1, 3, and 6 months following the COVID infection. Three months after the COVID infection, elevated FVIII was recorded in 44% of younger versus 65% of older individuals, p = 0.05, respectively, and 30 versus 57% (p = 0.008) 6 months post-COVID infection. With a focus on clinical presentation, a higher number of patients with moderate/severe COVID had elevated FVIII activity, but a statistically significant difference was observed only for the 6 months (32% mild vs. 53% moderate/severe, p = 0.041) post-infection time point. Following a COVID infection, an increase in FVIII activity suggests a continued hypercoagulable state in the post-COVID period and correlates with elevated D-dimer levels. This increase in FVIII is more pronounced in patients with moderate/severe clinical picture and those patients older than 50 years.

Investigation of Serum Albumin as a Dynamic Treatment-Specific Surrogate for Outcomes in Patients With Myelofibrosis Treated With Ruxolitinib.

Ruxolitinib improves splenomegaly and disease-related symptoms in most patients with myelofibrosis (MF), and it has been associated with a survival benefit in higher-risk patients with splenomegaly. Spleen volume reduction has been associated with a survival benefit in ruxolitinib-treated patients; however, its use as a surrogate is limited. We hypothesized that an anti-inflammatory response to ruxolitinib would correlate with improved patient outcomes. We interrogated serum albumin, an acute phase reactant and marker of nutritional status in 590 patients with MF and analyzed differential trajectories of albumin on the basis of ruxolitinib treatment. Additionally, we assessed the prognostic role of baseline albumin and change in albumin. We found that serum albumin levels tend to decrease in patients with MF; however, this tendency is abrogated by ruxolitinib treatment. To that end, baseline serum albumin level correlates with overall survival (OS) in patients with MF, independent of the variables that comprise the dynamic international prognostic scoring system; however, this correlation is limited to ruxolitinib-naïve patients. In ruxolitinib-treated patients, the change in serum albumin after ruxolitinib treatment, rather than the baseline value, is associated with improved OS, a finding not seen in ruxolitinib-naïve patients. These findings suggest that serum albumin, a ubiquitously available laboratory value, has specific relevance in patients with MF and reflects therapeutic response to ruxolitinib.