Biology subjects

Curtis, B. C.

Publications and source records attributed to Curtis, B. C..

6 recordsLinked to original sources

bioRxiv2026

A single-cell transcriptomic atlas of the periventricular proliferative zone in the late gestation fetal brain in the pigtail macaque

BackgroundThe fetal brain undergoes rapid changes in late gestation, when waves of neurogenesis and gliogenesis shape cortical circuitry. The periventricular proliferative region and adjacent white matter are enriched in neuroprogenitor cells, newborn neurons, and interneurons, which is challenging to study in the late gestation human fetal brain. The nonhuman primate (NHP) provides a powerful translational model to overcome this limitation, given its close similarity to human neurodevelopmental trajectories. The study objective was to construct a single-cell RNA-Seq (scRNA-Seq) atlas of the late-gestation fetal brain of the pigtail macaque (Macaca nemestrina), focused on the periventricular proliferative zone. MethodsA sample of the lateral ventricular wall, subventricular zone, and overlying white/gray matter was dissociated into single cells and processed through the 10X Genomics pipeline, followed by SoupX removal of ambient RNA, and Seurats pipeline to aggregate, cluster and annotate single-cell populations. Monocle3 was used to determine pseudotime and map lineage progression. ResultsThis analysis captured diverse populations of neuroprogenitors, newborn neurons, developing lineages of excitatory and inhibitory neurons, oligodendrocyte and astrocyte lineages, and resident immune and endothelial cells. ConclusionsSingle-cell populations from the third-trimester nonhuman primate fetal brain are highly similar to those in the human fetus. This late-gestation single-cell atlas of the periventricular proliferative zone provides a unique reference for progenitor, neuronal, glial, vascular, and immune cell states during a critical window of primate neurodevelopment, enabling mechanistic interrogation of how inflammatory, infectious, or hypoxic insults disrupt vulnerable neurogenic niches.

genomics↗

bioRxiv2025

Maternal Influenza A Virus Infection Induces Antiviral and Immune Dysregulation in the Placenta and Fetus Without Vertical Transmission

Influenza A virus (IAV) infection during pregnancy is associated with stillbirth and preterm birth, but the degree to which IAV alters placental and fetal immunity is poorly understood. The objective of our study was to determine the immunologic impact of maternal IAV infection on the placenta and fetus in a pigtail macaque (Macaca nemestrina) model. Pregnant pigtail macaques were inoculated with 107 plaque forming units (PFU) of IAV [A/California/07/2009 (H1N1)] and underwent necropsy 5 days post-infection (N=11). Results were compared to uninfected historical controls (N=16). IAV inoculation induced maternal pneumonia in all cases. Stillbirth occurred in 18% (2/11) of IAV-infected pregnancies, but not in controls. While vertical transmission was not observed, low-level IAV viral RNA was detected in two placentas. In the placenta, maternal IAV infection was associated with increased IL-1{beta}, IL-18, and IFN-{beta} levels, and an upregulated type I interferon (IFN) transcriptional response. IAV infection was also associated with significantly higher frequencies of intermediate and non-classical monocytes, plasmacytoid dendritic cells, CD4 T cells, and NKT cells in the fetus (lung, lymph node, blood). Although placental immune and transcriptional perturbations were rarely correlated with maternal IAV disease indicators (e.g., maternal lung viral load/IFN-/IFN-{beta}/IL-6), there were consistent and significant correlations between these metrics and perturbed immune cell populations in the fetus (CD4+ and CD8+ T cells, plasmacytoid dendritic cells, monocyte sub-populations). Maternal IAV infection disrupted both placental and fetal immune environments, but only fetal immune alterations correlated with maternal lung disease severity. One Sentence SummaryMaternal influenza A virus infection in pregnant pigtail macaques dysregulates placental and fetal immunity, with disease severity correlating strongly with fetal, but not placental, immune perturbations.

systems biology↗

bioRxiv2025

A single-cell transcriptomic atlas of the pigtail macaque placenta in late gestation

The placenta is a complex organ with multiple immune and non-immune cell types that promote fetal tolerance and facilitate the transfer of nutrients and oxygen. The nonhuman primate (NHP) is a key experimental model for studying human pregnancy complications, in part due to similarities in placental structure, which makes it essential to understand how single-cell populations compare across the human and NHP maternal-fetal interface. We constructed a single-cell RNA-Seq (scRNA-Seq) atlas of the placenta from the pigtail macaque (Macaca nemestrina) in the third trimester, comprising three different tissues at the maternal-fetal interface: the chorionic villi (placental disc), chorioamniotic membranes, and the maternal decidua. Each tissue was separately dissociated into single cells and processed through the 10X Genomics and Seurat pipeline, followed by aggregation, unsupervised clustering, and cluster annotation. Next, we determined the maternal-fetal origins of cell populations and analyzed single-cell RNA trajectory, Gene Ontology enrichment, and cell-cell communication. Single-cell populations in the pigtail macaque were strikingly similar in their identity and frequency to those found in the human placenta, including cells from trophoblast, stromal cell, immune, and macrophage lineages. An advantage of our approach was the deep sequencing of three tissues at the maternal-fetal interface, which yielded a rich diversity of common and rare single-cell populations. The third-trimester pigtail macaque single-cell atlas enables the identification of cellular subclusters analogous to those in humans and provides a powerful resource for understanding experimental perturbations on the NHP placenta.

genomics↗

bioRxiv2024

Enhancer AAVs for targeting spinal motor neurons and descending motor pathways in rodents and macaque

Experimental access to cell types within the mammalian spinal cord is severely limited by the availability of genetic tools. To enable access to lower motor neurons (LMNs) and LMN subtypes, we generated single cell multiome datasets from mouse and macaque spinal cords and discovered putative enhancers for each neuronal population. We cloned these enhancers into adeno-associated viral vectors (AAVs) driving a reporter fluorophore and functionally screened them in mouse. We extensively characterized the most promising candidate enhancers in rat and macaque and developed an optimized pan LMN enhancer virus. Additionally, we generated derivative viruses expressing iCre297T recombinase or ChR2-EYFP for labeling and functional studies, and we created a single vector with combined enhancer elements to achieve simultaneous labeling of layer 5 extratelencephalic projecting (ET) neurons and LMNs. This unprecedented LMN toolkit will enable future investigations of cell type function across species and potential therapeutic interventions for human neurodegenerative diseases.

neuroscience↗

bioRxiv2024

Genome-wide CRISPRa screens nominate modulators of CAR T cell survival within distinct tumor cytokine milieus

Chimeric Antigen Receptor (CAR) T cell therapy has revolutionized the treatment of B cell malignancies and translating this success to other cancers remains an ongoing clinical objective. Next-generation T cell products in development aim to genetically modulate many facets of cell behavior, for which gene-nominating platforms provide a useful framework for prioritization. Among competing screening approaches, CRISPR activation (CRISPRa) technology permits gain-of-function (GoF) gene surveys at genome-wide scale, but routine implementation in primary T cells has been stymied by high cell requirements ([~]107 - 108) and abbreviated activity. Here, we describe a novel cell manufacturing schema using an all-in-one transposon-based gene delivery system coupled with CAR-restricted cell expansion to generate yields (109) of primary T cells bearing CAR and CRISPRa transgenes that are well above the threshold needed for genome-scale screening. CRISPRa activity is sustained via the inclusion of divergent, duplicate Elongation Factor 1 core/human T-cell leukemia virus (EF1-HTLV) hybrid promoters; while guide RNA representation is preserved through late lentiviral transduction, thus preventing bottlenecking and premature candidate pruning. CRISPRa-CAR T cells manufactured via this pipeline retain potent on-target gene-overexpression (>85% target+) across varied cell subsets (e.g. Tim-3+Lag3+ or serial-challenge) and timescales (>14 days). When deployed to survival-based genome-wide selection landscapes, CRISPRa-CAR pools nominate known and novel endogenous genes capable of enhancing CD8+ CAR T survival in cytokine-rich (e.g. MYC, FUT6, IRF4, GSE1) and cytokine-depleted (e.g. CSF2RB, STAT6, IRF4, GSE1) settings of tumor challenge. This system will have broad utility for therapy-enhancing gene discovery.

immunology↗

bioRxiv2023

Rationally designed modular STAT-activating scaffolds enforce cell-intrinsic transcriptional programs augmenting the anti-tumor potency of CAR T cells

Chimeric antigen receptor (CAR)-expressing T cells can mediate anti-tumor responses in a variety of preclinical models and clinical settings, however, strategies to enhance anti-tumor potency is the subject of intense investigation. Signals emanating from gamma-c cytokine receptors modulate the transcriptional state of activated T cells impacting proliferation, survival, differentiation, and effector functioning through the STAT family of transcription factors. Design of ligand-independent cell-intrinsic cytokine STAT activation scaffolds is a conceptually attractive strategy to provide CAR T cells with a surrogate for exogenous cytokine support. Here, we designed a series of ligand-autonomous STAT inducer (LASI) scaffolds comprised of an extracellular identification tag, a homodimerizing transmembrane domain, and a membrane proximal IL7R Box1 domain followed by STAT5 and/or STAT3 docking sequences derived from IL7R and IL21R, respectively. We constructed LASI scaffolds having STAT5 (LASI-5), STAT3 (LASI-3), and combined STAT5 and STAT3 (LASI-5+3) docking domains and then interrogated their impact in primary human CD8+ anti-CD19 (4-1BB:zeta) CAR T cells. While LASI-5 expression had limited effects on CAR T cells, LASI-3 transcriptional programming was found to be indispensable to achieving anti-tumor functional enhancement associated with limited terminal differentiation, heightened T cell proliferation in response to antigen, and dampened expression of exhaustion-associated genes. Moreover, CAR T cells supplemented with LASI-3 or 5+3 displayed superior potency against human leukemia tumors in NSG mice. LASI-5+3 mediated the highest magnitude of CAR T cell engraftment in vivo that evolved into a fatal lymphoproliferative syndrome. However, the same efficacy enhancement was achieved with LASI-3 without the lymphoproliferative complication. Our findings provide a rationale for utilization of constitutively expressed LASI-3 to enhance the anti-tumor potency of CAR T cells, the need to regulate the activity of LASI-5+3, and a generalizable scaffold design for studying additional combinations of STAT family transcription factors.

bioengineering↗