A Zika virus-specific IgM elicited in pregnancy exhibits ultrapotent neutralization.

Congenital Zika virus (ZIKV) infection results in neurodevelopmental deficits in up to 14% of infants born to ZIKV-infected mothers. Neutralizing antibodies are a critical component of protective immunity. Here, we demonstrate that plasma IgM contributes to ZIKV immunity in pregnancy, mediating neutralization up to 3 months post-symptoms. From a ZIKV-infected pregnant woman, we isolated a pentameric ZIKV-specific IgM (DH1017.IgM) that exhibited ultrapotent ZIKV neutralization dependent on the IgM isotype. DH1017.IgM targets an envelope dimer epitope within domain II. The epitope arrangement on the virion is compatible with concurrent engagement of all ten antigen-binding sites of DH1017.IgM, a solution not available to IgG. DH1017.IgM protected mice against viremia upon lethal ZIKV challenge more efficiently than when expressed as an IgG. Our findings identify a role for antibodies of the IgM isotype in protection against ZIKV and posit DH1017.IgM as a safe and effective candidate immunotherapeutic, particularly during pregnancy.

Maturation Pathway from Germline to Broad HIV-1 Neutralizer of a CD4-Mimic Antibody.

Antibodies with ontogenies from VH1-2 or VH1-46-germline genes dominate the broadly neutralizing response against the CD4-binding site (CD4bs) on HIV-1. Here, we define with longitudinal sampling from time-of-infection the development of a VH1-46-derived antibody lineage that matured to neutralize 90% of HIV-1 isolates. Structures of lineage antibodies CH235 (week 41 from time-of-infection, 18% breadth), CH235.9 (week 152, 77%), and CH235.12 (week 323, 90%) demonstrated the maturing epitope to focus on the conformationally invariant portion of the CD4bs. Similarities between CH235 lineage and five unrelated CD4bs lineages in epitope focusing, length-of-time to develop breadth, and extraordinary level of somatic hypermutation suggested commonalities in maturation among all CD4bs antibodies. Fortunately, the required CH235-lineage hypermutation appeared substantially guided by the intrinsic mutability of the VH1-46 gene, which closely resembled VH1-2. We integrated our CH235-lineage findings with a second broadly neutralizing lineage and HIV-1 co-evolution to suggest a vaccination strategy for inducing both lineages.

A compact stem-loop DNA aptamer targets a uracil-binding pocket in the SARS-CoV-2 nucleocapsid RNA-binding domain.

SARS-CoV-2 nucleocapsid (N) protein is a structural component of the virus with essential roles in the replication and packaging of the viral RNA genome. The N protein is also an important target of COVID-19 antigen tests and a promising vaccine candidate along with the spike protein. Here, we report a compact stem-loop DNA aptamer that binds tightly to the N-terminal RNA-binding domain of SARS-CoV-2 N protein. Crystallographic analysis shows that a hexanucleotide DNA motif (5'-TCGGAT-3') of the aptamer fits into a positively charged concave surface of N-NTD and engages essential RNA-binding residues including Tyr109, which mediates a sequence-specific interaction in a uracil-binding pocket. Avid binding of the DNA aptamer allows isolation and sensitive detection of full-length N protein from crude cell lysates, demonstrating its selectivity and utility in biochemical applications. We further designed a chemically modified DNA aptamer and used it as a probe to examine the interaction of N-NTD with various RNA motifs, which revealed a strong preference for uridine-rich sequences. Our studies provide a high-affinity chemical probe for the SARS-CoV-2 N protein RNA-binding domain, which may be useful for diagnostic applications and investigating novel antiviral agents.

The emergence of the calvarial hematopoietic niche in health and disease.

The diploë region of skull has recently been discovered to act as a myeloid cell reservoir to the underlying meninges. The presence of ossified vascular channels traversing the inner skull of cortex provides a passageway for the cells to traffic from the niche, and CNS-derived antigens traveling through cerebrospinal fluid in a perivascular manner reaches the niche to signal myeloid cell egress. This review will highlight the recent findings establishing this burgeoning field along with the known role this niche plays in CNS aging and disease. It will further highlight the anatomical routes and physiological properties of the vascular structures these cells use for trafficking, spanning from skull to brain parenchyma.

Activation of ATF3 via the integrated stress response pathway regulates innate immune response to restrict Zika virus.

Zika virus (ZIKV) is a re-emerging mosquito-borne flavivirus that can have devastating health consequences. The developmental and neurological effects of a ZIKV infection arise in part from the virus triggering cellular stress pathways and perturbing transcriptional programs. To date, the underlying mechanisms of transcriptional control directing viral restriction and virus-host interaction are understudied. Activating Transcription Factor 3 (ATF3) is a stress-induced transcriptional effector that modulates the expression of genes involved in a myriad of cellular processes, including inflammation and antiviral responses, to restore cellular homeostasis. While ATF3 is known to be upregulated during ZIKV infection, the mode by which ATF3 is activated, and the specific role of ATF3 during ZIKV infection is unknown. In this study, we show via inhibitor and RNA interference approaches that ZIKV infection initiates the integrated stress response pathway to activate ATF4 which in turn induces ATF3 expression. Additionally, by using CRISPR-Cas9 system to delete ATF3, we found that ATF3 acts to limit ZIKV gene expression in A549 cells. We also determined that ATF3 enhances the expression of antiviral genes such as STAT1 and other components in the innate immunity pathway to induce an ATF3-dependent anti-ZIKV response. Our study reveals crosstalk between the integrated stress response and innate immune response pathways and highlights an important role for ATF3 in establishing an antiviral effect during ZIKV infection. IMPORTANCE: Zika virus (ZIKV) is a re-emerging mosquito-borne flavivirus that co-opts cellular mechanisms to support viral processes that can reprogram the host transcriptional profile. Such viral-directed transcriptional changes and the pro- or anti-viral outcomes remain understudied. We previously showed that ATF3, a stress-induced transcription factor, is significantly upregulated in ZIKV-infected mammalian cells, along with other cellular and immune response genes. We now define the intracellular pathway responsible for ATF3 activation and elucidate the impact of ATF3 expression on ZIKV infection. We show that during ZIKV infection, the integrated stress response pathway stimulates ATF3 which enhances the innate immune response to antagonize ZIKV infection. This study establishes a link between viral-induced stress response and transcriptional regulation of host defense pathways and thus expands our knowledge of virus-mediated transcriptional mechanisms and transcriptional control of interferon-stimulated genes during ZIKV infection.

Aspergillus fumigatus transcription factor ZfpA regulates hyphal development and alters susceptibility to antifungals and neutrophil killing during infection.

Hyphal growth is essential for host colonization during Aspergillus infection. The transcription factor ZfpA regulates A. fumigatus hyphal development including branching, septation, and cell wall composition. However, how ZfpA affects fungal growth and susceptibility to host immunity during infection has not been investigated. Here, we use the larval zebrafish-Aspergillus infection model and primary human neutrophils to probe how ZfpA affects A. fumigatus pathogenesis and response to antifungal drugs in vivo. ZfpA deletion promotes fungal clearance and attenuates virulence in wild-type hosts and this virulence defect is abrogated in neutrophil-deficient zebrafish. ZfpA deletion also increases susceptibility to human neutrophils ex vivo while overexpression impairs fungal killing. Overexpression of ZfpA confers protection against the antifungal caspofungin by increasing chitin synthesis during hyphal development, while ZfpA deletion reduces cell wall chitin and increases caspofungin susceptibility in neutrophil-deficient zebrafish. These findings suggest a protective role for ZfpA activity in resistance to the innate immune response and antifungal treatment during A. fumigatus infection.

Frequent first-trimester pregnancy loss in rhesus macaques infected with African-lineage Zika virus.

In the 2016 Zika virus (ZIKV) pandemic, a previously unrecognized risk of birth defects surfaced in babies whose mothers were infected with Asian-lineage ZIKV during pregnancy. Less is known about the impacts of gestational African-lineage ZIKV infections. Given high human immunodeficiency virus (HIV) burdens in regions where African-lineage ZIKV circulates, we evaluated whether pregnant rhesus macaques infected with simian immunodeficiency virus (SIV) have a higher risk of African-lineage ZIKV-associated birth defects. Remarkably, in both SIV+ and SIV- animals, ZIKV infection early in the first trimester caused a high incidence (78%) of spontaneous pregnancy loss within 20 days. These findings suggest a significant risk for early pregnancy loss associated with African-lineage ZIKV infection and provide the first consistent ZIKV-associated phenotype in macaques for testing medical countermeasures.

Variants in the Late Cornified Envelope Gene Locus Are Associated With Elevated T-helper 17 Responses in Patients With Postinfectious Lyme Arthritis.

BACKGROUND: Postinfectious Lyme arthritis (LA) is associated with dysregulated immunity and autoreactive T- and B-cell responses in joints. Here we explored the role of host genetic variation in this outcome. METHODS: The frequency of 253 702 single-nucleotide polymorphisms (SNPs) was determined in 147 patients with LA (87 with postinfectious LA and 60 with antibiotic-responsive LA), and for comparison in 90 patients with erythema migrans or the general population (n = 2504). Functional outcome of candidate SNPs was assessed by evaluating their impact on clinical outcome and on immune responses in blood and synovial fluid in patients with LA. RESULTS: Six SNPs associated with late cornified envelope (LCE3) genes were present at greater frequency in patients with postinfectious LA compared to those with antibiotic-responsive LA (70% vs 30%; odds ratio, 2; P < .01). These SNPs were associated with heightened levels of inflammatory Th17 cytokines in serum but lower levels of interleukin 27, a regulatory cytokine, implying that they may contribute to dysregulated Th17 immunity in blood. Moreover, in patients with postinfectious LA, the levels of these Th17 mediators correlated directly with autoantibody responses in synovial fluid, providing a possible link between LCE3 SNPs, maladaptive systemic Th17 immunity, and autoreactive responses in joints. CONCLUSIONS: Variation in the LCE3 locus, a known genetic risk factor in psoriasis and psoriatic arthritis, is associated with dysregulated systemic Th17 immunity and heightened autoantibody responses in joints. These findings underscore the importance of host genetic predisposition and systemic Th17 immunity in the pathogenesis of postinfectious (antibiotic-refractory) Lyme arthritis.

Trans-epithelial fluid flow and mechanics of epithelial morphogenesis.

Active fluid transport across epithelial monolayers is emerging as a major driving force of tissue morphogenesis in a variety of healthy and diseased systems, as well as during embryonic development. Cells use directional transport of ions and osmotic gradients to drive fluid flow across the cell surface, in the process also building up fluid pressure. The basic physics of this process is described by the osmotic engine model, which also underlies actin-independent cell migration. Recently, the trans-epithelial fluid flux and the hydraulic pressure gradient have been explicitly measured for a variety of cellular and tissue model systems across various species. For the kidney, it was shown that tubular epithelial cells behave as active mechanical fluid pumps: the trans-epithelial fluid flux depends on the hydraulic pressure difference across the epithelial layer. When a stall pressure is reached, the fluid flux vanishes. Hydraulic forces generated from active fluid pumping are important in tissue morphogenesis and homeostasis, and could also underlie multiple morphogenic events seen in other developmental contexts. In this review, we highlight findings that examined the role of trans-epithelial fluid flux and hydraulic pressure gradient in driving tissue-scale morphogenesis. We also review organ pathophysiology due to impaired fluid pumping and the loss of hydraulic pressure sensing at the cellular scale. Finally, we draw an analogy between cellular fluidic pumps and a connected network of water pumps in a city. The dynamics of fluid transport in an active and adaptive network is determined globally at the systemic level, and transport in such a network is best when each pump is operating at its optimal efficiency.

Structural and Functional Characterization of a Novel Class A Flavin Monooxygenase from Bacillus niacini.

A gene cluster responsible for the degradation of nicotinic acid (NA) in Bacillus niacini has recently been identified, and the structures and functions of the resulting enzymes are currently being evaluated to establish pathway intermediates. One of the genes within this cluster encodes a flavin monooxygenase (BnFMO) that is hypothesized to catalyze a hydroxylation reaction. Kinetic analyses of the recombinantly purified BnFMO suggest that this enzyme catalyzes the hydroxylation of 2,6-dihydroxynicotinic acid (2,6-DHNA) or 2,6-dihydroxypyridine (2,6-DHP), which is formed spontaneously by the decarboxylation of 2,6-DHNA. To understand the details of this hydroxylation reaction, we determined the structure of BnFMO using a multimodel approach combining protein X-ray crystallography and cryo-electron microscopy (cryo-EM). A liganded BnFMO cryo-EM structure was obtained in the presence of 2,6-DHP, allowing us to make predictions about potential catalytic residues. The structural data demonstrate that BnFMO is trimeric, which is unusual for Class A flavin monooxygenases. In both the electron density and coulomb potential maps, a region at the trimeric interface was observed that was consistent with and modeled as lipid molecules. High-resolution mass spectral analysis suggests that there is a mixture of phosphatidylethanolamine and phosphatidylglycerol lipids present. Together, these data provide insights into the molecular details of the central hydroxylation reaction unique to the aerobic degradation of NA in Bacillus niacini.

Hypertension increases sympathetic neuron activity by enhancing intraganglionic cholinergic collateral connections.

Autonomic dysregulation, including sympathetic hyperactivity, is a common feature of hypertension (HT) and other cardiovascular diseases. The CNS plays a role in driving chronic sympathetic activation in disease, but several lines of evidence suggest that neuroplasticity in the periphery may also contribute. The potential contribution of postganglionic sympathetic neurons to sustained sympathetic hyperactivity is not well understood. We recently discovered that noradrenergic sympathetic neurons in the stellate ganglion (SG) have excitatory cholinergic collateral connections to other neurons within the ganglion. We hypothesize that remodelling of these neurons and increased cholinergic collateral transmission contributes to sustained sympathetic hyperactivity in cardiovascular diseases, including HT. To test that hypothesis, we examined the activity of sympathetic neurons in isolated SG under control conditions and after 1 week of HT induced by peripheral angiotensin II infusion, using whole-cell patch clamp recordings. Despite the absence of central inputs, we observed elevated spontaneous activity and synaptic transmission in sympathetic SG neurons from hypertensive mice that required generation of action potentials. Genetically disrupting cholinergic transmission in noradrenergic neurons decreased basal neuronal activity and prevented angiotensin II-mediated enhancement of activity. Similar changes in activity, driven by increased collateral transmission, were identified in cardiac projecting neurons and neurons projecting to brown adipose tissue. These changes were not driven by altered A-type K+ currents. This suggests that HT stimulates increased activity throughout the intraganglionic network of collateral connections, contributing to the sustained sympathetic hyperactivity characteristic in cardiovascular disease. KEY POINTS: Sympathetic neurons in ganglia isolated from angiotensin II-treated hypertensive mice are more active than neurons from control mice despite the absence of central activation. The enhanced activity is the result of a ganglionic network of cholinergic collaterals, rather than altered intrinsic excitability. Increased neuronal activity was observed in both cardiac neurons and brown adipose tissue-projecting neurons, which are not involved in cardiovascular homeostasis.

Reciprocal regulatory balance within the CLEC16A-RNF41 mitophagy complex depends on an intrinsically disordered protein region.

CLEC16A is an E3 ubiquitin ligase that regulates mitochondrial quality control through mitophagy and is associated with over 20 human diseases. CLEC16A forms a complex with another E3 ligase, RNF41, and a ubiquitin-specific peptidase, USP8; however, regions that regulate CLEC16A activity or the assembly of the tripartite mitophagy regulatory complex are unknown. Here, we report that CLEC16A contains an internal intrinsically disordered protein region (IDPR) that is crucial for CLEC16A function and turnover. IDPRs lack a fixed secondary structure and possess emerging yet still equivocal roles in protein stability, interactions, and enzymatic activity. We find that the internal IDPR of CLEC16A is crucial for its degradation. CLEC16A turnover was promoted by RNF41, which binds and acts upon the internal IDPR to destabilize CLEC16A. Loss of this internal IDPR also destabilized the ubiquitin-dependent tripartite CLEC16A-RNF41-USP8 complex. Finally, the presence of an internal IDPR within CLEC16A was confirmed using NMR and CD spectroscopy. Together, our studies reveal that an IDPR is essential to control the reciprocal regulatory balance between CLEC16A and RNF41, which could be targeted to improve mitochondrial health in disease.

Heterogeneity of the group B streptococcal type VII secretion system and influence on colonization of the female genital tract.

Type VIIb secretion systems (T7SSb) in Gram-positive bacteria facilitate physiology, interbacterial competition, and/or virulence via EssC ATPase-driven secretion of small ɑ-helical proteins and toxins. Recently, we characterized T7SSb in group B Streptococcus (GBS), a leading cause of infection in newborns and immunocompromised adults. GBS T7SS comprises four subtypes based on variation in the C-terminus of EssC and the repertoire of downstream effectors; however, the intraspecies diversity of GBS T7SS and impact on GBS-host interactions remains unknown. Bioinformatic analysis indicates that GBS T7SS loci encode subtype-specific putative effectors, which have low interspecies and inter-subtype homology but contain similar domains/motifs and therefore may serve similar functions. We further identify orphaned GBS WXG100 proteins. Functionally, we show that GBS T7SS subtype I and III strains secrete EsxA in vitro and that in subtype I strain CJB111, esxA1 appears to be differentially transcribed from the T7SS operon. Furthermore, we observe subtype-specific effects of GBS T7SS on host colonization, as CJB111 subtype I but not CNCTC 10/84 subtype III T7SS promotes GBS vaginal colonization. Finally, we observe that T7SS subtypes I and II are the predominant subtypes in clinical GBS isolates. This study highlights the potential impact of T7SS heterogeneity on host-GBS interactions.

Stability-Based Proteomics for Investigation of Structured RNA-Protein Interactions.

RNA-protein interactions are essential to RNA function throughout biology. Identifying the protein interactions associated with a specific RNA, however, is currently hindered by the need for RNA labeling or costly tiling-based approaches. Conventional strategies, which commonly rely on affinity pull-down approaches, are also skewed to the detection of high affinity interactions and frequently miss weaker interactions that may be biologically important. Reported here is the first adaptation of stability-based mass spectrometry methods for the global analysis of RNA-protein interactions. The stability of proteins from rates of oxidation (SPROX) and thermal protein profiling (TPP) methods are used to identify the protein targets of three RNA ligands, the MALAT1 triple helix (TH), a viral stem loop (SL), and an unstructured RNA (PolyU), in LNCaP nuclear lysate. The 315 protein hits with RNA-induced conformational and stability changes detected by TPP and/or SPROX were enriched in previously annotated RNA-binding proteins and included new proteins for hypothesis generation. Also demonstrated are the orthogonality of the SPROX and TPP approaches and the utility of the domain-specific information available with SPROX. This work establishes a novel platform for the global discovery and interrogation of RNA-protein interactions that is generalizable to numerous biological contexts and RNA targets.

Hypertension-induced heart failure disrupts cardiac sympathetic innervation.

About 26 million people worldwide live with heart failure (HF), and hypertension is the primary cause in 25% of these cases. Autonomic dysfunction and sympathetic hyperactivity accompany cardiovascular diseases, including HF. However, changes in cardiac sympathetic innervation in HF are not well understood. We hypothesized that cardiac sympathetic innervation is disrupted in hypertension-induced HF. Male and female C57BL6/J mice were infused with Angiotensin II (AngII) for 4 weeks to generate hypertension leading to HF; controls were infused with saline. AngII-treated mice displayed HF phenotype including reduced cardiac function, hypertrophy, and fibrosis. AngII-treated mice also had significantly reduced sympathetic nerve density in the left ventricle, intraventricular septum, and right ventricle. In the left ventricle, the subepicardium remained normally innervated, while the subendocardium was almost devoid of sympathetic nerves. Loss of sympathetic fibers led to loss of norepinephrine content in the left ventricle. Several potential triggers for axon degeneration were tested and ruled out. AngII-treated mice had increased premature ventricular contractions after isoproterenol and caffeine injection. Although HF can induce a cholinergic phenotype and neuronal hypertrophy in stellate ganglia, AngII treatment did not induce a cholinergic phenotype or activation of trophic factors in this study. Cardiac neurons in the left stellate ganglion were significantly smaller in AngII-treated mice, while neurons in the right stellate were unchanged. Our findings show that AngII-induced HF disrupts sympathetic innervation, particularly in the left ventricle. Further investigations are imperative to unveil the mechanisms of denervation in HF and to develop neuromodulatory therapies for patients with autonomic imbalance.

Comparison of wild-type KT2440 and genome-reduced EM42 Pseudomonas putida strains for muconate production from aromatic compounds and glucose.

Pseudomonas putida KT2440 is a robust, aromatic catabolic bacterium that has been widely engineered to convert bio-based and waste-based feedstocks to target products. Towards industrial domestication of P. putida KT2440, rational genome reduction has been previously conducted, resulting in P. putida strain EM42, which exhibited characteristics that could be advantageous for production strains. Here, we compared P. putida KT2440- and EM42-derived strains for cis,cis-muconic acid production from an aromatic compound, p-coumarate, and in separate strains, from glucose. To our surprise, the EM42-derived strains did not outperform the KT2440-derived strains in muconate production from either substrate. In bioreactor cultivations, KT2440- and EM42-derived strains produced muconate from p-coumarate at titers of 45 g/L and 37 g/L, respectively, and from glucose at 20 g/L and 13 g/L, respectively. To provide additional insights about the differences in the parent strains, we analyzed growth profiles of KT2440 and EM42 on aromatic compounds as the sole carbon and energy sources. In general, the EM42 strain exhibited reduced growth rates but shorter growth lags than KT2440. We also observed that EM42-derived strains resulted in higher growth rates on glucose compared to KT2440-derived strains, but only at the lowest glucose concentrations tested. Transcriptomics revealed that genome reduction in EM42 had global effects on transcript levels and showed that the EM42-derived strains that produce muconate from glucose exhibit reduced modulation of gene expression in response to changes in glucose concentrations. Overall, our results highlight that additional studies are warranted to understand the effects of genome reduction on microbial metabolism and physiology, especially when intended for use in production strains.

Pain trajectories after bilateral orthotopic lung transplantation surgery performed via a clamshell incision.

INTRODUCTION: The nature, intensity, and progression of acute pain after bilateral orthotopic lung transplantation (BOLT) performed via a clamshell incision has not been well investigated. We aimed to describe acute pain after clamshell incisions using pain trajectories for the study cohort, in addition to stratifying patients into separate pain trajectory groups and investigating their association with donor and recipient perioperative variables. METHODS: After obtaining IRB approval, we retrospectively included all patients ≥18 years old who underwent primary BOLT via clamshell incision at a single center between January 1, 2017, and June 30, 2022. We modeled the overall pain trajectory using pain scores collected over the first seven postoperative days and identified separate pain trajectory classes via latent class analysis. RESULTS: Three hundred one adult patients were included in the final analysis. Three separate pain trajectory groups were identified, with most patients (72.8%) belonging to a well-controlled, stable pain trajectory. Uncontrolled pain was either observed in the early postoperative period (10%), or in the late postoperative period (17.3%). Late postoperative peaking trajectory patients were younger (p = .008), and sicker with a higher lung allocation score (p = .005), receiving preoperative mechanical ventilation (p < .001), or VV-ECMO support (p < .001). CONCLUSION: Despite the extensive nature of a clamshell incision, most pain trajectories in BOLT patients had a well-controlled stable pain profile. The benign nature of pain profiles in our patient population may be attributed to the routine institutional practice of early thoracic epidural analgesia for BOLT patients unless contraindicated.

Characterization of natural killer and cytotoxic T-cell immune infiltrates in pancreatic ductal adenocarcinoma.

BACKGROUND AND OBJECTIVES: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with poor response to systemic therapies, including immunotherapy. Given the immunotherapeutic potential of natural killer (NK) cells, we evaluated intratumoral NK cell infiltrates along with cytotoxic T cells in PDAC to determine their association with patient outcomes. METHODS: We analyzed tumors from 93 PDAC patients treated from 2012 to 2020. Predictor variables included tumor-infiltrating lymphocytes (TILs), T-cell markers (CD3, CD8, CD45RO), NK marker (NKp46), and NK inhibitory marker (major histocompatibility complex class I [MHC-I]) by immunohistochemistry. Primary outcome variables were recurrence-free survival (RFS) and overall survival (OS). RESULTS: Mean TILs, CD3, and NKp46 scores were 1.3 ± 0.63, 20.6 ± 17.5, and 3.1 ± 3.9, respectively. Higher expression of CD3 and CD8 was associated with higher OS, whereas NK cell infiltration was not associated with either RFS or OS. There was a tight positive correlation between MHC-I expression and all T-cell markers, but not with NKp46. CONCLUSIONS: Overall NK cell infiltrates were low in PDAC and did not predict clinical outcomes, whereas T-cell infiltrates did. Further characterization of the immune infiltrate in PDAC, including inhibitory signals and suppressive cell types, may yield better biomarkers of prognosis and immune targeting in this refractory disease.

Differential Roles for IL-1α and IL-1ß in Pseudomonas aeruginosa Corneal Infection.

Pseudomonas aeruginosa is an important cause of dermal, pulmonary, and ocular disease. Our studies have focused on P. aeruginosa infections of the cornea (keratitis) as a major cause of blinding microbial infections. The infection leads to an influx of innate immune cells, with neutrophils making up to 90% of recruited cells during early stages. We previously reported that the proinflammatory cytokines IL-1α and IL-1ß were elevated during infection. Compared with wild-type (WT), infected Il1b-/- mice developed more severe corneal disease that is associated with impaired bacterial killing as a result of defective neutrophil recruitment. We also reported that neutrophils are an important source of IL-1α and IL-1ß, which peaked at 24 h postinfection. To examine the role of IL-1α compared with IL-1ß in P. aeruginosa keratitis, we inoculated corneas of C57BL/6 (WT), Il1a-/-, Il1b-/-, and Il1a-/-Il1b-/- (double-knockout) mice with 5 × 104 ExoS-expressing P. aeruginosa. Il1b-/- and double-knockout mice have significantly higher bacterial burden that was consistent with delayed neutrophil and monocyte recruitment to the corneas. Surprisingly, Il1a-/- mice had the opposite phenotype with enhanced bacteria clearance compared with WT mice. Although there were no significant differences in neutrophil recruitment, Il1a-/- neutrophils displayed a more proinflammatory transcriptomic profile compared to WT with elevations in C1q expression that likely caused the phenotypic differences observed. To our knowledge, our findings identify a novel, non-redundant role for IL-1α in impairing bacterial clearance.

Animal models for partial heart transplantation.

BACKGROUND: Partial heart transplantation delivers growing heart valve implants by transplanting the part of the heart containing the necessary heart valve only. In contrast to heart transplantation, partial heart transplantation spares the native ventricles. This has important implications for partial heart transplant biology, including the allowable ischemia time, optimal graft preservation, primary graft dysfunction, immune rejection, and optimal immunosuppression. AIMS: Exploration of partial heart transplant biology will depend on suitable animal models. Here we review our experience with partial heart transplantation in rodents, piglets, and non-human primates. MATERIALS & METHODS: This review is based on our experience with partial heart transplantation using over 100 rodents, over 50 piglets and one baboon. RESULTS: Suitable animal models for partial heart transplantation include rodent heterotopic partial heart transplantation, piglet orthotopic partial heart transplantation, and non-human primate partial heart xenotransplantation. DISCUSSION: Rodent models are relatively cheap and offer extensive availability of research tools. However, rodent open-heart surgery is technically not feasible. This limits rodents to heterotopic partial heart transplant models. Piglets are comparable in size to children. This allows for open-heart surgery using clinical grade equipment for orthoptic partial heart transplantation. Piglets also grow rapidly, which is useful for studying partial heart transplant growth. Finally, nonhuman primates are immunologically most closely related to humans. Therefore, nonhuman primates are most suitable for studying partial heart transplant immunobiology and xenotransplantation. CONCLUSIONS: Animal research is a privilege that is contingent on utilitarian ethics and the 3R principles of replacement, reduction and refinement. This privilege allows the research community to seek fundamental knowledge about partial heart transplantation, and to apply this knowledge to enhance the health of children who require partial heart transplants.