Biology subjects

McCall, A. S.

Publications and source records attributed to McCall, A. S..

9 recordsLinked to original sources

bioRxiv2026

A 4D GelMA-Cad/Gelatin Hydrogel for High-Resolution DMD Bioprinting of Vascularized Soft Tissue Models

Digital micromirror device (DMD)-based bioprinting enables high-resolution fabrication of hydrogel-based tissue constructs, but print fidelity and hydrogel mechanics are typically coupled through polymer concentration, light exposure, and crosslinking density. This creates a materials constraint for vascular bioprinting, where constructs must preserve microscale architecture during fabrication while supporting multicellular remodeling, lumen formation, and long-term tissue organization. To meet these requirements, we developed a 4D GelMA-Cad/gelatin hydrogel with time-dependent mechanics that maintains print fidelity while providing a soft, cell-permissive matrix. GelMA-Cad provides a covalently photocrosslinked, bioactive network containing an N-cadherin-mimetic peptide, while uncrosslinked gelatin serves as a transient matrix-modifying component that diffuses from the hydrogel after fabrication, producing further softening without bulk degradation of the GelMA-Cad network. We characterized hydrogel chemistry with NMR, quantified time-dependent changes in stiffness, visualized pore morphology with cryo-SEM, and evaluated the platform in retinal and pulmonary constructs. The 4D hydrogel supports 3D-patterned accuracy, cell viability, inner blood-retinal barrier vasculature, and vascularized lung-alveolar tissue models. Single-cell RNA-sequencing of the pulmonary vascular-alveolar model demonstrated compartment-specific transcriptional responses, including enrichment of lung capillary-associated endothelial features and AT1-like epithelial states. Overall, this platform decouples DMD print fidelity from post-print matrix mechanics for vascularized soft tissue bioprinting and tissue modeling.

bioengineering↗

bioRxiv2026

IPF distal lung epithelial cells acquire a DNA methylation signature consistent with activation of basal cell transcriptional programs

Idiopathic pulmonary fibrosis (IPF) is a progressive fibrotic interstitial lung disease associated with failed alveolar epithelial repair with an expansion of aberrant airway-like epithelium in the alveolar space leading to lung function decline usually resulting in death within 3-5 years of diagnosis. While single-cell and spatial transcriptomic approaches have been used to characterize disease-emergent cell populations, less is known about the regulation of transcriptional programs that drive failed alveolar epithelial cell repair in IPF. DNA methylation is a fundamental layer of gene regulation that stabilizes differentiated cell identity; however, changes in methylation in the epithelial compartment in IPF have not been studied. To identify novel epigenetic mediators of epithelial cell dysfunction, we performed high-resolution DNA methylation profiling of purified distal lung epithelial cells from 10 age-matched control and 14 IPF lungs using Oxford Nanopore Technologies (ONT) whole-genome, long-read sequencing. We identified widespread methylome remodeling in the IPF lung epithelium, with 84% of differentially methylated regions (DMRs) hypomethylated. DMRs were largely found outside of promoters, with 88% outside of 3 kb from the transcription start site (TSS), consistent with altered distal regulatory element activity. DMRs were enriched for transcription factor (TF) binding site motifs and gene associations consistent with dysregulation of polycomb repressive complex 2 (PRC2) and increases in p63 activity. DNA methylation at DMRs associated with p63 target genes, including KRT5, and genes implicated in failed epithelial repair, including MUC5B, MMP7, and S100A2 inversely correlated with gene expression. Consistent with the observed dysregulation of the IPF epithelial methylome, an experimental co-culture model of alveolar type II epithelial cell (AT2s) to basal-like epithelial cell transdifferentiation revealed widespread hypomethylation. Further, sites associated with NKX2.1 and FOXA1/2 binding, TFs involved in alveolar fate maintenance, were hypermethylated, suggesting loss of epigenetic regulation of alveolar identity. Together, these data implicate DNA methylation in the failed alveolar epithelial repair processes in IPF, potentially providing future therapeutic strategies by identifying putative regulatory elements associated with aberrant transcriptional programs.

molecular biology↗

bioRxiv2025

Loss of Regulator of telomere elongation helicase (Rtel1) impairs alveolar epithelial cell progenitor function but does not exacerbate experimental fibrosis

Inherited loss-of-function mutations in the replication-associated helicase Regulator of telomere elongation helicase (RTEL1) are among the most frequent monogenic causes of familial pulmonary fibrosis (FPF), but the specific functional role of RTEL1 in lung development, homeostasis, and injury-repair has not been determined. Using single-cell RNA-sequencing to define the cellular diversity and molecular programs following intratracheal bleomycin injury in mice, we observed that Rtel1 was most highly expressed in the lung epithelium, and its expression was reduced in PF epithelial cells compared to nonfibrotic controls. While global deletion of Rtel1 leads to embryonic lethality, constitutive deletion of Rtel1 in the lung epithelium (Rtel{Delta}epi) led to normal lung structure and architecture. Rtel1{Delta}epi mice exhibited accelerated telomere attrition and impaired AEC progenitor potential ex-vivo. Despite decreased AEC progenitor function, no differences in lung fibrosis were observed in Rtel{Delta}epi mice compared to controls following single-dose or repetitive intratracheal bleomycin. Together, these data implicate RTEL1 as a regulator of AEC progenitor function but this level of progenitor dysfunction is not alone sufficient to cause or exacerbate experimental lung fibrosis.

cell biology↗

bioRxiv2025

Dehydration promotes intracellular lipid synthesis and accumulation

Lipids can be considered a water reservoir used to offset dehydration stress as their oxidation by the mitochondria generates water. However, whether dehydration and the ensuing hypertonic stress directly regulate lipid synthesis is unknown. We found that hypertonic stress decreases cellular oxygen consumption, increases intracellular lipid synthesis, and favors glutamine oxidation as a carbon precursor for lipid synthesis via remodeling mitochondrial metabolism. These findings provide a mechanism whereby cellular dehydration leads to intracellular lipid accumulation, functionally linking water availability to lipid storage.

physiology↗

bioRxiv2025

Sustained Yap/Taz activation promotes aberrant alveolar epithelial cell differentiation and drives persistent fibrotic remodeling

YAP/TAZ signaling is required for initiation of lung alveolar repair, yet previous studies in idiopathic pulmonary fibrosis (IPF) predicted increased YAP/TAZ signaling in alveolar epithelial cells (AECs). We investigated whether persistent YAP/TAZ AEC signaling contributes to failed epithelial repair and persistent fibrotic remodeling. In IPF lungs, we identified increased YAP+/TAZ+ AECs and increased expression of YAP/TAZ transcriptional targets compared to donor control lungs. In human lung organoids, pharmacological YAP/TAZ activation resulted in phenotype shifts of AECs into aberrant transitional states. In mice with Yap/Taz activation (YTactive) resulting from deletion of Hippo-kinases Stk3/4 in alveolar-type 2 (AT2) cells, resulted in persistent fibrotic remodeling at 28- and 56-days post-bleomycin injury. Gene promoter activity associated with transitional cell markers (Krt19, Hopx, and Runx2) was increased in YTactive AT2 cells. Immunofluorescent staining showed a loss of AT2 associated Cebpa and increased Krt19 in YTactive lineage traced AT2 cells 28 days post-injury. Inhibition of Yap/Taz using Verteporfin resulted in improved lung repair in YTactive mouse lungs, including increased Cebpa and decreased Krt19+ transitional cells. These findings demonstrate sustained Yap/Taz activation drives abnormal alveolar repair and persistent fibrotic remodeling. Blocking aberrant persistent Yap/Taz activity promotes adaptive repair and has potential as a therapeutic strategy for PF.

molecular biology↗

bioRxiv2024

Anatomically and Physiologically Accurate Engineered Neurovascular Unit and Blood-Brain Barrier Model Using Microvessels Isolated from Postmortem Human Brain Tissue

Brain vasculature is a complex and heterogeneous physiological structure that serves specialized roles in maintaining brain health and homeostasis. There is substantial interest in developing representative human models of the brain vasculature for drug screening and disease modeling applications. Many contemporary strategies have focused on culturing neurovascular cell types in hydrogels and microdevices, but it remains challenging to achieve anatomically relevant vascular structures that have physiologically similar function to their in vivo counterparts. Here, we present a strategy for isolating microvessels from cryopreserved human cortical tissue and culturing these vessels in a biomimetic gelatin-based hydrogel contained in a microfluidic device. We provide histological evidence of arteriole and capillary architectures within hydrogels, as well as anastomosis to the hydrogel edges allowing lumen perfusion. In capillaries, we demonstrate restricted diffusion of a 10 kDa dextran, indicating intact passive blood-brain barrier function. We anticipate this bona fide human brain vasculature-on-a-chip will be useful for various biotechnology applications.

bioengineering↗

bioRxiv2023

Image-based spatial transcriptomics identifies molecular niche dysregulation associated with distal lung remodeling in pulmonary fibrosis

The human lung is structurally complex, with a diversity of specialized epithelial, stromal and immune cells playing specific functional roles in anatomically distinct locations, and large-scale changes in the structure and cellular makeup of this distal lung is a hallmark of pulmonary fibrosis (PF) and other progressive chronic lung diseases. Single-cell transcriptomic studies have revealed numerous disease-emergent/enriched cell types/states in PF lungs, but the spatial contexts wherein these cells contribute to disease pathogenesis has remained uncertain. Using sub-cellular resolution image-based spatial transcriptomics, we analyzed the gene expression of more than 1 million cells from 19 unique lungs. Through complementary cell-based and innovative cell-agnostic analyses, we characterized the localization of PF-emergent cell-types, established the cellular and molecular basis of classical PF histopathologic disease features, and identified a diversity of distinct molecularly-defined spatial niches in control and PF lungs. Using machine-learning and trajectory analysis methods to segment and rank airspaces on a gradient from normal to most severely remodeled, we identified a sequence of compositional and molecular changes that associate with progressive distal lung pathology, beginning with alveolar epithelial dysregulation and culminating with changes in macrophage polarization. Together, these results provide a unique, spatially-resolved characterization of the cellular and molecular programs of PF and control lungs, provide new insights into the heterogeneous pathobiology of PF, and establish analytical approaches which should be broadly applicable to other imaging-based spatial transcriptomic studies.

genomics↗

bioRxiv2023

Hypoxia-inducible factor-2 (HIF2) regulates alveolar regeneration after repetitive injury

Idiopathic Pulmonary Fibrosis (IPF) is a progressive and often fatal chronic respiratory disease thought to result from repetitive injury and failed repair of the lung alveoli, and recent studies have identified a number of disease-emergent intermediate/transitional cell states in the IPF lung supporting this concept. In this study, we found that persistent activation of hypoxia-inducible factor (HIF)-signaling in airway-derived, repair-associated cell types/states is a hallmark of dysfunctional epithelial repair in the IPF lung epithelium and experimental models of recurrent lung epithelial injury. Disrupting Hif-signaling attenuated experimental lung fibrosis, reduced mucous-secretory cell polarization, and promoted functional alveolar regeneration following repetitive injury. Mouse and human organoid studies demonstrated that small-molecule-based HIF2 inhibition promoted alveolar epithelial cell proliferation and maturation while preventing the emergence of maladaptive intermediate/transitional states analogous to those in IPF. Together, these studies indicate that targeted HIF2-inhibition represents a novel and effective therapeutic strategy to promote functional lung regeneration, and could be readily translated into human studies of IPF and other chronic interstitial lung diseases with disease modifying effect. One sentence summaryInhibiting hypoxia-inducible-factor 2 (HIF2) promotes functional lung alveolar epithelial repair following recurrent injury.

cell biology↗

bioRxiv2023

Epithelial Yap/Taz are required for functional alveolar regeneration following acute lung injury

A hallmark of idiopathic pulmonary fibrosis (IPF) and other interstitial lung diseases is dysregulated repair of the alveolar epithelium. The Hippo pathway effector transcription factors YAP and TAZ have been implicated as essential for type 1 and type 2 alveolar epithelial cell (AT1 and AT2) differentiation in the developing lung, yet aberrant activation of YAP/TAZ is a prominent feature of the dysregulated alveolar epithelium in IPF. In these studies, we sought to define the functional role of YAP/TAZ activity during alveolar regeneration. We demonstrate that Yap and Taz are normally activated in AT2 cells shortly after injury, and deletion of Yap/Taz in AT2 cells led to pathologic alveolar remodeling, failure of AT2 to AT1 cell differentiation, increased collagen deposition, exaggerated neutrophilic inflammation, and increased mortality following injury induced by a single dose of bleomycin. Loss of Yap/Taz activity prior to a LPS injury prevented AT1 cell regeneration, led to intra-alveolar collagen deposition, and resulted in persistent innate inflammation. Together these findings establish that AT2 cell Yap/Taz activity is essential for functional alveolar epithelial repair and prevention of fibrotic remodeling.

cell biology↗