Emerging Technologies in Cardiac Pacing.

Cardiac pacing to treat bradyarrhythmias has evolved in recent decades. Recognition that a substantial proportion of pacemaker-dependent patients can develop heart failure due to electrical and mechanical dyssynchrony from traditional right ventricular apical pacing has led to development of more physiologic pacing methods that better mimic normal cardiac conduction and provide synchronized ventricular contraction. Conventional biventricular pacing has been shown to benefit patients with heart failure and conduction system disease but can be limited by scarring and fibrosis. His bundle pacing and left bundle branch area pacing are novel techniques that can provide more physiologic ventricular activation as an alternative to conventional or biventricular pacing. Leadless pacing has emerged as another alternative pacing technique to overcome limitations in conventional transvenous pacemaker systems. Our objective is to review the evolution of cardiac pacing and explore these new advances in pacing strategies.

Structural Basis for NusA Stabilized Transcriptional Pausing.

Transcriptional pausing by RNA polymerases (RNAPs) is a key mechanism to regulate gene expression in all kingdoms of life and is a prerequisite for transcription termination. The essential bacterial transcription factor NusA stimulates both pausing and termination of transcription, thus playing a central role. Here, we report single-particle electron cryo-microscopy reconstructions of NusA bound to paused E. coli RNAP elongation complexes with and without a pause-enhancing hairpin in the RNA exit channel. The structures reveal four interactions between NusA and RNAP that suggest how NusA stimulates RNA folding, pausing, and termination. An asymmetric translocation intermediate of RNA and DNA converts the active site of the enzyme into an inactive state, providing a structural explanation for the inhibition of catalysis. Comparing RNAP at different stages of pausing provides insights on the dynamic nature of the process and the role of NusA as a regulatory factor.

A designed Copper Histidine-brace enzyme for oxidative depolymerization of polysaccharides as a model of lytic polysaccharide monooxygenase.

The "Histidine-brace" (His-brace) copper-binding site, composed of Cu(His)2 with a backbone amine, is found in metalloproteins with diverse functions. A primary example is lytic polysaccharide monooxygenase (LPMO), a class of enzymes that catalyze the oxidative depolymerization of polysaccharides, providing not only an energy source for native microorganisms but also a route to more effective industrial biomass conversion. Despite its importance, how the Cu His-brace site performs this unique and challenging oxidative depolymerization reaction remains to be understood. To answer this question, we have designed a biosynthetic model of LPMO by incorporating the Cu His-brace motif into azurin, an electron transfer protein. Spectroscopic studies, including ultraviolet-visible (UV-Vis) absorption and electron paramagnetic resonance, confirm copper binding at the designed His-brace site. Moreover, the designed protein is catalytically active towards both cellulose and starch, the native substrates of LPMO, generating degraded oligosaccharides with multiturnovers by C1 oxidation. It also performs oxidative cleavage of the model substrate 4-nitrophenyl-D-glucopyranoside, achieving a turnover number ~9% of that of a native LPMO assayed under identical conditions. This work presents a rationally designed artificial metalloenzyme that acts as a structural and functional mimic of LPMO, which provides a promising system for understanding the role of the Cu His-brace site in LPMO activity and potential application in polysaccharide degradation.

Protective effect of Cyclo(His-Pro) on peritoneal fibrosis through regulation of HDAC3 expression.

Peritoneal dialysis is a common treatment for end-stage renal disease, but complications often force its discontinuation. Preventive treatments for peritoneal inflammation and fibrosis are currently lacking. Cyclo(His-Pro) (CHP), a naturally occurring cyclic dipeptide, has demonstrated protective effects in various fibrotic diseases, yet its potential role in peritoneal fibrosis (PF) remains uncertain. In a mouse model of induced PF, CHP was administered, and quantitative proteomic analysis using liquid chromatography-tandem mass spectrometry was employed to identify PF-related protein signaling pathways. The results were further validated using human primary cultured mesothelial cells. This analysis revealed the involvement of histone deacetylase 3 (HDAC3) in the PF signaling pathway. CHP administration effectively mitigated PF in both peritoneal tissue and human primary cultured mesothelial cells, concurrently regulating fibrosis-related markers and HDAC3 expression. Moreover, CHP enhanced the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) while suppressing forkhead box protein M1 (FOXM1), known to inhibit Nrf2 transcription through its interaction with HDAC3. CHP also displayed an impact on spleen myeloid-derived suppressor cells, suggesting an immunomodulatory effect. Notably, CHP improved mitochondrial function in peritoneal tissue, resulting in increased mitochondrial membrane potential and adenosine triphosphate production. This study suggests that CHP can significantly prevent PF in peritoneal dialysis patients by modulating HDAC3 expression and associated signaling pathways, reducing fibrosis and inflammation markers, and improving mitochondrial function.

A human immune system (HIS) mouse model that dissociates roles for mouse and human FcR+ cells during antibody-mediated immune responses.

Human immune system (HIS) mice provide a model to study human immune responses in vivo. Currently available HIS mouse models may harbor mouse Fc Receptor (FcR)-expressing cells that exert potent effector functions following administration of human Ig. Previous studies showed that the ablation of the murine FcR gamma chain (FcR-γ) results in loss of antibody-dependent cellular cytotoxicity and antibody-dependent cellular phagocytosis in vivo. We created a new FcR-γ-deficient HIS mouse model to compare host (mouse) versus graft (human) effects underlying antibody-mediated immune responses in vivo. FcR-γ-deficient HIS recipients lack expression and function of mouse activating FcRs and can be stably and robustly reconstituted with human immune cells. By screening blood B-cell depletion by rituximab Ig variants, we found that human FcγRs-mediated IgG1 effects, whereas mouse activating FcγRs were dominant in IgG4 effects. Complement played a role as an IgG1 variant (IgG1 K322A) lacking complement binding activity was largely ineffective. Finally, we provide evidence that FcγRIIIA on human NK cells could mediate complement-independent B-cell depletion by IgG1 K322A. We anticipate that our FcR-γ-deficient HIS model will help clarify mechanisms of action of exogenous administered human antibodies in vivo.

Crystal structure of HPPD inhibitor sensitive protein from Oryza sativa.

4-hydroxyphenylpyruvate dioxygenase (HPPD) Inhibitor Sensitive 1 (HIS1) is an endogenous gene of rice, conferring broad-spectrum resistance to ß-triketone herbicides. Similar genes, known as HIS1-like genes (HSLs), exhibit analogous functions and can complement the herbicide-resistant characteristics endowed by HIS1. The identification of HIS1 and HSLs represents a valuable asset, as the intentional pairing of herbicides with resistance genes emerges as an effective strategy for crop breeding. Encoded by HIS1 is a Fe(II)/2-oxoglutarate-dependent oxygenase responsible for detoxifying ß-triketone herbicides through hydroxylation. However, the precise structure supporting this function remains unclear. This work, which determined the crystal structure of HIS1, reveals a conserved core motif of Fe(II)/2-oxoglutarate-dependent oxygenase and pinpoints the crucial residue dictating substrate preference between HIS1 and HSL.

Prognostic benefits of His-Purkinje capture in physiological pacemakers for bradycardia.

INTRODUCTION: Clinical outcomes of long-term ventricular septal pacing (VSP) without His-Purkinje capture remain unknown. This study evaluated the differences in clinical outcomes between conduction system pacing (CSP), VSP, and right ventricular pacing (RVP). METHODS: Consecutive patients with bradycardia indicated for pacing from 2016 to 2022 were prospectively followed for the clinical endpoints of heart failure (HF)-hospitalizations and all-cause mortality at 2 years. VSP was defined as septal pacing due to unsuccessful CSP implant or successful CSP followed by loss of His-Purkinje capture within 90 days. RESULTS: Among 1016 patients (age 73.9 ± 11.2 years, 47% female, 48% atrioventricular block), 612 received RVP, 335 received CSP and 69 received VSP. Paced QRS duration was similar between VSP and RVP, but both significantly longer than CSP (p < .05). HF-hospitalizations occurred in 130 (13%) patients (CSP 7% vs. RVP 16% vs. VSP 13%, p = .001), and all-cause mortality in 143 (14%) patients (CSP 7% vs. RVP 19% vs. VSP 9%, p < .001). The association of pacing modality with clinical events was limited to those with ventricular pacing (Vp) > 20% (pinteraction < .05). Adjusting for clinical risk factors among patients with Vp > 20%, VSP (adjusted hazard ratio [AHR]: 4.74, 95% confidence interval [CI]: 1.57-14.36) and RVP (AHR: 3.08, 95% CI: 1.44-6.60) were associated with increased hazard of HF-hospitalizations, and RVP (2.52, 95% CI: 1.19-5.35) with increased mortality, compared to CSP. Clinical endpoints did not differ between VSP and RVP with Vp > 20%, or amongst groups with Vp < 20%. CONCLUSION: Conduction system capture is associated with improved clinical outcomes. CSP should be preferred over VSP or RVP during pacing for bradycardia.

Narrow QRS ectopy with concealed connections from a para-Hisian origin to the proximal left fascicles.

INTRODUCTION: Catheter ablation of ectopy originating from the vicinity of the His bundle can be challenging. METHODS AND RESULTS: We report a case of a 33-year-old man with narrow QRS ectopy with preferential conduction from a para-Hisian origin to the proximal left fascicles, which was successfully eliminated by radiofrequency ablation in the right coronary cusp, guided by ultrahigh-resolution mapping of the His bundle, bundle branch, and fascicular electrograms. CONCLUSION: Some narrow QRS ectopy may originate from the vicinity of the conduction system, instead of the "true" conduction system, and have concealed connections from its origin to the conduction system.

CLDN1 Arg81His founder variant causes ichthyosis, leukocyte vacuoles, alopecia, and sclerosing cholangitis (ILVASC) syndrome in Moroccan Jews.

Neonatal ichthyosis and sclerosing cholangitis syndrome (NISCH), also known as ichthyosis, leukocyte vacuoles, alopecia, and sclerosing cholangitis (ILVASC), is an extremely rare disease of autosomal recessive inheritance, resulting from loss of function of the tight junction protein claudin-1. Its clinical presentation is highly variable, and is characterized by liver and ectodermal involvement. Although most ILVASC cases described to date were attributed to homozygous truncating variants in CLDN1, a single missense variant CLDN1 p.Arg81His, associated with isolated skin ichthyosis phenotype, has been recently reported in a family of Moroccan Jewish descent. We now describe seven patients with ILVASC, originating from four non consanguineous families of North African Jewish ancestry (including one previously reported family), harboring CLDN1 p.Arg81His variant, and broaden the phenotypic spectrum attributed to this variant to include teeth, hair, and liver/bile duct involvement, characteristic of ILVASC. Furthermore, we provide additional evidence for pathogenicity of the CLDN1 p.Arg81His variant by transmission electron microscopy of the affected skin, revealing distorted tight junction architecture, and show through haplotype analysis in the vicinity of the CLDN1 gene, that this variant represents a founder variant in Jews of Moroccan descent with an estimated carrier frequency of 1:220.

Conduction system pacing: how far are we from the "electrical" bypass?

Conduction system pacing is an alternative practice to conventional right ventricular apical pacing. It is a method that maintains physiologic ventricular activation, based on a correct pathophysiological basis, in which the pacing lead bypasses the lesion of the electrical fibers and the electrical impulse transmits through the intact adjacent conduction system. For this reason, it might be reasonably characterized by the term "electrical bypass" compared to the coronary artery bypass in revascularization therapy. In this review, reference is made to the sequence of events in which conventional right ventricular pacing may cause adverse outcomes. Furthermore, there is a reference to alternative strategies and pacing sites. Interest focuses on the modalities for which there are data from the literature, namely for the right ventricular (RV) septal pacing, the His bundle pacing (HBP), and the left bundle branch pacing (LBBP). A more extensive reference is about the HBP, for which there are the most updated data. We analyze the considerations that limit HBP-wide application in three axes, and we also present the data for the implantation and follow-up of these patients. The indications with their most important studies to date are then described in detail, not only in their undoubtedly positive findings but also in their weak aspects, because of which this pacing mode has not yet received a strong recommendation for implementation. Finally, there is a report on LBBP, focusing mainly on its points of differentiation from HBP.

Proton Domino Reactions at an Imidazole Relay Control the Oxidation of a TyrZ-His190 Artificial Mimic of Photosystem II.

A close mimic of P680 and the TyrosineZ-Histidine190 pair in photosystem II (PS II) has been synthesized using a ruthenium chromophore and imidazole-phenol ligands. The intramolecular oxidation of the ligands by the photoproduced Ru(III) species is characterized by a small driving force, very similar to PS II where the complexity of kinetics was attributed to the reversibility of electron transfer steps. Laser flash photolysis revealed biphasic kinetics for ligand oxidation. The fast phase (τ<50 ns) corresponds to partial oxidation of the imidazole-phenol ligand, proton transfer within the hydrogen bond, and formation of a neutral phenoxyl radical. The slow phase (5-9 µs) corresponds to full oxidation of the ligand which is kinetically controlled by deprotonation of the distant 1-nitrogen of the imidazolium. These results show that imidazole with its two protonatable sites plays a special role as a proton relay in a 'proton domino' reaction.

Synthesis, Structure and Reactivity of a Mononuclear N,N,O-Bound Fe(II) α-Keto-Acid Complex.

A bulky, tridentate phenolate ligand (ImPh2 NNOtBu ) was used to synthesise the first example of a mononuclear, facial, N,N,O-bound iron(II) benzoylformate complex, [Fe(ImPh2 NNOtBu )(BF)] (2). The X-ray crystal structure of 2 reveals that the iron centre is pentacoordinate (τ=0.5), with a vacant site located cis to the bidentate BF ligand. The Mössbauer parameters of 2 are consistent with high-spin iron(II), and are very close to those reported for α-ketoglutarate-bound non-heme iron enzyme active sites. According to NMR and UV-vis spectroscopies, the structural integrity of 2 is retained in both coordinating and non-coordinating solvents. Cyclic voltammetry studies show that the iron centre has a very low oxidation potential and is more prone to electrochemical oxidation than the redox-active phenolate ligand. Complex 2 reacts with NO to form a S=3 /2 {FeNO}7 adduct in which NO binds directly to the iron centre, according to EPR, UV-vis, IR spectroscopies and DFT analysis. Upon O2 exposure, 2 undergoes oxidative decarboxylation to form a diiron(III) benzoate complex, [Fe2 (ImPh2 NNOtBu )2 (µ2 -OBz)(µ2 -OH)2 ]+ (3). A small amount of hydroxylated ligand was also observed by ESI-MS, hinting at the formation of a high-valent iron(IV)-oxo intermediate. Initial reactivity studies show that 2 is capable of oxygen atom transfer reactivity with O2 , converting methyl(p-tolyl)sulfide to sulfoxide.

Optimized expression and purification of a human adenosine deaminase in E. coli and characterization of its Asp8Asn variant.

Homo sapiens adenosine deaminase isoform 1 (HsADA1) hydrolyzes adenosine and 2-deoxyadenosine as a key step in the purine nucleoside salvage pathway. Some HsADA1 mutations have severe deleterious effects, as is the case in a severe combined immunodeficiency resulting from loss of enzyme activity (ADA-SCID). Other mutations that reduce enzyme activity, for instance the Asp8Asn (D8N) variant, do not cause ADA-SCID but are correlated with other consequences to health. To ease further study of HsADA1 and its variants, we optimized an inexpensive, recombinant expression process in an Escherichia coli host through multiplexed parameter testing enabled by a lysate-based microtiter plate assay. We demonstrate the importance of gene codon usage, induction time and temperature, and alcohol supplementation towards improving enzyme yield to a final titer of 5 mg per liter of culture. We further show that use of a double-histidine-tag (his-tag) system greatly improves purity. We then utilize our expression and purification framework to produce the HsADA1 D8N variant, which had previously not been purified to homogeneity. We confirm that the D8N variant is ∼30% less active than the wildtype HsADA1 and show that it better retains its activity in human serum. Additionally, we show that both HsADA1 and the D8N variant have heightened activity in serum, driven in part by a previously undescribed phenomenon involving albumin. Therefore, this work presents a valuable process to produce HsADA1 that allows for insights into it and its variants' behavior. We also confirm the utility of lysate-based activity assays towards finding optimal E. coli expression conditions for enzymes and show how fusing his-tags in tandem can enhance product purity.

Soluble expression of recombinant human interleukin-2 in Escherichia coli and its facile production.

Recombinant human interleukin-2 (rhIL-2) represents one of the most difficult-to-produce cytokines in E. coli due to its extreme hydrophobicity and high tendency to formation of inclusion bodies. Refolding of rhIL-2 inclusion bodies always represents cumbersome downstream processes and low production efficiency. Herein, we disclosed a fusion strategy for efficiently soluble expression and facile production of rhIL-2 in E. coli Origami B (DE3) host. A two-tandem SUMO fusion partner (His-2SUMO) with a unique SUMO protease cleavage site at C-terminus was devised to fuse with the N-terminus of rhIL-2 and the fusion protein (His-2SUMO-rhIL-2) was almost completely expressed in a soluble from. The fusion partner could be efficiently removed by Ulp1 cleavage and the rhIL-2 was simply produced by a two-step Ni-NTA affinity chromatography with a considerable purity and whole recovery. The eventually obtained rhIL-2 was well-characterized and the results showed that the purified rhIL-2 exhibits a compact and ordered structure. Although the finally obtained rhIL-2 exists in a soluble aggregates form and the aggregation probably has been occurred during expression stage, the soluble rhIL-2 aggregates remain exhibit comparable bioactivity with the commercially available rhIL-2 drug formulation.

Novel melanin-derived stationary phase for immobilized metal ion affinity chromatography in recombinant His-tagged protein purification.

The matrix of the stationary phase is a crucial element in affinity chromatography for protein purification. Various materials, including polymer or magnetic materials, have been employed as the matrix in the purification of His-tagged protein. Here, for the first time, we utilized a combination of melanin and alginate, both natural polymer materials, to synthesize Ni-melanin/alginate (Ni-M/A) beads for His-tagged protein purification. We investigated the binding of His-tagged Mpro on the Ni-M/A beads, referred to as Ni-M/A-Mpro, and assessed the elution efficiency of Mpro from the beads. Our examination involved FTIR, EDS, XRD, SDS-PAGE, and Western blotting methods. FTIR spectra revealed notable changes in the stretching patterns and intensities of hydroxyl, amine, carbonyl, imine and amide chemical groups, when Mpro protein was present in the Ni-M/A sample. XRD spectra demonstrated the occurrence of two Nickel peaks at 35-40 deg and 40-45 deg in Ni-M/A, but only one nickel peak at 35-40 deg in Ni-M/A-Mpro, indicating the binding of Mpro on the Nickel ions. EDS analysis reported a decrease in the concentration of Nickel on the surface of Ni-M/A from 16% to 7% when Mpro protein was loaded into the stationary phase. Importantly, our data indicated that the purity of the His-tagged protein Mpro after purification reached 97% after just one-step purification using the Ni-M/A stationary phase. Moreover, the binding capacity of Ni-M/A for Mpro was approximately 5.2 mg/g with recovery efficiency of 40%. Our results suggested Ni-M/A as a highly potential solid phase for affinity chromatography in the purification of His-tagged protein.

Histidine dephosphorylation of the Gß protein GPB-1 promotes axon regeneration in C. elegans.

Histidine phosphorylation is an emerging noncanonical protein phosphorylation in animals, yet its physiological role remains largely unexplored. The protein histidine phosphatase (PHPT1) was recently identified for the first time in mammals. Here, we report that PHIP-1, an ortholog of PHPT1 in Caenorhabditis elegans, promotes axon regeneration by dephosphorylating GPB-1 Gß at His-266 and inactivating GOA-1 Goα signaling, a negative regulator of axon regeneration. Overexpression of the histidine kinase NDK-1 also inhibits axon regeneration via GPB-1 His-266 phosphorylation. Thus, His-phosphorylation plays an antiregenerative role in C. elegans. Furthermore, we identify a conserved UNC-51/ULK kinase that functions in autophagy as a PHIP-1-binding protein. We demonstrate that UNC-51 phosphorylates PHIP-1 at Ser-112 and activates its catalytic activity and that this phosphorylation is required for PHIP-1-mediated axon regeneration. This study reveals a molecular link from ULK to protein histidine phosphatase, which facilitates axon regeneration by inhibiting trimeric G protein signaling.

Mid-term performance of His bundle pacing and usefulness of backup leads.

Ventricular backup leads may be considered in selected patients with His bundle pacing (HBP), but it remains unknown to what extent this is useful. A total of 184 HBP patients were studied. At last follow-up, 147 (79.9%) patients retained His bundle capture at programmed output. His bundle pacing lead revision was performed in 5/36 (13.9%) patients without a backup lead and in 3/148 (2.0%) patients with a backup lead (P = 0.008). One patient without a backup lead had syncope due to atrial oversensing. Thus, implantation of ventricular backup leads may avoid lead revision and adverse events in selected HBP patients.

The clinical anatomy of the atrioventricular conduction axis.

It is axiomatic that the chances of achieving accurate capture of the conduction axis and its fascicles will be optimized by equally accurate knowledge of the relationship of the components to the recognizable cardiac landmarks, and we find it surprising that acknowledged experts should continue to use drawings that fall short in terms of anatomical accuracy. The accuracy achieved by Sunao Tawara (1906) in showing the location of the atrioventricular conduction axis is little short of astounding. Our purpose in bringing this to current attention is to question the need of the experts to have produced such inaccurate representations, since the findings of Tawara have been extensively endorsed in very recent years. The recent studies do no more than point to the amazing accuracy of the initial account of Tawara. At the same time, we draw attention to the findings described in the middle of the 20th century by Ivan Mahaim (1947). These observations have tended to be ignored in recent accounts. They are, perhaps, of equal significance to those seeking specifically to pace the left fascicles of the branching atrioventricular bundle.

Validating the Delone and Mclean's model in a developing country's infectious disease pandemic context.

BACKGROUND: This study aimed at validating the updated DeLone and McLean's information systems success model (D&MISS) in a developing country's infectious disease pandemic preparedness and response context. The findings from this study are relevant to inform policies and actions for enhancing developing countries' the Health Information System's (HIS) performance, and specifically to improve their future pandemic readiness and response. The study sought to respond to a key research question: to what extent can the D&MISS model provide evidence to enhance the HIS's infectious disease pandemic readiness and response in developing countries? METHOD: A cross-sectional study design that involved a multi-stage probability sampling approach to select eligible healthcare workers was applied. Conducted in Nigeria and Liberia, 576 primary healthcare workers, out of the proposed 600, participated, representing a response rate of 96%. The D&MISS model served as the theoretical underpinning for this study, and nine hypothesized relationships were stated before the study based on the interconnectedness of the model's six dimensions. Structural Equation Modelling (SEM) data analysis using the Partial Least Square approach was used to determine if hypothesized relationships were supported. RESULTS: 70% of the observed variance in the Net Benefit construct was explained by the predictive influence of the Use and User Satisfaction constructs. The Use construct had a slightly more substantial predictive influence than the User Satisfaction construct. Eight of the nine hypothesized relationships were supported, except for the relationship between Information Quality and Use. The relationships between System Quality and Use and User Satisfaction and Net Benefit had the highest beta coefficient, statistically significant at p < 0.05. CONCLUSION AND RELEVANCE: The D&MISS model demonstrated its relevance in providing evidence on the gaps of the HISs regarding future pandemic preparedness and response. However, from a future research opportunity, its enhancement and modifications with context-specific dimensions peculiar to developing countries will improve its ability to provide more context-specific evidence to improve pandemic preparedness and response for developing countries.

His potential injury as the end point of screwing by a continuous recording technique in His bundle pacing: A case report.

BACKGROUND: His bundle pacing (HBP) engaged electrical activation of both ventricles by stimulating the His-Purkinje network, which could avoid marked ventricles dyssynchrony. The lead was given three to five clockwise rotations at the site with the His potential to anchor the interventricular septum. In 2018, the Multicenter His Bundle Pacing Collaborative Working Group recommended that the His bundle capture threshold should be lower than 2.5 V/1 ms in non-pacing-dependent patients, and pacing-dependent patients should have a lower adjacent ventricular capture threshold as self-backup. Therefore, to avoid safety issues such as loss of capture caused by increased threshold, we believe that more stringent criteria should be adopted in patients with atrioventricular block (AVB). In previous studies, the connection cable needed to be disconnected during the screwing. When the procedure was finished, the performer found that the patients with His bundle injury could obtain a lower threshold than those without His bundle injury. Although no studies of new bundle branch block (BBB) or AVB by the acute His bundle injury was reported. However, It is worrying that the damage of His bundle seems random during the procedure. How to balance avoiding severe injury with a lower capture threshold? At present, we report a case of light His injury and lower His capture threshold under continuous intracardiac electrocardiogram monitoring.