Faculty Bibliography

Pathogens have evolved to be highly adapted to their natural host. Community-associated methicillin-resistant Staphylococcus aureus USA300, for instance, is a lineage responsible for the epidemic of skin and soft tissue infections (SSTIs) in humans. Owing to its human tropism, mechanisms that enabled the rise of USA300 as a major skin pathogen remain incompletely defined. By leveraging a rodent-adapted strain of S. aureus, we developed a natural model of SSTIs. We found that LukMF', a pore-forming leukocidin homolog to the human-specific LukSF-PV toxin, drives skin pathology in mice. LukMF' lyses neutrophils via the chemokine receptor CCR1, which in turn fuels inflammatory pathology and microbial survival within the infectious nidus. Ablation of CCR1, depletion of neutrophils, or vaccination with LukMF' all protected mice from skin pathology. Thus, these data support epidemiological studies linking leukocidins with human SSTIs and highlight the power of natural models to unearth potential targets to curtail infections.

Skin, hair, and eye (oculocutaneous) color is due to melanin, a pigment produced by melanocytes. This review considers processes required for pigmentation and the complex genetic network that regulates them. The first requisite is migration of neural crest-derived melanoblasts, which populate various embryonic sites, then differentiate into melanocytes or seed stem cell niches. Differentiation is marked by expression of genes essential for melanogenesis, which takes place in melanosomes and involves conversion of tyrosine into melanin. Melanosome biogenesis requires premelanosome maturation through coordinated delivery of melanogenic enzymes such as tyrosinase (TYR), structural proteins, and transporters that establish an intraluminal environment conducive to melanogenesis. Sorting of proteins through endolysosomal pathways and delivery to melanosomes is facilitated by trafficking protein complexes. Finally, melanin is transferred to keratinocytes to protect against ultraviolet light. Numerous pigment-related disorders result from disruption of these pathways, including Waardenburg syndrome caused by melanoblast migration disruption, oculocutaneous albinism presenting with absent/reduced melanogenesis, and melanoma resulting from dysregulation of proliferation/survival. Genetic variants also determine normal color variation, which is pronounced across populations that, historically, lived in different geographical regions. This variation, shaped by genetic factors, environmental influences, and evolutionary pressures, underpins the wide range of pigmentation phenotypes seen today.

After birth, tissues grow until they reach adult size, with each organ exhibiting unique cellular dynamics, growth patterns, and stem or non-stem cell sources. Using multiscale experimental and computational approaches, we found that aortic enlargement follows distinct growth principles, scaling with the vertebral column. Expansion proceeds via two temporally coordinated, spatially stochastic waves of proliferation aligned with blood flow, each with unique cell-cycle kinetics, with the first wave featuring cycles as short as 6 h. Single-cell RNA sequencing revealed increased fatty acid metabolism accompanying cell enlargement. Mathematical modeling and experiments showed that endothelial cell extrusion is essential for maintaining homeostatic aortic size as it adjusts for proliferation excess. Using a genetic model of achondroplasia, we mechanistically demonstrated that the aorta preserves proper scaling by increasing cell extrusion while keeping proliferation rates intact. These findings provide a blueprint of the principles orchestrating aortic growth, which relies entirely on the proliferation of resident differentiated cells. A record of this paper's transparent peer review process is included in the supplemental information.

AIM/OBJECTIVE:To investigate how psychosocial stress contributes to accelerated thrombosis, focusing on platelet activation and hyperreactivity. The specific objective was to identify novel platelet regulators involved in stress-mediated thrombosis, with a particular emphasis on the tetraspanin CD37. METHODS AND RESULTS/RESULTS:To explore how stress contributes to platelet hyperreactivity, platelets were isolated from (1) mice that experienced chronic variable stress and stress-free controls (n=8/group) and (2) human subjects with self-reported high and no stress levels (n=18/group), followed by RNA-sequencing. By comparing mutually expressed transcripts, a subset of genes differentially expressed following psychosocial stress was identified in both human and mouse platelets. In both mice and humans, platelet CD37 positively associates with platelet aggregation responses that underlie thrombosis, with Cd37-/- platelets exhibiting impaired integrin αIIbβ3 signaling, characterized by reduced platelet fibrinogen spreading and decreased agonist-induced αIIbβ3 activation. Consistent with a role for CD37 in regulating platelet activation responses, chimeric mice that received Cd37-/- bone marrow experienced a significantly increased time to vessel occlusion in the carotid artery FeCl3 model compared to mice reconstituted with wild-type bone marrow. CD37 deficiency did not alter hemostasis, as platelet count, coagulation metrics, prothrombin time, and partial thromboplastin time did not differ in Cd37-/- mice relative to wild-type mice. Consistent with this, bleeding time did not differ between wild-type and Cd37-/- mice following tail tip transection. CONCLUSIONS:This study provides new insights into the platelet-associated mechanisms underlying stress-mediated thrombosis. Identifying CD37 as a novel regulator of platelet activation responses offers potential therapeutic targets for reducing the thrombotic risk associated with psychosocial stress. The findings also contribute to understanding how psychosocial stress accelerates thrombotic events and underscore the importance of platelet activation in this process.

Atherosclerotic cardiovascular diseases (ASCVD) remain the leading cause of death globally. Animal and human studies link psychological stress-related disorders to ASCVD. Despite this accumulating evidence linking stress to increased cardiovascular disease (CVD) risk, it remains unclear whether stress impairs the benefits of standard risk-reduction therapies, of which lipid-lowering remains the most common, or whether this increased risk is driven by systemic inflammatory states. We tested the hypothesis that psychological stress limits the benefits of lipid lowering on resolving inflammation in atherosclerotic plaques by combining two established mouse models, namely one in which levels of atherogenic LDL cholesterol (LDL-C) can be lowered after plaques develop, and the other a model of chronic social defeat stress (CSDS). Here we show that mice susceptible to CSDS ("SUS") had attenuated benefits of LDL-C lowering compared to control (CON) or resilient (RES) mice. Moreover, in SUS mice (vs. CON or RES) there was heightened inflammation in the plaque macrophages, with evidence that this was a result of re-programming in the bone marrow (BM) of the precursors of macrophages, namely monocytes. Remarkably, these observations aligned with human imaging studies, in which LDL-C lowering therapy was not as effective in reducing either systemic or arterial inflammation in subjects with higher (vs. lower) neural imaging measures of psychological stress. In summary, the integration of the mouse model and human data provides important mechanistic and clinical insights into the crucial role of chronic stress in ASCVD, highlighting that this common risk factor impairs the anti-atherosclerotic benefits of lipid-lowering medications and may represent an important determinant of residual ASCVD risk.

The intestinal secretory lineage is thought to comprise four distinct cell types derived from one Atoh1⁺ progenitor, but the mechanisms that distinguish Paneth and goblet cells are unclear. Bhattacharya et al.¹ argue that these cells are instead phenotypic manifestations of a common terminal Atoh1⁺ cell, actively shaped by niche-derived signals.

BACKGROUND:/calmodulin-dependent protein kinase II and selected for probe collection. RESULTS:This approach significantly reduced the amount of FFPE tissue needed for robust single population RNA-seq. We demonstrate ~20 identified L3 or L5 pyramidal neurons or one lamina-specific cortical ribbon from a single 5µm thick section is sufficient to generate robust RNA-seq reads. Bioinformatic analysis of neurons and ribbons showed notable similarities and differences reflective of the single neuron and multiple admixed cell types within the former and latter, respectively. Comparison with existing methods Protocols exist for DSP of postmortem human FFPE brain tissue. However, this new approach enables profiling small groups of ~14-21 pyramidal neurons using the GeoMx DSP platform. CONCLUSIONS:This optimized DSP assay provides high resolution RNA-seq data demonstrating utility and versatility of the GeoMx platform for individually characterized neurons and isolated cortical ribbons within postmortem FFPE human brain tissue for downstream analyses.

Alzheimer's disease (AD) is characterized by the deposition of full-length and truncated amyloid beta (Aβ) species within brain parenchyma and cerebral vessels. However, Aβ truncated species remain understudied. Here, we present a protocol for culture and characterization of a mouse monoclonal antibody targeting N-terminally truncated proteoforms starting at position 4. We describe a detailed procedure for hybridoma culture, antibody collection, and isolation via affinity chromatography. We then describe steps for target validation via dot blot, as well as potential applications. For complete details on the use and execution of this protocol, please refer to Cabrera et al. and Rostagno et al.¹

BACKGROUND:Synovial fluid (SF) biomarkers demonstrate time-dependent variation after acute knee injury, and it is postulated that persistently elevated inflammatory markers may mediate worse long-term outcomes. PURPOSE/OBJECTIVE:This study investigated the relationship between biomarkers in SF at the time of meniscectomy and long-term patient-reported outcomes in patients with acute versus chronic meniscal injuries. STUDY DESIGN/METHODS:Cohort study; Level of evidence, 3. METHODS:This retrospective analysis included patients who underwent knee SF aspiration on the day of arthroscopic meniscectomy between October 2011 and October 2020 with minimum 4-year follow-up. SF aspirated from the operative knee was analyzed for 10 pro- and anti-inflammatory biomarkers. Patients completed the visual analog scale for pain, Lysholm Knee Questionnaire, Tegner Activity Scale, and Knee injury and Osteoarthritis Outcome Score-Physical Function Short-form (KOOS-PS) before surgery and at follow-up. Patients were categorized as having acute (<6 weeks) or chronic (>1 year) symptoms. K-means clustering analysis was performed using biomarker levels to group patients into distinct cohorts. RESULTS:= .020) than the low-inflammation cohort. CONCLUSION/CONCLUSIONS:In patients with chronic meniscal injury, those with a more proinflammatory SF biomarker profile at the time of meniscectomy had worse outcomes than those who had a low inflammatory profile. In acute meniscal injuries, most patients demonstrate a high inflammatory profile, which was not associated with a difference in long-term outcomes.