Biology subjects

Mooney, D.

Publications and source records attributed to Mooney, D..

10 recordsLinked to original sources

bioRxiv2026

Programmable antigen-presenting scaffolds expand rare tumor-antigen-specific T cells from peripheral blood

Adoptive cellular therapies can be effective against solid tumors but rely on the identification and expansion of rare tumor-reactive T cells (TRTs). Here, we utilize antigen-presenting cell-mimetic scaffolds (APC-ms), a modular biomaterial platform that presents defined peptide-HLA and co-stimulatory ligands on a fluid lipid bilayer to selectively recover rare antigen-specific T cells from peripheral blood. Independent variation of peptide-HLA, co-stimulatory ligands, and cytokine support defined a formulation that enriched MART-1-specific T cells to above 2% of CD8+ T cells in 35 of 50 HLA-A*02:01+ healthy donors from low or undetectable baseline frequencies. Compared with mo-DCs, APC-ms achieved greater fold expansion of MART-1-specific T cells with increased functional avidity and preservation of an early differentiation phenotype, and generated a higher number of MART-1-specific clonotypes. In five of eight patients with treatment-naive melanoma, APC-ms could enrich MART-1-specific T cells to a median of 13.7% of CD8+ cells (range 0.30 - 57.2%) Moreover, APC-ms could expand T cells specific for an HLA-A*24:01-restricted personal neoantigen that were previously undetectable in peripheral blood of a patient with melanoma. These findings establish APC-ms as a tunable platform for the recovery of rare TRTs from peripheral blood, supporting its further development for therapeutic TCR discovery and personalized adoptive cell therapy.

immunology↗

bioRxiv2026

A Microgel-Based Platform for Tunable Expansion and Function of γδ T-cells

Current {gamma}{delta} T-cell expansion protocols often sacrifice functionality for yield and largely ignore the context of activation. Here we utilize a tunable alginate microgel system functionalized with anti-CD3 and co-stimulatory antibodies (CD28 or CD2) to investigate the impact of biochemical signaling and substrate mechanics on {gamma}{delta} T-cell activation. Microgel-mediated expansion was compared to conventional soluble antibodies and TransAct beads. The microgels enhanced {gamma}{delta} T-cell expansion compared to soluble antibodies, allowed for controlled tuning of differentiation state, and promoted higher NKG2D, IFN-{gamma} and TNF- expression levels. Functionally, microgel-expanded {gamma}{delta} T-cells exhibited superior cytotoxicity against both solid and liquid tumor targets. This system also allowed elucidation of the differences in stimulation requirements for various donors, based on the starting phenotype. These findings establish a tunable platform for engineering {gamma}{delta} T-cells with improved therapeutic potential. Significance Statement{gamma}{delta} T-cells have shown promising therapeutic effects when used for T cell-based immunotherapy to treat solid tumor. However, achieving rapid expansion of {gamma}{delta} T-cells while maintaining their functionality remains a major challenge, especially given the heterogeneous responses from donors. We demonstrate that a tunable microgel system with flexible presentation of stimulatory cues improves {gamma}{delta} T-cell expansion while preserving cytotoxic function and reveal how starting phenotypes influence responses to activation. These understandings will provide design rationale to enable patient-specific treatment for optimal therapeutic outcomes.

bioengineering↗

bioRxiv2026

Viscoelastic Niches Shape γδ T-Cell Phenotype and Effector Function

{gamma}{delta} T-cells, which are predominantly enriched in epithelial tissues, have been used in cancer therapy because of their capacity for rapid cytotoxicity in an MHC-independent manner. Current paradigms are largely agnostic to the role of tissue mechanical cues in regulating {gamma}{delta} T-cell function. Here, we investigated the role of matrix viscoelasticity in modulating {gamma}{delta} T-cell migration, differentiation state, phenotype, and function. Using a tunable collagen-based gel system, we found that encapsulation in highly-elastic (slow-relaxing) matrices preserved a less differentiated phenotype, as evidenced by CD27 and CD45RA expression. Slow-relaxing matrices also increased expression of Fas and PD-1, while decreasing expression of CD11a. Despite increased PD-1 expression, these cells remained functional, as demonstrated by high levels of TNF- and IFN-{gamma} relative to PD-1-negative cells. Proteomic analysis revealed that {gamma}{delta} T-cells respond to changes in viscoelasticity through actin remodeling and shifts in metabolic machinery. Overall, when compared to non-encapsulated cells (2D culture), encapsulated {gamma}{delta} T-cells showed increased expression of cytotoxic programs. Functionally, cells encapsulated in slow-relaxing gels showed improved control of tumor growth in an aggressive HCT116 tumor model. Together, these findings establish matrix viscoelasticity as an important regulator of post-thymic {gamma}{delta} T-cell differentiation and function.

bioengineering↗

bioRxiv2025

A novel human blood-brain barrier model reveals pericytes as critical regulators of viral neuroinvasion

The blood-brain barrier (BBB) plays a vital role in regulating the passage of biomolecules between the bloodstream and the central nervous system (CNS) while also protecting the CNS from pathogens. Pericytes reside at the interface between the endothelial cells that form the vessel walls and the brain parenchyma. These cells are critical for maintaining BBB integrity and play key roles in regulating vessel permeability, blood flow, and immune cell migration. In this study, we developed a novel serum-free protocol to generate neural crest cell-derived pericytes (NCC-PCs) from human pluripotent stem cells (hPSCs). These NCC-PCs enhance BMEC barrier function and can be co-cultured with hPSC-derived brain microvascular endothelial cells (BMECs) in a contact co-culture BBB model that recapitulates the in vivo cellular interactions at the BBB. We used this model to evaluate the pathological consequences of BBB cell infection by highly neuroinvasive flaviviruses. Our results identify a previously undescribed role for NCC-PCs in maintaining BMEC barrier integrity during infection and reducing the spread of viral infection to the CNS.

microbiology↗

bioRxiv2023

SARS-CoV-2 infection of human pluripotent stem cell-derived vascular cells reveals smooth muscle cells as key mediators of vascular pathology during infection

Although respiratory symptoms are the most prevalent disease manifestation of infection by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), nearly 20% of hospitalized patients are at risk for thromboembolic events. This prothrombotic state is considered a key factor in the increased risk of stroke, which is observed clinically during both acute infection and long after symptoms clear. Here we develop a model of SARS-CoV-2 infection using human-induced pluripotent stem cell-derived endothelial cells (ECs), pericytes (PCs), and smooth muscle cells (SMCs) to recapitulate the vascular pathology associated with SARS-CoV-2 exposure. Our results demonstrate that perivascular cells, particularly SMCs, are a susceptible vascular target for SARS-CoV-2 infection. Utilizing RNA sequencing, we characterize the transcriptomic changes accompanying SARS-CoV-2 infection of SMCs, PCs, and ECs. We observe that infected SMCs shift to a pro-inflammatory state and increase the expression of key mediators of the coagulation cascade. Further, we show human ECs exposed to the secretome of infected SMCs produce hemostatic factors that contribute to vascular dysfunction, despite not being susceptible to direct infection. The findings here recapitulate observations from patient sera in human COVID-19 patients and provide mechanistic insight into the unique vascular implications of SARS-CoV-2 infection at a cellular level.

microbiology↗

bioRxiv2022

Optimizing CpG spatial distribution with DNA origami for Th1-polarized therapeutic vaccination

Multivalent presentation of ligands often enhances receptor activation and downstream signaling. DNA origami offers precise nanoscale spacing of ligands, a potentially useful feature for therapeutic nanoparticles. Here we introduce a "square block" DNA origami platform to explore the importance of spacing of CpG oligonucleotides, which engage Toll-like receptors and thereby act as danger signals for dendritic cells. Through in vitro cell-culture studies and in vivo tumor-treatment models, we demonstrate that square blocks induce Th1 immune polarization when CpG is spaced at 3.5 nm. We observe that this DNA origami vaccine enhances DC activation, antigen cross-presentation, CD8 T cell activation, Th1-polarized CD4 activation and NK cell activation. The vaccine also synergizes effectively with anti-PD-L1 for improved cancer immunotherapy in melanoma and lymphoma models and induces long-term T cell memory. Our results suggest that DNA origami may serve as an advanced vaccine platform for controlling adjuvant spacing and co-delivering antigens. One Sentence SummaryThis study developed a DNA origami-based cancer vaccine (DoriVac) that co-delivers antigen and CpG immune adjuvant with an optimal nanospacing for Th1 immune polarization.

synthetic biology↗

bioRxiv2022

Development of a physiological insulin resistance model in human stem cell-derived adipocytes

Adipocytes are key regulatory cells of human metabolism, and their dysfunction in insulin signaling is central to metabolic diseases such as type II diabetes mellitus (T2D). However, the progression of insulin resistance that leads to T2D is still poorly understood. This limited understanding is due, in part, to the dearth of suitable models of insulin signaling in human adipocytes. Traditionally, in vitro adipocyte models fail to recapitulate in vivo insulin signaling, possibly due to exposure to supraphysiological nutrient and hormone conditions. Here, we have developed a sensitization protocol for human pluripotent stem cell-derived adipocytes that uses physiologically relevant nutrient conditions to produce a potent signaling response comparable to in vivo adipocytes. After systematically optimizing conditions, this protocol allows for robust insulin-stimulated glucose uptake and transcriptional insulin response. Furthermore, exposure of these sensitized adipocytes to physiologically relevant hyperinsulinemic conditions dampens insulin-stimulated glucose uptake and dysregulates transcription of insulin-responsive genes. Overall, this sensitization methodology provides a novel platform for the mechanistic study of insulin signaling and resistance using human pluripotent stem cell-derived adipocytes. TeaserA new protocol to generate hPSC-adipocytes that respond to physiological insulin levels and can model diabetes.

cell biology↗

bioRxiv2022

Immune-responsive biodegradable scaffolds for enhancing neutrophil regeneration

Neutrophils are essential effector cells for mediating rapid host defense and their insufficiency arising from therapy-induced side-effects, termed neutropenia, can lead to immunodeficiency-associated complications. In autologous hematopoietic stem cell transplantation (HSCT), neutropenia is a complication that limits therapeutic efficacy. Here, we report the development and in vivo evaluation of an injectable, biodegradable hyaluronic acid (HA)-based scaffold, termed HA cryogel, with myeloid responsive degradation behavior. In mouse models of immune deficiency, we show that the infiltration of functional myeloid-lineage cells, specifically neutrophils, is essential to mediate HA cryogel degradation. Post-HSCT neutropenia in recipient mice delayed degradation of HA cryogels by up to 3 weeks. We harnessed the neutrophil-responsive degradation to sustain the release of granulocyte colony stimulating factor (G-CSF) from HA cryogels. Sustained release of G-CSF from HA cryogels enhanced post-HSCT neutrophil recovery, comparable to pegylated G-CSF, which, in turn, accelerated cryogel degradation. HA cryogels are a potential approach for enhancing neutrophils and concurrently assessing immune recovery in neutropenic hosts.

bioengineering↗

bioRxiv2022

Matrix viscoelasticity controls spatio-temporal tissue organization

The spatio-temporal patterning of multicellular tissues is driven by the collective dynamics of cell proliferation and active movement. These processes are mediated by the extracellular matrix environment via a combination of biomolecular and physical cues. Here we show that the passive viscoelastic properties of the matrix that encapsulate a proliferating ball of cells (e.g. a developing organoid) play a critical role in guiding tissue organization in space and time. By varying the viscoelasticity of well-defined model matrices, we show how a spheroidal tissue of breast epithelial cells breaks symmetry and forms finger-like protrusions that invade the matrix. A computational model allows us to recapitulate these observations and leads to a phase diagram that demarcates the regions of morphological stability and instability as a function of matrix viscoelasticity, tissue viscosity, cell motility and cell division rate. Experiments that use biomolecular manipulations to independently vary these parameters confirm our predictions. To further test our theory, we also study the self-organization of an in-vitro intestinal organoid and show that the morphological changes of this system also fits within our paradigm. Altogether, our studies demonstrate the role of stress relaxation mechanisms in determining the dynamics of tissue growth and the symmetry breaking instabilities associated with branching, a fundamental process in morphogenesis and oncogenesis, and suggest ways of controlling tissue form using the extracellular matrix.

biophysics↗

bioRxiv2021

Induced reprogramming of adult murine cardiomyocytes to pluripotency in vivo

Partial cell reprogramming has been demonstrated in certain mouse tissues by in situ overexpression of Oct3/4, Klf4, Sox2 and cMyc (OKSM) transcription factors, and can induce rejuvenation and/or augment regeneration. Reprogramming of adult cardiomyocytes has been elusive until recently, but its success could help overcome the lack of endogenous regenerative capacity of the mammalian myocardium. Here, we generated cardiomyocyte-specific, doxycycline-inducible, reprogrammable mice and demonstrated that sustained OKSM induction reprograms cardiomyocytes fully into teratoma-forming pluripotent cells. However, we also showed that cyclic OKSM upregulation induces significant decrease of epigenetic age in the cardiomyocytes without de-differentiation or reacquisition of pluripotency. In mice with progressive heart failure, cardiomyocyte epigenetic rejuvenation correlated with stabilization of systolic heart function. These findings confirm that OKSM can reprogram adult mouse cardiomyocytes to different states depending on the duration of their expression, and provide further evidence that partially reprogrammed cardiomyocytes may contribute to ameliorate cardiac disease.

developmental biology↗