Faculty Bibliography

Microsporidia are single-celled intracellular parasites that cause opportunistic diseases in humans. Encephalitozoon intestinalis is a prevalent human-infecting species that invades the small intestine. Macrophages are potential reservoirs of infection, and dissemination to other organ systems is also observed. The macrophage response to infection and the developmental trajectory of the parasite are not well studied. Here we use single cell RNA sequencing to investigate transcriptional changes in both the parasite and the host during E. intestinalis infection of human macrophages in vitro. The parasite undergoes large transcriptional changes throughout the life cycle, providing a blueprint for parasite development. While a small population of infected macrophages mount a response, most remain transcriptionally unchanged, suggesting that the majority of parasites may avoid host detection. The stealthy microsporidian lifestyle likely allows these parasites to harness macrophages for replication. Together, our data provide insights into the host response in primary human macrophages and the E. intestinalis developmental program.

The continuity of a species depends on germ cells. Germ cells are different from all the other cell types of the body (somatic cells) as they are solely destined to develop into gametes (sperm or egg) to create the next generation. In this review, we will touch on 4 areas of embryonic germ cell development in Drosophila melanogaster: the assembly and function of germplasm, which houses the determinants for germ cell specification and fate and the mitochondria of the next generation; the process of pole cell formation, which will give rise to primordial germ cells (PGCs); the specification of pole cells toward the PGC fate; and finally, the migration of PGCs to the somatic gonadal precursors, where they, together with somatic gonadal precursors, form the embryonic testis and ovary.

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.

Astrocytes are a highly abundant glial cell type and perform critical homeostatic functions in the central nervous system. Like neurons, astrocytes have many discrete heterogeneous subtypes. The subtype identity and functions are, at least in part, associated with their anatomical location and can be highly restricted to strategically important anatomical domains. Here, we report that astrocytes forming the glia limitans superficialis, the outermost border of the brain and spinal cord, are a highly specialized astrocyte subtype and can be identified by a single marker: myocilin (Myoc). We show that glia limitans superficialis astrocytes cover the entire brain and spinal cord surface, exhibit an atypical morphology, and are evolutionarily conserved from zebrafish, rodents, and non-human primates to humans. Identification of this highly specialized astrocyte subtype will advance our understanding of CNS homeostasis and potentially be targeted for therapeutic intervention to combat peripheral inflammatory effects on the CNS.

High-dimensional multiplexed imaging can reveal the spatial organization of tumour tissues at the molecular level. However, owing to the scale and information complexity of the imaging data, it is challenging to discover and thoroughly characterize the heterogeneity of tumour microenvironments. Here we show that self-supervised representation learning on data from imaging mass cytometry can be leveraged to distinguish morphological differences in tumour microenvironments and to precisely characterize distinct microenvironment signatures. We used self-supervised masked image modelling to train a vision transformer that directly takes high-dimensional multiplexed mass-cytometry images. In contrast with traditional spatial analyses relying on cellular segmentation, the vision transformer is segmentation-free, uses pixel-level information, and retains information on the local morphology and biomarker distribution. By applying the vision transformer to a lung-tumour dataset, we identified and validated a monocytic signature that is associated with poor prognosis.

Embryonic development in many species, including case reports in humans, can be temporarily halted before implantation during a process called diapause. Facultative diapause occurs under conditions of maternal metabolic stress such as nursing. While molecular mechanisms of diapause have been studied, a natural inducing factor has yet to be identified. Here, we show that oxytocin induces embryonic diapause in mice. We show that gestational delays were triggered during nursing or optogenetic stimulation of oxytocin neurons simulating nursing patterns. Mouse blastocysts express oxytocin receptors, and oxytocin induced delayed implantation-like dispersion in cultured embryos. Last, oxytocin receptor-knockout embryos transferred into wild-type surrogates had low survival rates during diapause. Our results indicate that oxytocin coordinates timing of embryonic development with uterine progression through pregnancy, providing an evolutionarily conserved mechanism for ensuring successful reproduction.

INTRODUCTION/UNASSIGNED:Alkyl nitrites ("poppers") are used recreationally for sexual enhancement, muscle relaxation, and euphoria. However, they can be toxic and cause adverse reactions such as methemoglobinemia. While inhalation is the typical route of usage, the New York City Poison Center has noted an increase in calls related to ingestion. Given the uncertainty of usage instructions at the point of sale, our study aimed to assess instructions provided to consumers about alkyl nitrite use and to evaluate the proximity and co-sale of alkyl nitrites with similarly appearing energy drink shots. METHODS/UNASSIGNED:We conducted a cross-sectional convenience sample survey of smoke shops, cannabis dispensaries, and exotic shops within the catchment area of an urban poison center. Plain clothes "investigators" (i.e., the researchers) visited these retailers and followed a predetermined protocol and script to request information regarding the availability and usage of alkyl nitrites. Additionally, the researchers attempted to visually assess the proximity of alkyl nitrites to similarly appearing energy drink shots during their visit. RESULTS/UNASSIGNED:drinks and alkyl nitrites were located near these energy drink shots in twenty (39%) of these fifty-one stores. DISCUSSION/UNASSIGNED:Many commercial alkyl nitrite retailers in our survey area lack knowledge or provide potentially inaccurate information regarding the use of alkyl nitrites. Additionally, alkyl nitrites are often sold alongside commercial energy drinks, potentially increasing the risk of incidental ingestion. CONCLUSIONS/UNASSIGNED:Further research is necessary to determine the impact of these patterns of sale and potential misinformation. Discussing preliminary results of our survey with the New York City Department of Health led to the rapid creation of an informational poster and local outreach. Clinicians should report cases of alkyl nitrite use to their regional poison center to allow for more targeted and timely public health intervention.

Microsporidia are divergent fungal pathogens that employ a unique harpoon-like apparatus called the polar tube (PT) to invade host cells. The long PT is fired out of the microsporidian spore over the course of just a few hundred milliseconds. Once fired, the PT is thought to pierce the plasma membrane of a target cell and act as a conduit for the transfer of the parasite into the host cell, which initiates infection. The PT architecture and its association with neighboring organelles within the parasite cell remain poorly understood. Here, we use cryoelectron tomography to investigate the structural cell biology of the PT in dormant spores from the human-infecting microsporidian species, Encephalitozoon intestinalis. Segmentation and subtomogram averaging of the PT reveal at least four layers: two protein-based layers surrounded by a membrane layer and filled with a dense core. Regularly spaced protein filaments form the structural skeleton of the PT. Combining cryoelectron tomography with cellular modeling, we propose a model for the three-dimensional organization of the polaroplast, an organelle that surrounds the PT and is continuous with the outermost, membranous layer of the PT. Our results reveal the ultrastructure of the microsporidian invasion apparatus in situ, laying the foundation for understanding infection mechanisms.