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Although biologics have been revolutionizing the treatment of inflammatory bowel diseases (IBD) over the past decade, a significant number of patients still fail to benefit from these drugs. Overcoming the nonresponse to biologics is one of the top challenges in IBD treatment. In this study, we revealed that hyaluronan (HA), an extracellular matrix (ECM) component in the gut, is associated with nonresponsiveness to infliximab and vedolizumab therapy in patients with IBD. In murine colitis models, inhibition of HA synthase 2-mediated (HAS2-mediated) HA synthesis sensitized the therapeutic response to infliximab. Mechanistically, HA induced the expression of MMP3 in colonic fibroblasts by activating STAT3 signaling, thereby mediating the proteolytic cleavage of multiple IgG1 biologics. Finally, we found that macrophage-derived factors upregulated HAS2 expression in fibroblasts, thereby contributing to infliximab nonresponse. In summary, we identified a pathogenic connection between abnormal ECM remodeling and biologics nonresponse and provided insights for the precise therapy for IBD.

Macrophages play a crucial role in promoting perfusion recovery and revascularization after ischemia through antiinflammatory polarization, a process essential for the treatment of peripheral artery disease (PAD). Mitochondrial dynamics, particularly regulated by the fission protein DRP1, are closely linked to macrophage metabolism and inflammation. However, the role of DRP1 in reparative neovascularization remains unexplored. Here, we show that DRP1 expression was increased in F4/80+ macrophages within ischemic muscle on day 3 after hind limb ischemia (HLI), an animal model of PAD. Mice lacking Drp1 in myeloid cells exhibited impaired limb perfusion recovery, angiogenesis, and muscle regeneration after HLI. These effects were associated with increased proinflammatory M1-like macrophages, p-NF-κB, and TNF-α, and reduced antiinflammatory M2-like macrophages and p-AMPK in ischemic muscle of myeloid Drp1-/- mice. In vitro, Drp1-deficient macrophages under hypoxia serum starvation (HSS), an in vitro PAD model, demonstrated enhanced glycolysis via reducing p-AMPK as well as mitochondrial dysfunction, and excessive mitochondrial ROS production, resulting in increased proinflammatory M1-gene and reduced antiinflammatory M2-gene expression. Conditioned media from HSS-treated Drp1-/- macrophages exhibited increased proinflammatory cytokine secretion, leading to suppressed angiogenesis in endothelial cells. Thus, macrophage DRP1 deficiency under ischemia drives proinflammatory metabolic reprogramming and macrophage polarization, limiting revascularization in experimental PAD.

Urinary neutrophils are a hallmark of urinary tract infection (UTI), yet the mechanisms governing their activation, function, and efficacy in controlling infection remain incompletely understood. Tamm-Horsfall glycoprotein (THP), the most abundant protein in urine, uses terminal sialic acids to bind an inhibitory receptor and dampen neutrophil inflammatory responses. We hypothesized that neutrophil modulation is an integral part of THP-mediated host protection. In a UTI model, THP-deficient mice showed elevated urinary tract bacterial burdens, increased neutrophil recruitment, and more severe tissue histopathological changes compared with WT mice. Furthermore, THP-deficient mice displayed impaired urinary NETosis during UTI. To investigate the effect of THP on NETosis, we coupled in vitro fluorescence-based NET assays, proteomic analyses, and standard and imaging flow cytometry with peripheral human neutrophils. We found that THP increases proteins involved in respiratory chain, neutrophil granules, and chromatin remodeling pathways; enhances NETosis in an ROS-dependent manner; and drives NET-associated morphologic features including nuclear decondensation. These effects were observed only in the presence of a NETosis stimulus and could not be solely replicated with equivalent levels of sialic acid alone. We conclude that THP is a critical regulator of NETosis in the urinary tract, playing a key role in host defense against UTI.

The hERG1 potassium channel conducts the cardiac repolarizing current, IKr. hERG1 has emerged as a therapeutic target for cardiac diseases marked by prolonged actional potential duration (APD). Unfortunately, many hERG1 activators display off-target and proarrhythmic effects that limit their therapeutic potential. A Per-Arnt-Sim (PAS) domain in the hERG1 N-terminus reduces IKr by slowing channel activation and promoting inactivation. Disrupting PAS activity increases IKr and shortens APD in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). We thus hypothesized that the hERG1 PAS domain could represent a therapeutic target to reduce arrhythmogenic potential in a long QT syndrome (LQTS) background. To test this, we measured the antiarrhythmic capacity of a PAS-disabling single-chain variable fragment antibody, scFv2.10, in a hiPSC-CM line derived from a Jervell and Lange Nielsen syndrome (JLN) patient. JLN is a severe form of LQTS caused by autosomal recessive mutations in KCNQ1. The patient in this study carried compound heterozygous mutations in KCNQ1. Corresponding JLN hiPSC-CMs displayed prolonged APD and early after depolarizations (EADs). Disrupting PAS with scFv2.10 increased IKr, shortened APD, and reduced the incidence of EADs. These data demonstrate that the hERG1 PAS domain could serve as a therapeutic target to treat disorders of cardiac electrical dysfunction.

Background: The immunogenicity of current influenza vaccines need improvement. Inactivated influenza and COVID-19 mRNA vaccines can be co-administered but randomized controlled trial data is lacking on whether the two vaccines are more immunogenic if given in the same or opposite arms. Murine studies suggest mRNA vaccines can adjuvant influenza vaccines when co-formulated and delivered together.

Methods: We randomly assigned 56 adults to receive the Afluria quadrivalent inactivated influenza and Moderna monovalent SARS-CoV-2 XBB.1.5 mRNA vaccines, either in opposite arms or both in the same arm at the same site. The primary endpoint was the difference in median combined serum haemagglutination inhibition titre to the H1, H3 and B-Vic vaccine influenza strains after vaccination.

Results: We found no significant difference in haemagglutination inhibition antibody levels between the groups (p = 0.30), with the same arm group having a 1.26-fold higher titre than the opposite arm group. There was no difference in analyses of antibodies to individual influenza strains, nor in nasal or saliva antibody levels. While both binding and neutralising antibody titres against SARS-CoV-2 were not significantly different between groups post-vaccination, there was a higher fold-change in BA.5 and ancestral strain neutralising antibodies in the opposite arm group.

Conclusion: Influenza vaccination is equivalently immunogenic if given in same or opposite arms as the SARS-CoV-2 vaccine, but it may be preferable to administer the SARS-CoV-2 vaccine at a different site to influenza vaccines.

Trial registration: Australian New Zealand Clinical Trials Registry ACTRN12624000445572.

Funding: Australian National Health and Medical Research Council and Medical Research Future Fund.

High apolipoprotein B-containing (apoB-containing) low-density lipoproteins (LDLs) and low apoA1-containing high-density lipoproteins (HDLs) are associated with atherosclerotic cardiovascular diseases. In search of a molecular regulator that could simultaneously and reciprocally control both LDL and HDL levels, we screened a microRNA (miR) library using human hepatoma Huh-7 cells. We identified miR-541-3p that both significantly decreases apoB and increases apoA1 expression by inducing mRNA degradation of 2 different transcription factors, Znf101 and Casz1. We found that Znf101 enhances apoB expression, while Casz1 represses apoA1 expression. The hepatic knockdown of Casz1 in mice increased plasma apoA1, HDL, and cholesterol efflux capacity. The hepatic knockdown of Zfp961, an ortholog of Znf101, reduced lipogenesis and production of triglyceride-rich lipoproteins and atherosclerosis, without causing hepatic lipid accumulation. This study identifies hepatic Znf101/Zfp961 and Casz1 as potential therapeutic targets to alter plasma lipoproteins and reduce atherosclerosis without causing liver steatosis.

The oral mucosa undergoes daily insults, and stem cells in the epithelial basal cell layer regenerate gingiva tissue to maintain oral health. The Iroquois Homeobox 1 (IRX1) protein is expressed in the stem cell niches in human/mouse oral epithelium and mesenchyme under homeostasis. We found that Irx1+/- heterozygous (Het) mice have delayed wound closure, delayed morphological changes of regenerated epithelium, and defective keratinocyte proliferation and differentiation during wound healing. RNA-Seq analyses between WT and Irx1+/- mice at 3 days postinjury (dpi) found impaired epithelial migration and decreased keratinocyte-related genes upon injury. IRX1-expressing cells are found in the gingival epithelial basal cell layer, a stem cell niche for gingival maintenance. IRX1-expressing cells are also found in cell niches in the underlying stroma. IRX1 activates SOX9 in the transient amplifying layer to increase cell proliferation, and EGF signaling is activated to induce cell migration. Krt14CreERT lineage tracing experiments reveal defects in the stratification of the Irx1+/- HET mouse oral epithelium. IRX1 is primed at the base of the gingiva in the basal cell layer of the oral epithelium, facilitating rapid and scarless wound healing through activating SOX9 and the EGF signaling pathway.

The impact of diet-induced maternal obesity on offspring airway hyperresponsiveness was studied in a diversity outbred mouse model that mirrors human genetic diversity. Female mice were started on high-fat or regular diet 8 weeks before breeding and throughout pregnancy and lactation. After weaning, all offspring were fed a regular diet. By 12 weeks, body weight and fat were increased in offspring of high-fat diet-fed dams, which was accompanied by metabolic dysfunction and hyperinsulinemia. This was followed by increased epithelial sensory innervation and increased bronchoconstriction to inhaled 5-hydroxytryptamine at 16 weeks. Bronchoconstriction was nerve mediated and blocked by vagotomy or atropine. A high-fat diet before pregnancy exerted the most influence on offspring airway physiology. Maternal obesity induced metabolic dysfunction and hyperinsulinemia, resulting in hyperinnervation and subsequent increased reflex-mediated hyperresponsiveness in their offspring. This is relevant to our understanding of asthma inheritance, considering the genetic diversity of humans.

The availability and integration of electrophysiological and molecular data from the living brain is critical in understanding and diagnosing complex human disease. Intracranial stereo electroencephalography (SEEG) electrodes used for identifying the seizure focus in patients with epilepsy could enable the integration of such multimodal data. Here, we report multimodal profiling of epileptic brain activity via explanted depth electrodes (MoPEDE), a method that recovers extensive protein-coding transcripts, including cell type markers, DNA methylation, and short variant profiles from explanted SEEG electrodes matched with electrophysiological and radiological data allowing for high-resolution reconstructions of brain structure and function. We found gene expression gradients that corresponded with the neurophysiology-assigned epileptogenicity index but also outlier molecular fingerprints in some electrodes, potentially indicating seizure generation or propagation zones not detected during electroclinical assessments. Additionally, we identified DNA methylation profiles indicative of transcriptionally permissive or restrictive chromatin states and SEEG-adherent differentially expressed and methylated genes not previously associated with epilepsy. Together, these findings validate that RNA profiles and genome-wide epigenetic data from explanted SEEG electrodes offer high-resolution surrogate molecular landscapes of brain activity. The MoPEDE approach has the potential to enhance diagnostic decisions and deepen our understanding of epileptogenic network processes in the human brain.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease 2019 (COVID-19), has emerged as a global pandemic pathogen with high mortality. While treatments have been developed to reduce morbidity and mortality of COVID-19, more antivirals with broad-spectrum activities are still needed. Here, we identified lonafarnib (LNF), a Food and Drug Administration-approved inhibitor of cellular farnesyltransferase (FTase), as an effective anti-SARS-CoV-2 agent. LNF inhibited SARS-CoV-2 infection and acted synergistically with known anti-SARS antivirals. LNF was equally active against diverse SARS-CoV-2 variants. Mechanistic studies suggested that LNF targeted multiple steps of the viral life cycle. Using other structurally diverse FTase inhibitors and a LNF-resistant FTase mutant, we demonstrated a key role of FTase in the SARS-CoV-2 life cycle. To demonstrate in vivo efficacy, we infected SARS-CoV-2-susceptible humanized mice expressing human angiotensin-converting enzyme 2 (ACE2) and treated them with LNF. LNF at a clinically relevant dose suppressed the viral titer in the respiratory tract and improved pulmonary pathology and clinical parameters. Our study demonstrated that LNF, an approved oral drug with excellent human safety data, is a promising antiviral against SARS-CoV-2 that warrants further clinical assessment for treatment of COVID-19 and potentially other viral infections.