Analysis of Calcium-Sensing Receptor Signaling Using Dual Luciferase Assays.

Luciferases catalyze a reaction that involves the emission of light, a phenomenon referred to as "bioluminescence". The calcium-sensing receptor (CaSR), a G protein-coupled receptor (GPCR), induces characteristic signaling pathways that stimulate extracellular signal-regulated kinase 1/2 (ERK1/2) and Ca2+ mobilization from the endoplasmic reticulum. ERK1/2 causes an activation of the serum response element (SRE), whereas Ca2+ causes an activation of the nuclear factor of activated T-cells response element (NFAT-RE). Transfection of cells with a vector containing a firefly luciferase reporter gene under the control of the SRE or NFAT-RE allows the monitoring of ERK1/2 activation and Ca2+ mobilization, respectively, by measuring luminescence. In a dual luciferase assay, firefly luminescence is normalized by co-transfecting an internal control vector, which contains a constitutively active promoter driving the expression of a second luciferase, namely, Renilla luciferase, whose activity can be quantified within the same sample. Here, a protocol for the analysis of CaSR signaling using dual luciferase assays in HEK293 cells is provided. The assays can, for example, be used to investigate functional consequences of mutations in the CaSR gene.

Regulated Cell Death Pathways in Pathological Cardiac Hypertrophy.

Cardiac hypertrophy is characterized by an increased volume of individual cardiomyocytes rather than an increase in their number. Myocardial hypertrophy due to pathological stimuli encountered by the heart, which reduces pressure on the ventricular walls to maintain cardiac function, is known as pathological hypertrophy. This eventually progresses to heart failure. Certain varieties of regulated cell death (RCD) pathways, including apoptosis, pyroptosis, ferroptosis, necroptosis, and autophagy, are crucial in the development of pathological cardiac hypertrophy. This review summarizes the molecular mechanisms and signaling pathways underlying these RCD pathways, focusing on their mechanism of action findings for pathological cardiac hypertrophy. It intends to provide new ideas for developing therapeutic approaches targeted at the cellular level to prevent or reverse pathological cardiac hypertrophy.

The impact of resilience on academic performance with a focus on mature learners.

BACKGROUND: Resilience is an essential psychological trait that empowers individuals to adapt and thrive in the face of challenges. Although it is acknowledged that health professionals need to possess high levels of resilience, there has been limited research comparing how different groups of health students, particularly school leaver undergraduates and mature age graduate entry students, develop resilience in their coursework. METHODS: This study combines both objective (academic grades with validated survey results) and subjective data (interviews) to compare how resilience is related to academic coursework performance for two groups of pharmacy students: the mature age graduate entry (GE, N = 64) learners and school leaver undergraduate (UG, N = 208) learners. We employed a sequential explanatory mixed methods design using surveys, academic performance data and semi-structured interviews. The survey tapped constructs related to resilience (burnout, stressors and coping methods) while the interviews elicited a more nuanced understanding of individual and environmental factors. RESULTS: Although there was no statistical difference in burnout experience between the two groups, GE students exhibited more positive resilience attitudes than UG when selecting resilience statements on the survey. Both cohorts indicated in the survey that engaging in distraction activities (physical exercise, sleeping, listening to music, anything other than the stressor) was their most preferred method of relieving stress. Within UG student survey responses, those who indicated support from partners, friends and family had better academic performance, while those who did not report coping methods did worse academically. The three key environmental factors we identified that contributed to both undergraduate and graduate entry resilience were workload, feedback provision and psychosocial support. CONCLUSIONS: Currently, there is still a need for resilience programs geared at academic success to be implemented in higher education. This study provides objective evidence of academic success coupled with exploration into the nuances of resilience amongst different student groups. It not only highlights the differing resilience development strategies and burnout coping mechanisms in emerging health professionals, but showcases the juxtaposition of two different learner groups (UG and GE students) within a discipline. The cross-cohort facilitation of learning as in the discipline-specific strategies identified can help both groups develop resilience and inform future innovations. By comparing mature-age graduate students and younger-in-age undergraduate students, we identified a wider range of strategies and more positive attitudes to burnout in mature-age students. Health and clinical educators in university health degrees, clinical placements and clinical workplaces can develop effective training materials based on findings from this study to 1) assist undergraduate younger-age health students with developing resilience and 2) further refine mature-age health students' and practicing health professionals' resilience in today's fast-paced clinical workplaces.

Cocaine- and Amphetamine-Regulated Transcript Peptide in the Central Nervous System of the Gecko, Hemidactylus leschenaultii: Molecular Characterization, Neuroanatomical Organization, and Regulation by Neuropeptide Y.

Neuropeptide cocaine- and amphetamine-regulated transcript (CART) is widely expressed in the brains of teleosts, amphibians, birds, and mammals and has emerged as a conserved regulator of energy balance across these vertebrate phyla. However, as yet, there is no information on CART in the reptilian brain. We characterized the cDNA encoding CART and mapped CART-containing elements in the brain of gecko, Hemidactylus leschenaultii (hl) using a specific anti-CART antiserum. We report a 683-bp hlcart transcript containing a 336-bp open reading frame, which encodes a putative 111-amino acid hl-preproCART. The 89-amino acid hl-proCART generated from hl-preproCART produced two putative bioactive hl-CART-peptides. These bioactive CART-peptides were > 93% similar with those in rats/humans. Although reverse transcription-polymerase chain reaction (RT-PCR) detected hlcart-transcript in the brain, CART-containing neurons/fibers were widely distributed in the telencephalon, diencephalon, mesencephalon, rhombencephalon, spinal cord, and retina. The mitral cells in olfactory bulb, neurons in the paraventricular, periventricular, arcuate (Arc), Edinger-Westphal, and brainstem nuclei were intensely CART-positive. In view of antagonistic roles of neuropeptide Y (NPY) and CART in energy balance in the framework of mammalian hypothalamus, we probed CART-NPY interaction in the hypothalamus of H. leschenaultii. Double immunofluorescence showed a dense NPY-innervation of Arc CART neurons. Ex vivo hypothalamic slices treated with NPY/NPY-Y1-receptor agonist significantly reduced hlcart-mRNA levels in the Arc-containing tissues and CART-ir in the dorsal-Arc. However, CART-ir in ventral-Arc was unaffected. NPY via Y1-receptors may regulate energy balance by inhibiting dArc CART neurons. This study on CART in a reptilian brain fills the current void in literature and underscores the conserved feature of the neuropeptide across the entire vertebrate phyla.

Self-Regulated Learning and Learning Outcomes in Undergraduate and Graduate Medical Education: A Meta-Analysis.

Self-regulated learning (SRL) plays a pivotal role in medical education. There is a pressing need for a meta-analytical review to comprehensively evaluate the effect sizes of SRL strategies across diverse learning outcomes and levels of medical trainees. A meta-analysis was executed by searching five databases and resulted in 61 studies that met our inclusion criteria. A three-level meta-analysis was performed to examine the association between SRL strategies and various levels of learning outcomes (i.e., affective, cognitive, and behavioral learning outcomes). Moderator analyses were conducted using meta-regression, considering factors such as types of learning outcomes, training levels, SRL subscales, and quality of the studies. The analysis yielded an average effect size of .212, using Pearson's correlation coefficient, demonstrating a positive and significant association between SRL strategies and overall learning outcomes. Although our moderator analyses indicated that SRL subscales and study quality did not significantly influence the relationship between SRL strategies and learning outcomes, SRL strategies had a more pronounced effect on affective outcomes than on test scores, behavioral outcomes, and mental health outcomes. In addition, the association between SRL strategies and learning outcomes were significantly higher among the clerkship phase of undergraduate medical education than the pre-clerkship phase.

MC-LR disrupts dopamine synthesis in the substantia nigra of midbrain by enhancing the chaperone-mediated autophagy pathway through direct binding to ERK2.

Microcystins are environmental toxins produced by freshwater cyanobacteria. Microcystin-LR (MC-LR) is one of the most abundant and harmful isomers. MC-LR poses a serious threat to human health. MC-LR could penetrate the blood-brain barrier of mice and accumulate in the substantia nigra (SN) of the midbrain, leading to a reduction in dopamine levels and Parkinson's disease (PD)-like motor dysfunction in mice. The reduction in dopamine levels is a key factor contributing to movement disorders in humans with PD. Dopamine is synthesized in the dopaminergic neurons of the SN by the actions of tyrosine hydroxylase (TH) and dihydroxyphenylalanine decarboxylase (DDC). In this study, we found that MC-LR could enter dopaminergic neurons in the SN and directly bound to extracellular signal-regulated kinase 2 (ERK2), enhancing ERK2 stability. ERK2 further enhanced the transcriptional activity of Heat Shock Protein Family A Member 8 (HSPA8) and promoted the expression of Heat shock cognate 71 kDa protein (HSC70), which in turn amplified the chaperone-mediated autophagy (CMA) pathway and accelerated the degradation of TH and DDC. This affected the dopamine synthesis process, resulting in a significant reduction in dopamine levels. The study is the first to reveal that ERK2 was a direct target of MC-LR, and further enhanced CMA affecting dopamine synthesis, which has important theoretical and practical significance for environmental safety management.

Regulated Medical Waste Reduction in the Dermatology Clinic.

INTRODUCTION: The disposal of regulated medical waste (RMW) in the healthcare setting can be both costly and environmentally harmful. Prior studies have found large amounts of waste disposed of in RMW containers are inappropriately placed. Few studies to date have investigated the efficacy of waste reduction practices in the dermatology setting. METHODS: This study aims to evaluate the effectiveness of a practice-wide intervention in reducing RMW in the outpatient dermatology setting. By performing daily waste audits and two concurrent educational interventions, the amount of RMW produced and percent of appropriately placed RMW will be measured. Further analysis will occur by comparing pre-intervention values to post-intervention values. RESULTS: The percentage of waste properly placed in RMW containers prior to any intervention was 11%. Following both educational interventions, the percentage of waste properly placed in RMW containers increased by 56.1% (CI 43.7-68.5%) and the percentage of total waste produced that was identified and disposed of as RMW decreased by 6.0% (95% CI 1.2-10.8%). CONCLUSION: Our study provides practical data for dermatology providers to make small changes which can result in significant reductions of regulated medical waste, potentially providing benefits to the environment and cost-savings.

AFB1-responsive mesoporous silica nanoparticles for AFB1 quantification based on aptamer-regulated release of SERS reporter.

In this study, we propose a novel surface-enhanced Raman scattering (SERS) method for quantifying aflatoxin B1 (AFB1). This method relies on the target-triggered release of a SERS reporter from aptamer-sealed aminated mesoporous silica nanoparticles (MSNs). These MSNs were synthesized to accommodate 4-mercaptophenylboronic acid (4-MPBA) within their well-defined micropores, which were subsequently sealed with AFB1 aptamers. Upon specific binding of AFB1 to its aptamer, the conformational change in the aptamer is regulated by the presence of the target. Consequently, a positive linear relationship between the AFB1 concentration and the 4-MPBA SERS signal was observed. Under optimal conditions, the method exhibited a good linear relationship over the range of 0.1 to 5 ng/mL AFB1, with a limit of detection (LOD) of 0.03 ng/mL. This strategy was validated using wheat samples, yielding results comparable to high performance liquid chromatography-fluorescence detector (P > 0.05), confirming its reliability for detecting AFB1 in complex food matrices.

Comprehensive analysis of multiple regulated cell death risk signatures in lung adenocarcinoma.

Background: Regulated cell death (RCD) has considerable impact on tumor progress and sensitivity of treatment. Lung adenocarcinoma (LUAD) show a high resistance for conventional radiotherapies and chemotherapies. Currently, regulation of cancer cell death has been emerging as a new promising therapeutic avenue for LUAD patients. However, the crosstalk in each pattern RCD is unclear. Methods: We integrated collected the hub-genes of 12 RCD subroutines and compressively analyzed these hub-genes synergistic effect in LUAD. The characters of RCD genes expression and prognosis were developed in The Cancer Genome Atlas (TCGA)-LUAD data. We developed and validated an RCD risk model based on TCGA and GSE70294 data set, respectively. Functional annotation and tumor immunotherapy based on the risk model were also investigated. Results: 28 RCD-related genes and two LUAD molecular cluster were identified. Survival analysis revealed that the prognosis in high-risk group was worser than those in low-risk group. Functional enrichment analysis indicated that the RCD risk model correlated with immune responses. Further analysis indicated that the high-risk group in RCD risk model exhibited an immunosuppressive microenvironment and a lowly immunotherapy responder ratio. Conclusions: We present an RCD risk model which have a promising ability in predicting LUAD prognosis and immunotherapy response.

A review on synthetic inhibitors of dual-specific tyrosine phosphorylation-regulated kinase 1A (DYRK1A) for the treatment of Alzheimer's disease (AD).

Alzheimer's disease (AD) is a complex disorder that is influenced by a number of variables, such as age, gender, environmental factors, disease, lifestyle, infections, and many more. The main characteristic of AD is the formation of amyloid plaque and neurofibrillary tangles (NFT), which are caused by various reasons such as inflammation, impairment of neurotransmitters, hyperphosphorylation of tau protein, generation of toxic amyloid beta (Aß) 40/42, oxidative stress, etc. Protein kinases located in chromosome 21, namely dual-specific tyrosine phosphorylation-regulated kinase 1A (DYRK1A), play an essential role in the pathogenesis of AD. DYRK1A stimulates the Aß peptide aggregation and phosphorylation of tau protein to generate the NFT formation that causes neurodegeneration. Thus, DYRK1A is associated with AD, and inhibition of DYRK1A has the potential to treat AD. In this review, we discussed the pathophysiology of AD, various factors responsible for AD, and the role of DYRK1A in AD. We have also discussed the latest therapeutic potential of DYRK1A inhibitors for neurogenerative disease, along with their structure-activity relationship (SAR) studies. This article provides valuable information for guiding the future discovery of novel and target-specific DYRK1A inhibitors over other kinases and their structural optimization to treat AD.

Knock-out of the major regulator Flo8 in Komagataella phaffii results in unique host strain performance for methanol-free recombinant protein production.

Flo8 is a main transcriptional regulator of flocculation and pseudohyphal growth in yeast. Disruption of FLO8 in the popular recombinant protein production host Komagataella phaffii (Pichia pastoris) prevents pseudohyphal growth and reduces cell-to-surface adherence, making the mutant an interesting platform for research and industry. However, knowledge of the physiological impact of the mutation remained scarce. In-depth analysis of transcriptome data from FLO8-deficient K. phaffii revealed that Flo8 affects genes involved in cell cycle, mating, respiration, and catabolite repression additionally to flocculation targets. One gene with considerably increased expression in flo8 was GTH1, encoding a high-affinity glucose transporter in K. phaffii. Its promoter (PG1) was previously established as a strong, glucose-regulatable alternative to methanol-induced promoters. PG1 and its improved derivatives PG1-3, D-PGS4 and D-PGS5, proved to be promising candidates for controlling recombinant protein production in the FLO8-deficient background. In small-scale screenings, PG13-controlled intracellular EGFP levels were 2.8-fold higher, and yields of different secreted recombinant proteins were up to 4.8-fold increased. The enhanced productivity of the flo8 mutant in combination with the PG1 variants was transferrable to glucose-limited fed-batch processes and could largely be attributed to higher transcriptional activity of the promoter, leading to a much higher productivity per chromosomally integrated gene copy. K. phaffii flo8 has many advantageous characteristics, such as reduced surface growth and increased transcriptional strength of glucose-regulatable promoters. These features turn the flo8 strain into a valuable new base strain for various experimental designs and establish flo8 as an excellent strain background for methanol-free recombinant protein production processes.

Enablers of and Barriers to Self-Regulated and Self-Directed Aptitudes of Learning (SELF-ReDiAL) in Health Professional Education: A Systematic Review and Meta-analysis.

Introduction: Recently, accrediting organizations have focused on developing lifelong learners who possess self-regulated and self-directed aptitudes of learning (hereinafter SELF-ReDiAL or in short, SR). This meta-analysis aimed to identify factors which promote or deter SR in health professionals. Methods: Original studies which, by using self-reports, evaluated enablers of and barriers to SR in health professionals (dentistry, medicine, nursing, and pharmacology), and were indexed in Scopus® and PubMed® databases from 1 January 2000 to 31 August 2022, were retrieved. Major themes suggested to affect SR were determined, and enablers or barriers related to those themes, were considered eligible for inclusion. Studies were excluded if they were not in English, and if the full text could not be retrieved. Results: From 149 identified papers, 43 studies were subsequently included in the meta-analysis. The strongest effect was generated by wellbeing (d = 0.806; 95% confidence of interval [CI]: 0.296, 1.316). In respect of teaching method, problem-based learning (d = 0.590; 95%CI: 0.375, 0.806), team-based learning (d = 0.382; 95%CI: 0.232, 0.531), and flipped classroom (d = 0.095; 95%CI: -0.088, 0.279) showed positive effects on SR, whereas lectures were negatively associated with SR (d = -0.079; 95%CI: - 0.389, 0.230). Further, the analysis showed that SR generally decreases during the first year of enrolment (d = -0.144; 95%CI: - 0.284, -0.004), while no major change in SR is observed in the second year (d = 0.027; 95%CI: -0.044, 0.099). Conclusions: Considering that student wellbeing had the strongest observed effect on SR, policy-setting and governing bodies should incorporate methods which improve wellbeing to develop health professionals with high levels of SR. Supplementary Information: The online version contains supplementary material available at 10.1007/s40670-024-02068-y.

Modulatory Effects of Regulated Cell Death: An Innovative Preventive Approach for the Control of Mastitis.

Mastitis is a common disease worldwide that affects the development of the dairy industry due to its high incidence and complex etiology. Precise regulation of cell death and survival plays a critical role in maintaining internal homeostasis, organ development, and immune function in organisms, and regulatory abnormalities are a common mechanism of various pathological changes. Recent research has shown that regulated cell death (RCD) plays a crucial role in mastitis. The development of drugs to treat cell death and survival abnormalities that can be widely used in mastitis treatment has important clinical significance. This paper will review the molecular mechanisms of apoptosis, autophagy, pyroptosis, ferroptosis, and necroptosis and their regulatory roles in mastitis to provide a new perspective for the targeted treatment of mastitis.

Nano-Pulse Stimulation Therapy Initiates Regulated Cell Death in Skin, Unlike Bovie Radiofrequency Ablation and Cryoablation.

Background: This study describes a unique new bioelectric approach for clearing skin lesions and illustrates the clinical and histological differences between this new method and the standards of cryoablation and Bovie® radiofrequency ablation (RFA). Objectives: To determine the advantage of stimulating regulated cell death with nanosecond pulsed electric fields over the necrosis response elicited by thermal ablation modalities. Methods: Human abdominal skin was treated with cryoablation, Bovie® RFA, and nano-pulse stimulation (NPS) therapy four times before an abdominoplasty procedure was performed to collect skin for histology. The clinical appearance and corresponding histology of each treatment were documented over time and compared. Results: NPS therapy triggered regulated cell death as indicated by the appearance of activated Caspase-3 at 2 h post treatment and the absence of nuclear staining 1 day post treatment. Epidermal regeneration follows without impacting the noncellular dermis in contrast to cryoablation and Bovie® RFA which trigger necrosis and often cause scarring, inflammation, or permanent pigmentary changes. The main differences between NPS therapy and other ablation modalities are the level of fibrosis, amount of scarring, elastic fiber concentration, and inflammation. An analysis of the skin thickness 30 days after the treatment indicates that NPS-treated skin is the most similar to untreated skin but cryoablated and RF-ablated skin were 2- and 3.5-fold thicker, respectively, suggesting that they initiate necrosis rather than regulated cell death. Conclusions: We conclude that NPS therapy is a unique nonthermal modality that may be applied for clearing benign skin lesions by initiating the skin's own programmed cell death pathway instead of necrosis as used by cryoablation and Bovie® RFA.

The CellFX Percutaneous Electrode System for Nanosecond Pulsed Field Ablation.

The purpose of this study was to demonstrate the safety and performance of the CellFX Percutaneous Electrode for delivering nanosecond pulsed field ablation (nsPFA) energy to soft tissues. Three different porcine tissue types were treated, namely, liver, kidney, and skeletal muscle, at treatment levels of three times greater than clinical treatment levels. The histological characteristics of the ablation zone for each of these tissues compared with that of radiofrequency (RF) ablation on day 0 and 2 days post-treatment. Ablation zone dimensions were measured during gross necropsy after tetrazolium chloride staining and compared between the nsPFA and RF groups at 2 days post-ablation. The CellFX system successfully achieved ablation and necrosis of all treatment sites in all target tissues. No evidence of thermal effects or collagen degeneration was found at any of the nsPFA treatment sites. Overall systemic tolerability was evidenced by the absence of clinically significant changes in urinalysis and serum chemistry before and after treatments.

Chrysene contribution to bronchial asthma: Activation of TRPA1 disrupts bronchial epithelial barrier via ERK pathway.

BACKGROUND: Elevated polycyclic aromatic hydrocarbon (PAH) levels are associated with exacerbation of asthma. Chrysene is one of the most prevalent unsubstituted PAHs in the environment. Transient receptor potential ankyrin 1 (TRPA1) can be used as a chemoreceptor to detect inhaled stimuli and plays an important role in the occurrence and deterioration of asthma. Whether exposure to a high concentration of chrysene in the environment can activate TRPA1 and contribute to the development of asthma, potentially through the dysfunction of the bronchial epithelial barrier, remains unclear. METHODS: A cell-based assay was performed to verify the downregulation of the expression of E-cadherin and tight junction (TJ) proteins by chrysene in bronchial epithelial cells to explore the role of chrysene-mediated TRPA1 activation in the regulation of TJ protein expression through the extracellular signal-regulated protein kinase (ERK) pathway. Animal tests were conducted to determine whether chrysene could enhance airway hyperresponsiveness (AHR) induced by house dust mites (HDMs) and disrupt barrier function, thereby contributing to asthma. RESULTS: The cell-based assay revealed that chrysene could disrupt the function of the bronchial epithelial barrier and decrease the expression levels of E-cadherin, zonula occludens-1 (ZO-1), occludin, and claudin-5 through the ERK pathway. Chrysene induced airway epithelial barrier dysfunction primarily through TRPA1 instead of transient receptor potential vanilloid 1. TRPA1 knockdown was able to attenuate chrysene-induced downregulation of TJ protein expression and downregulate ERK activation (p-ERK). Compared with exposure to HDM alone, coexposure to chrysene and HDM resulted in an increased incidence of AHR, disruption of barrier function, and eosinophilic inflammatory responses in a mouse model of asthma. Coexposure to chrysene and HDM increased TRPA1 expression. The animal test verified that the TRPA1 inhibitor HC030031 could suppress chrysene and HDM-induced asthma in mice. CONCLUSIONS: Our findings showed that chrysene contributed to the breakdown of the function of the bronchial epithelial barrier through the TRPA1-ERK axis and therefore acted as an adjuvant to contribute to asthma.

Exploring the Use of Intracellular Chelation and Non-Iron Metals to Program Ferroptosis for Anticancer Application.

The discovery of regulated cell death (RCD) revolutionized chemotherapy. With caspase-dependent apoptosis initially being thought to be the only form of RCD, many drug development strategies aimed to synthesize compounds that turn on this kind of cell death. While yielding a variety of drugs, this approach is limited, given the acquired resistance of cancers to these drugs and the lack of specificity of the drugs for targeting cancer cells alone. The discovery of non-apoptotic forms of RCD is leading to new avenues for drug design. Evidence shows that ferroptosis, a relatively recently discovered iron-based cell death pathway, has therapeutic potential for anticancer application. Recent studies point to the interrelationship between iron and other essential metals, copper and zinc, and the disturbance of their respective homeostasis as critical to the onset of ferroptosis. Other studies reveal that several coordination complexes of non-iron metals have the capacity to induce ferroptosis. This collective knowledge will be assessed to determine how chelation approaches and coordination chemistry can be engineered to program ferroptosis in chemotherapy.

Trace Ru Incorporation Boosted Co2P Nanorods for Superior Water Electrolysis and Substrate-Paired Electrolysis Toward Value-Added Chemicals in Alkaline Medium.

Electro-valorization of biomass-derived chemicals has ensured sustainable production of value-added products, an effective approach for reducing carbon footprint, through renewable energy. Electrochemical oxidation and reduction reactions in aqueous media using H2O as a potential source for active hydrogenated and oxygenated species fulfill the purpose. In this study, Ru─Co2P nanorods are explored as a bifunctional electrocatalyst toward valorization of Organics at basic media. The in-situ electrogenerated Co3+ and Co4+ species act as active oxidants toward product selectivity. An overpotential of 68 mV is found for hydrogen evolution reaction (1 m NaOH) with Ru─Co2P. Further, used as cathode, Ru─Co2P effectively reduces furfuraldehyde to furfuryl alcohol and p-nitrophenol to p-aminophenol. Ru doping enables ease of formation of active species both for reduction and oxidation, faster charge transfer between catalyst to absorbates. Density Functional Theory calculation establishes Ru incorporation in Co2P surface results in enhanced adsorption of substrates. Ru doping modulates the electronic structure of Co2P which changes the density of states resulting in faster water dissociation and water splitting. To reach 10 mA cm-2 current density only 1.6 V is required for water electrolysis, whereas 1.4 V is enough for substrate-paired electrolysis with simultaneous oxidation of benzyl alcohol and reduction of p-nitro phenol.

Insufficient S-sulfhydration of serum and glucocorticoid-regulated kinase 1 participates in hyperhomocysteinemia-induced liver injury.

BACKGROUND & AIMS: Previous studies have established that hyperhomocysteinemia (HHcy) significantly contributes to the development of non-alcoholic steatohepatitis (NASH). Conversely, hydrogen sulfide (H2S) has shown potential in mitigating NASH. Despite these findings, it remains uncertain whether H2S can serve as a therapeutic agent against HHcy-induced liver damage. METHODS: Mice were fed a high-methionine diet to induce HHcy and HepG2 cells were exposed to homocysteine (Hcy). In both models, we assessed liver injury, H2S concentration, and autophagy levels. For rescue, sodium hydrosulfide (NaHS), an H2S donor, was used to test its potential in reversing hepatic pathological features induced by HHcy. RESULTS: 1) Hcy accumulation led to liver damage and increased autophagy. This was linked to insufficient S-sulfhydration of serum and glucocorticoid-regulated kinase 1 (SGK1) at Cys244 and Cys282, a crucial autophagy regulator. The deficiency in S-sulfhydration was resulted from downregulation of cystathionine-γ-lyase (CSE) and subsequent H2S decrease, leading to SGK1 inactivation. 2) Administration of NaHS reduced the liver damage caused by high Hcy levels and restored H2S levels, promoting the S-sulfhydration and activation of SGK1. 3) Pharmacological inhibition of SGK1 induced autosis, a specific type of cell death caused by overactivation of autophagy. Conversely, a constitutively active mutant of SGK1 (SGK1S422D) significantly decreased autophagy and improved cell viability. CONCLUSIONS: NaHS supplementation mitigates HHcy-induced liver injury by downregulating hepatic autophagy through the S-sulfhydration and activation of SGK1. This post-translational modification by H2S holds promise as a therapeutic approach for HHcy-induced liver injury.

High-throughput identification of calcium-regulated proteins across diverse proteomes.

Calcium ions play important roles in nearly every biological process, yet whole-proteome analysis of calcium effectors has been hindered by a lack of high-throughput, unbiased, and quantitative methods to identify protein-calcium engagement. To address this, we adapted protein thermostability assays in budding yeast, human cells, and mouse mitochondria. Based on calcium-dependent thermostability, we identified 2,884 putative calcium-regulated proteins across human, mouse, and yeast proteomes. These data revealed calcium engagement of signaling hubs and cellular processes, including metabolic enzymes and the spliceosome. Cross-species comparison of calcium-protein engagement and mutagenesis experiments identified residue-specific cation engagement, even within well-known EF-hand domains. Additionally, we found that the dienoyl-coenzyme A (CoA) reductase DECR1 binds calcium at physiologically relevant concentrations with substrate-specific affinity, suggesting direct calcium regulation of mitochondrial fatty acid oxidation. These discovery-based proteomic analyses of calcium effectors establish a key resource to dissect cation engagement and its mechanistic effects across multiple species and diverse biological processes.