A Method for Rigorously Selective Capture and Simultaneous Fluorescent Labeling of N-Terminal Glycine Peptides.

Although chemical methods for the selective derivatization of amino acid (AA) side chains in peptides and proteins are available, selective N-terminal labeling is challenging, especially for glycine, which has no side chain at the α-carbon position. We report here a double activation at glycine's α-methylene group that allows this AA to be differentiated from the other 19 AAs. A condensation reaction of dibenzoylmethane with glycine results in the formation of an imine, and subsequent tautomerization is followed by intramolecular cyclization, leading to the formation of a fluorescent pyrrole ring. Additionally, the approach exhibits compatibility with AAs possessing reactive side chains. Further, the method allows for selective pull-down assays of N-terminal glycine peptides from mixtures without prior knowledge of the N-terminal peptide distribution.

Single-Cell Transcriptional Signatures of Glomerular Disease in Transgenic Mice with APOL1 Variants.

BACKGROUND: APOL1 high-risk variants contribute to kidney disease among African-ancestry individuals. We sought to describe cell-specific APOL1 variants-induced pathways using two mouse models. METHODS: We characterized bacterial artificial chromosome (BAC)/APOL1 transgenic mice crossed with HIV-associated nephropathy (HIVAN) Tg26 mice and BAC/APOL1 transgenic mice given interferon-γ. RESULTS: Both mouse models showed more severe glomerular disease in APOL1-G1 compared to APOL1-G0 mice. Bulk RNA-seq of HIVAN model-glomeruli identified synergistic podocyte-damaging pathways activated by APOL1-G1 and by the HIV transgene. Single-nuclear RNA-seq revealed podocyte-specific patterns of differentially-expressed genes as a function of APOL1 alleles. Shared activated pathways, e.g. mTOR, and differentially-expressed genes, e.g. Ccn2, in podocytes in both models suggest novel markers of APOL1-associated kidney disease. HIVAN mouse-model podocyte single-nuclear RNA-seq data showed similarity to human focal segmental glomerulosclerosis glomerular RNA-seq data. Differential effects of the APOL1-G1 variant on the eukaryotic Initiation factor-2 pathway highlighted differences between the two models. CONCLUSIONS: These findings in two mouse models demonstrated both shared and distinct cell type-specific transcriptomic signatures induced by APOL1 variants. These findings suggest novel therapeutic opportunities for APOL1 glomerulopathies.

Controlled Bio-Orthogonal Catalysis Using Nanozyme-Protein Complexes via Modulation of Electrostatic Interactions.

Bio-orthogonal chemistry provides a powerful tool for drug delivery systems due to its ability to generate therapeutic agents in situ, minimizing off-target effects. Bio-orthogonal transition metal catalysts (TMCs) with stimuli-responsive properties offer possibilities for controllable catalysis due to their spatial-, temporal-, and dosage-controllable properties. In this paper, we fabricated a stimuli-responsive bio-orthogonal catalysis system based on an enhanced green fluorescent protein (EGFP)-nanozyme (NZ) complex (EGFP-NZ). Regulation of the catalytic properties of the EGFP-NZ complex was directly achieved by modulating the ionic strength of the solution. The dielectric screening introduced by salt ions allows the dissociation of the EGFP-NZ complex, increasing the access of substrate to the active site of the NZs and concomitantly increasing nanozyme activity. The change in catalytic rate of the NZ/EGFP = 1:1 complex was positively correlated with salt concentration from 0 mM to 150 mM.

TAD boundary deletion causes PITX2-related cardiac electrical and structural defects.

While 3D chromatin organization in topologically associating domains (TADs) and loops mediating regulatory element-promoter interactions is crucial for tissue-specific gene regulation, the extent of their involvement in human Mendelian disease is largely unknown. Here, we identify 7 families presenting a new cardiac entity associated with a heterozygous deletion of 2 CTCF binding sites on 4q25, inducing TAD fusion and chromatin conformation remodeling. The CTCF binding sites are located in a gene desert at 1 Mb from the Paired-like homeodomain transcription factor 2 gene (PITX2). By introducing the ortholog of the human deletion in the mouse genome, we recapitulate the patient phenotype and characterize an opposite dysregulation of PITX2 expression in the sinoatrial node (ectopic activation) and ventricle (reduction), respectively. Chromatin conformation assay performed in human induced pluripotent stem cell-derived cardiomyocytes harboring the minimal deletion identified in family#1 reveals a conformation remodeling and fusion of TADs. We conclude that TAD remodeling mediated by deletion of CTCF binding sites causes a new autosomal dominant Mendelian cardiac disorder.

Common dolphin whistle responses to experimental mid-frequency sonar.

Oceanic delphinids that occur in and around Navy operational areas are regularly exposed to intense military sonar broadcast within the frequency range of their hearing. However, empirically measuring the impact of sonar on the behavior of highly social, free-ranging dolphins is challenging. Additionally, baseline variability or the frequency of vocal state-switching among social oceanic dolphins during undisturbed conditions is lacking, making it difficult to attribute changes in vocal behavior to anthropogenic disturbance. Using a network of drifting acoustic buoys in controlled exposure experiments, we investigated the effects of mid-frequency (3-4 kHz) active sonar (MFAS) on whistle production in short-beaked (Delphinus delphis delphis) and long-beaked common dolphins (Delphinus delphis bairdii) in southern California. Given the complexity of acoustic behavior exhibited by these group-living animals, we conducted our response analysis over varying temporal windows (10 min- 5 s) to describe both longer-term and instantaneous changes in sound production. We found that common dolphins exhibited acute and pronounced changes in whistle rate in the 5 s following exposure to simulated Navy MFAS. This response was sustained throughout sequential MFAS exposures within experiments simulating operational conditions, suggesting that dolphins may not habituate to this disturbance. These results indicate that common dolphins exhibit brief yet clearly detectable acoustic responses to MFAS. They also highlight how variable temporal analysis windows-tuned to key aspects of baseline vocal behavior as well as experimental parameters related to MFAS exposure-enable the detection of behavioral responses. We suggest future work with oceanic delphinids explore baseline vocal rates a-priori and use information on the rate of change in vocal behavior to inform the analysis time window over which behavioral responses are measured.

Rational Engineering of Escherichia coli Nissle 1917 as Live Biotherapeutic to Degrade Uremic Toxin Precursors.

Uremic toxins (UTs) are microbiota-derived metabolites that accelerate the progression of kidney damage in patients with chronic kidney disease (CKD). One of the major UTs involved in CKD progression is p-cresol-sulfate (PCS), derived from dietary l-tyrosine (l-Tyr). Here, we engineered a probiotic strain of Escherichia coli Nissle 1917, to convert l-Tyr to the nontoxic compound p-coumaric acid via tyrosine ammonia lyase (TAL). First, a small metagenomic library was assessed to identify the TAL with the greatest whole-cell activity. Second, accessory genes implicated in the import of l-Tyr and export of PCA were overexpressed to enhance l-Tyr degradation by 106% and 56%, respectively. Last, random mutagenesis coupled to a novel selection and screening strategy was developed that identified a TAL variant with a 25% increase in whole-cell activity. Taken together, the final strain exhibits a 183% improvement over initial whole-cell activity and provides a promising candidate to degrade l-Tyr mediated PCS accumulation.

Functional Redundancy in Candida auris Cell Surface Adhesins Crucial for Cell-Cell Interaction and Aggregation.

Candida auris is an emerging nosocomial fungal pathogen associated with life-threatening invasive disease due to its persistent colonization, high level of transmissibility and multi-drug resistance. Aggregative and non-aggregative growth phenotypes for C. auris strains with different biofilm forming abilities, drug susceptibilities and virulence characteristics have been described. Using comprehensive transcriptional analysis we identified key cell surface adhesins that were highly upregulated in the aggregative phenotype during in vitro and in vivo grown biofilms using a mouse model of catheter infection. Phenotypic and functional evaluations of generated null mutants demonstrated crucial roles for the adhesins Als5 and Scf1 in mediating cell-cell adherence, coaggregation and biofilm formation. While individual mutants were largely non-aggregative, in combination cells were able to co-adhere and aggregate, as directly demonstrated by measuring cell adhesion forces using single-cell atomic force spectroscopy. This co-adherence indicates their role as complementary adhesins, which despite their limited similarity, may function redundantly to promote cell-cell interaction and biofilm formation. Functional diversity of cell wall proteins may be a form of regulation that provides the aggregative phenotype of C. auris with flexibility and rapid adaptation to the environment, potentially impacting persistence and virulence.

Outcome prioritization and preferences among older adults with cancer starting chemotherapy in a randomized clinical trial.

INTRODUCTION: Older adults with cancer facing competing treatments must prioritize between various outcomes. This study assessed health outcome prioritization among older adults with cancer starting chemotherapy. METHODS: Secondary analysis of a randomized trial addressing vulnerabilities in older adults with cancer. Patients completed three validated outcome prioritization tools: 1) Health Outcomes Tool: prioritizes outcomes (survival, independence, symptoms) using a visual analog scale; 2) Now vs. Later Tool: rates the importance of quality of life at three times-today versus 1 or 5 years in the future; and 3) Attitude Scale: rates agreement with outcome-related statements. The authors measured the proportion of patients prioritizing various outcomes and evaluated their characteristics. RESULTS: A total of 219 patients (median [range] age 71 [65-88], 68% with metastatic disease) were included. On the Health Outcomes Tool, 60.7% prioritized survival over other outcomes. Having localized disease was associated with choosing survival as top priority. On the Now vs. Later Tool, 50% gave equal importance to current versus future quality of life. On the Attitude Scale, 53.4% disagreed with the statement "the most important thing to me is living as long as I can, no matter what my quality of life is"; and 82.2% agreed with the statement "it is more important to me to maintain my thinking ability than to live as long as possible". CONCLUSION: Although survival was the top priority for most participants, some older individuals with cancer prioritize other outcomes, such as cognition and function. Clinicians should elicit patient-defined priorities and include them in decision-making.

Hepatic Sarcoidosis Found Incidentally in a Patient Presenting With Recurrent Clostridium difficile Infections.

A 50-year-old female who presented to our hospital for recurrent diarrhea was found to have worsening aminotransferase and alkaline phosphatase levels. Workup revealed lymphadenopathy and hepatomegaly prompting a biopsy of the liver and axillary lymph node, confirming a diagnosis of hepatic sarcoidosis. Our patient later developed cutaneous sarcoidosis. She is currently asymptomatic and is followed by gastroenterology, pulmonary, and dermatology. Recognition of extrapulmonary manifestations of sarcoidosis is important for proper management of patients. Treatment often requires a multidisciplinary approach when more than one organ system is involved.

A Janus carbaporphyrin pseudo-dimer.

Carbaporphyrin dimers, investigated for their distinctive electronic structures and exceptional properties, have predominantly consisted of systems containing identical subunits. This study addresses the associated knowledge gap by focusing on asymmetric carbaporphyrin dimers with Janus-like characteristics. The synthesis of a Janus-type carbaporphyrin pseudo-dimer 5 is presented. It displays antiaromatic characteristics on the fused side and nonaromatic behavior on the unfused side. A newly synthesized tetraphenylene (TPE) linked bis-dibenzihomoporphyrin 8 and a previously reported dibenzo[g,p]chrysene (DBC) linked bis-dicarbacorrole 9 were prepared as controls. Comprehensive analyses, including 1H NMR spectral studies, single crystal X-ray diffraction analyses, and DFT calculations, validate the mixed character of 5. A further feature of the Janus pseudo-dimer 5 is that it may be transformed into a heterometallic complex, with one side coordinating a Cu(III) center and the other stabilizing a BODIPY complex. This disparate regiochemical reactivity underscores the potential of carbaporphyrin dimers as versatile frameworks, with electronic features and site-specific coordination chemistry controlled through asymmetry. These findings position carbaporphyrin dimers as promising candidates for advances in electronic structure studies, coordination chemistry, materials science, and beyond.

Prevention of age-related truncation of γ-glutamylcysteine ligase catalytic subunit (GCLC) delays cataract formation.

A sharp drop in lenticular glutathione (GSH) plays a pivotal role in age-related cataract (ARC) formation. Despite recognizing GSH's importance in lens defense for decades, its decline with age remains puzzling. Our recent study revealed an age-related truncation affecting the essential GSH biosynthesis enzyme, the γ-glutamylcysteine ligase catalytic subunit (GCLC), at aspartate residue 499. Intriguingly, these truncated GCLC fragments compete with full-length GCLC in forming a heterocomplex with the modifier subunit (GCLM) but exhibit markedly reduced enzymatic activity. Crucially, using an aspartate-to-glutamate mutation knock-in (D499E-KI) mouse model that blocks GCLC truncation, we observed a notable delay in ARC formation compared to WT mice: Nearly 50% of D499E-KI mice remained cataract-free versus ~20% of the WT mice at their age of 20 months. Our findings concerning age-related GCLC truncation might be the key to understanding the profound reduction in lens GSH with age. By halting GCLC truncation, we can rejuvenate lens GSH levels and considerably postpone cataract onset.

Functional Redundancy in Candida auris Cell Surface Adhesins Crucial for Cell-Cell Interaction and Aggregation.

Candida auris is an emerging nosocomial fungal pathogen associated with life-threatening invasive disease due to its persistent colonization, high level of transmissibility and multi-drug resistance. Aggregative and non-aggregative growth phenotypes for C. auris strains with different biofilm forming abilities, drug susceptibilities and virulence characteristics have been described. Using comprehensive transcriptional analysis we identified key cell surface adhesins that were highly upregulated in the aggregative phenotype during in vitro and in vivo grown biofilms using a mouse model of catheter infection. Phenotypic and functional evaluations of generated null mutants demonstrated crucial roles for the adhesins Als5 and Scf1 in mediating cell-cell adherence, coaggregation and biofilm formation. While individual mutants were largely non-aggregative, in combination cells were able to co-adhere and aggregate, as directly demonstrated by measuring cell adhesion forces using single-cell atomic force spectroscopy. This co-adherence indicates their role as complementary adhesins, which despite their limited similarity, may function redundantly to promote cell-cell interaction and biofilm formation. Functional diversity of cell wall proteins may be a form of regulation that provides the aggregative phenotype of C. auris with flexibility and rapid adaptation to the environment, potentially impacting persistence and virulence.

A secondary mechanism of action for triazole antifungals in Aspergillus fumigatus mediated by hmg1.

Triazole antifungals function as ergosterol biosynthesis inhibitors and are frontline therapy for invasive fungal infections, such as invasive aspergillosis. The primary mechanism of action of triazoles is through the specific inhibition of a cytochrome P450 14-α-sterol demethylase enzyme, Cyp51A/B, resulting in depletion of cellular ergosterol. Here, we uncover a clinically relevant secondary mechanism of action for triazoles within the ergosterol biosynthesis pathway. We provide evidence that triazole-mediated inhibition of Cyp51A/B activity generates sterol intermediate perturbations that are likely decoded by the sterol sensing functions of HMG-CoA reductase and Insulin-Induced Gene orthologs as increased pathway activity. This, in turn, results in negative feedback regulation of HMG-CoA reductase, the rate-limiting step of sterol biosynthesis. We also provide evidence that HMG-CoA reductase sterol sensing domain mutations previously identified as generating resistance in clinical isolates of Aspergillus fumigatus partially disrupt this triazole-induced feedback. Therefore, our data point to a secondary mechanism of action for the triazoles: induction of HMG-CoA reductase negative feedback for downregulation of ergosterol biosynthesis pathway activity. Abrogation of this feedback through acquired mutations in the HMG-CoA reductase sterol sensing domain diminishes triazole antifungal activity against fungal pathogens and underpins HMG-CoA reductase-mediated resistance.

Jagged2 targeting in lung cancer activates anti-tumor immunity via Notch-induced functional reprogramming of tumor-associated macrophages.

Signaling through Notch receptors intrinsically regulates tumor cell development and growth. Here, we studied the role of the Notch ligand Jagged2 on immune evasion in non-small cell lung cancer (NSCLC). Higher expression of JAG2 in NSCLC negatively correlated with survival. In NSCLC pre-clinical models, deletion of Jag2, but not Jag1, in cancer cells attenuated tumor growth and activated protective anti-tumor T cell responses. Jag2-/- lung tumors exhibited higher frequencies of macrophages that expressed immunostimulatory mediators and triggered T cell-dependent anti-tumor immunity. Mechanistically, Jag2 ablation promoted Nr4a-mediated induction of Notch ligands DLL1/4 on cancer cells. DLL1/4-initiated Notch1/2 signaling in macrophages induced the expression of transcription factor IRF4 and macrophage immunostimulatory functionality. IRF4 expression was required for the anti-tumor effects of Jag2 deletion in lung tumors. Antibody targeting of Jagged2 inhibited tumor growth and activated IRF4-driven macrophage-mediated anti-tumor immunity. Thus, Jagged2 orchestrates immunosuppressive systems in NSCLC that can be overcome to incite macrophage-mediated anti-tumor immunity.

Cast immobilization duration for distal radius fractures, a systematic review.

PURPOSE: The optimal duration of immobilization for the conservative treatment of non- or minimally displaced and displaced distal radius fractures remains under debate. This research aims to review studies of these treatments to add evidence regarding the optimal immobilization period. METHODS: A comprehensive database search was conducted. Studies investigating and comparing short (< 3 weeks) versus long (> 3 weeks) immobilizations for the conservative treatment of distal radius fractures were included. The studies were evaluated for radiological and functional outcomes, including pain, grip strength, and range of motion. Two reviewers independently reviewed all studies and performed the data extraction. RESULTS: The initial database search identified 11.981 studies, of which 16 (involving 1.118 patients) were ultimately included. Patient-reported outcome measurements, grip strength, range of motion, and radiological outcomes were often better after shorter immobilization treatments. Radiological outcomes were better with longer immobilization in two studies and shorter immobilization in one study. Fourteen studies concluded that early mobilization is preferred, while the remaining two studies observed better outcomes with longer immobilization. The data were unsuitable for meta-analysis due to their heterogeneous nature. CONCLUSION: Shorter immobilization for conservatively treated distal radius fractures often yield equal or better outcomes than longer immobilizations. The immobilization for non- or minimally displaced distal radius fractures could therefore be shortened to 3 weeks or less. Displaced and reduced distal radius fractures cannot be immobilized shorter than 4 weeks due to the risk of complications. Future research with homogeneous groups could elucidate the optimal duration of immobilization.

Assessment of associations between inhaled formaldehyde and lymphohematopoietic cancer through integration of epidemiological and toxicological evidence with biological plausibility.

Formaldehyde is recognized as carcinogenic for portal of entry sites, though conclusions are mixed regarding lymphohematopoietic (LHP) cancers. This systematic review assesses the likelihood of a causal relationship between formaldehyde and LHP cancers by integrating components recommended by NASEM. Four experimental rodent bioassays and 16 observational studies in humans were included following implementation of the a priori protocol. All studies were assessed for risk of bias (RoB), and meta-analyses conducted on epidemiological studies, followed by a structured assessment of causation based on GRADE and Bradford Hill. RoB analysis identified systemic limitations precluding confidence in the epidemiological evidence due to inadequate characterization of formaldehyde exposure and a failure to adequately adjust for confounders or effect modifiers, thus suggesting that effect estimates are likely to be impacted by systemic bias. Mixed findings were reported in individual studies; meta-analyses did not identify significant associations between formaldehyde inhalation (when measured as ever/never exposure) and LHP outcomes, with meta-SMRs ranging from 0.50 to 1.51, depending on LHP subtype. No associations with LHP-related lesions were reported in reliable animal bioassays. No biologically plausible explanation linking inhalation of FA and LHP was identified, supported primarily by the lack of systemic distribution and in vivo genotoxicity. In conclusion, the inconsistent associations reported in a subset of the evidence were not considered causal when integrated with the totality of the epidemiological evidence, toxicological data, and considerations of biological plausibility. The impact of systemic biases identified herein could be quantitatively assessed to better inform causality and use in risk assessment.

Engineering Anomalously Large Electron Transport in Topological Semimetals.

Anomalous transport of topological semimetals has generated significant interest for applications in optoelectronics, nanoscale devices, and interconnects. Understanding the origin of novel transport is crucial to engineering the desired material properties, yet their orders of magnitude higher transport than single-particle mobilities remain unexplained. This work demonstrates the dramatic mobility enhancements result from phonons primarily returning momentum to electrons due to phonon-electron dominating over phonon-phonon scattering. Proving this idea, proposed by Peierls in 1932, requires tuning electron and phonon dispersions without changing symmetry, topology, or disorder. This is achieved by combining de Haas - van Alphen (dHvA), electron transport, Raman scattering, and first-principles calculations in the topological semimetals MX2 (M = Nb, Ta and X = Ge, Si). Replacing Ge with Si brings the transport mobilities from an order magnitude larger than single particle ones to nearly balanced. This occurs without changing the crystal structure or topology and with small differences in disorder or Fermi surface. Simultaneously, Raman scattering and first-principles calculations establish phonon-electron dominated scattering only in the MGe2 compounds. Thus, this study proves that phonon-drag is crucial to the transport properties of topological semimetals and provides insight to engineer these materials further.

Dual Layer vs Single Layer Woven EndoBridge Device in the Treatment of Intracranial Aneurysms: A Propensity Score-Matched Analysis.

BACKGROUND: The Woven EndoBridge (WEB) devices have been used for treating wide neck bifurcation aneurysms (WNBAs) with several generational enhancements to improve clinical outcomes. The original device dual-layer (WEB DL) was replaced by a single-layer (WEB SL) device in 2013. This study aimed to compare the effectiveness and safety of these devices in managing intracranial aneurysms. METHODS: A multicenter cohort study was conducted, and data from 1,289 patients with intracranial aneurysms treated with either the WEB SL or WEB DL devices were retrospectively analyzed. Propensity score matching was utilized to balance the baseline characteristics between the two groups. Outcomes assessed included immediate occlusion rate, complete occlusion at last follow-up, retreatment rate, device compaction, and aneurysmal rupture. RESULTS: Before propensity score matching, patients treated with the WEB SL had a significantly higher rate of complete occlusion at the last follow-up and a lower rate of retreatment. After matching, there was no significant difference in immediate occlusion rate, retreatment rate, or device compaction between the WEB SL and DL groups. However, the SL group maintained a higher rate of complete occlusion at the final follow-up. Regression analysis showed that SL was associated with higher rates of complete occlusion (OR: 0.19; CI: 0.04 to 0.8, p = 0.029) and lower rates of retreatment (OR: 0.12; CI: 0 to 4.12, p = 0.23). CONCLUSION: The WEB SL and DL devices demonstrated similar performances in immediate occlusion rates and retreatment requirements for intracranial aneurysms. The SL device showed a higher rate of complete occlusion at the final follow-up.

Association between occupational noise exposure level and pure-tone audiometry abnormalities among Metropolitan Manila Development Authority employees: A cross-sectional study.

Traffic enforcers are exposed to various occupational health and safety hazards, including noise pollution, which may lead to occupational hearing loss. This cross-sectional study aimed to estimate the prevalence of hearing loss and to assess the relationship between occupational noise exposure level (ONEL) and abnormalities in air conduction thresholds among Metropolitan Manila Development Authority (MMDA) employees along Epifanio delos Santos Avenue, Philippines. Eight-hour ONELs were measured among 108 participants working with greater than 5 years of service. Participants had hearing evaluations using pure tone audiometry (PTA) to calculate the prevalence of hearing loss. Generalized linear models with a Poisson distribution were fitted to estimate the association between ONEL and audiologic abnormalities, controlling for confounding factors. Approximately 16% of employees had hearing loss. The prevalence of hearing loss was higher with ONEL exposures greater than 85 A-weighted decibels (dBA), with traffic enforcers exposed to higher ONELs than office workers. ONELs greater than 85 dBA were related to audiologic abnormalities at different frequencies in PTA. The prevalence of audiologic abnormalities at 4000 Hz and 6000 Hz was 48% higher (adjusted prevalence ratio [aPR], 1.48; 95% CI, 1.12-1.96) and 25% higher (aPR, 1.25; 95% CI, 1.00-1.55), respectively, among participants with ONELs greater than 85 dBA than with ONELs less than or equal to 85 dBA. Participants exposed to ONELs greater than 85 dBA, more likely traffic enforcers, may have increased risk of audiologic abnormalities. Regular ONEL monitoring is warranted for occupational risk assessment of traffic enforcers. A hearing conservation program may need to be considered for this population. Additional studies are needed to determine trends in hearing deterioration among traffic enforcers.