Biology subjects

Bliley, J. M.

Publications and source records attributed to Bliley, J. M..

6 recordsLinked to original sources

bioRxiv2026

Multi-Modal photoFRESH: Light-Pipe Embedded Printing of Heterogeneous Hydrogel and Tissue Architectures

Recreating the complex spatial gradients and multi-material transitions of native tissues remains a fundamental challenge in 3D bioprinting. To address this, we introduce multi-modal photoFRESH, which integrates localized photochemistry into embedded printing by delivering light through a fiber-optic light-pipe. By tuning numerical aperture, print speed, and photoabsorber content, we achieve precise layer-by-layer control of crosslinking, stiffness, and bioorthogonal biomolecular tethering while preserving high print fidelity. Both photoactivatable support baths and extruded bioinks can be patterned, together with traditional FRESH printing. Utility of the platform is demonstrated by the fabrication of structurally complex tissue scaffolds and cellularized muscle constructs with distinct mechanical and biochemical domains. This multi-modal approach expands the boundaries of embedded bioprinting toward functional and heterogeneous tissue architectures.

bioengineering↗

bioRxiv2025

Engineering 3D Skeletal Muscle Tissue with Complex Multipennate Myofiber Architectures

The hierarchical architecture of skeletal muscle spans from microscale sarcomeres to macroscale myofibers and is integral to its contractile functionality. Pathologies such as volumetric muscle loss (VML) compromise this structure and destroy the native extracellular matrix (ECM), exceeding the regenerative capacity of endogenous repair mechanisms. Here, we present a novel method for tissue engineering biomimetic three-dimensional (3D) skeletal muscle with complex architectures by leveraging freeform reversible embedding of suspended hydrogels (FRESH) 3D bioprinting of the ECM. Collagen type I scaffolds mimicking diverse muscle architectures-- including parallel, unipennate, bipennate, multipennate, and convergent--were designed, FRESH printed, and seeded with C2C12 myoblasts to guide myogenesis. Engineered muscle tissues demonstrated scaffold-mediated alignment and fusion into functional myotubes, exhibiting contractile responses to electrical stimulation with architecture-dependent specific force of [~]1 kN/m2 and a positive force-frequency relationship. In vivo implantation further revealed scaffold-directed cellular and vascular organization, underscoring the translational potential of this approach. In summary, this study demonstrates the capability to use FRESH 3D bioprinting to engineer physiologically relevant muscle architectures, significantly advancing the design of functional muscle tissues for regenerative medicine and in vitro modeling applications.

bioengineering↗

bioRxiv2024

Transient Notch Activation Converts Pluripotent Stem Cell-Derived Cardiomyocytes towards a Purkinje Fiber Fate

Cardiac Purkinje fibers form the most distal part of the ventricular conduction system. They coordinate contraction and play a key role in ventricular arrhythmias. While many cardiac cell types can be generated from human pluripotent stem cells, methods to generate Purkinje fiber cells remain limited, hampering our understanding of Purkinje fiber biology and conduction system defects. To identify signaling pathways involved in Purkinje fiber formation, we analyzed single cell data from murine embryonic hearts and compared Purkinje fiber cells to trabecular cardiomyocytes. This identified several genes, processes, and signaling pathways putatively involved in cardiac conduction, including Notch signaling. We next tested whether Notch activation could convert human pluripotent stem cell-derived cardiomyocytes to Purkinje fiber cells. Following Notch activation, cardiomyocytes adopted an elongated morphology and displayed altered electrophysiological properties including increases in conduction velocity, spike slope, and action potential duration, all characteristic features of Purkinje fiber cells. RNA-sequencing demonstrated that Notch-activated cardiomyocytes undergo a sequential transcriptome shift, which included upregulation of key Purkinje fiber marker genes involved in fast conduction such as SCN5A, HCN4 and ID2, and downregulation of genes involved in contractile maturation. Correspondingly, we demonstrate that Notch-induced cardiomyocytes have decreased contractile force in bioengineered tissues compared to control cardiomyocytes. We next modified existing in silico models of human pluripotent stem cell-derived cardiomyocytes using our transcriptomic data and modeled the effect of several anti-arrhythmogenic drugs on action potential and calcium transient waveforms. Our models predicted that Purkinje fiber cells respond more strongly to dofetilide and amiodarone, while cardiomyocytes are more sensitive to treatment with nifedipine. We validated these findings in vitro, demonstrating that our new cell-specific in vitro model can be utilized to better understand human Purkinje fiber physiology and its relevance to disease.

developmental biology↗

bioRxiv2021

Recapitulate Human Cardio-pulmonary Co-development Using Simultaneous Multilineage Differentiation of Pluripotent Stem Cells

The extensive crosstalk between the developing heart and lung is pivotal for their proper morphogenesis and maturation. However, there remains a lack of model systems for investigating the critical cardio-pulmonary mutual interaction during human embryogenesis. Here, we reported a novel stepwise strategy for directing simultaneous induction of both mesoderm-derived cardiac and endoderm-derived lung epithelial lineages within a single differentiation of human pluripotent stem cells (hPSCs) via temporal specific tuning of WNT and TGF-{beta} signaling in the absence of exogenous growth factors. Using 3D suspension culture, we established concentric cardio-pulmonary micro-Tissues (Ts), and observed expedited alveolar maturation in the presence of cardiac accompany. Upon withdrawal of WNT agonist, the cardiac and pulmonary components within each dual-lineage T effectively segregated from each other with concurrent initiation of cardiac contraction. We expect our multilineage differentiation model to offer an experimentally tractable system for investigating human cardio-pulmonary interplay and tissue boundary formation during embryogenesis.

developmental biology↗

bioRxiv2020

Dynamic Loading of Human Engineered Heart Tissue Enhances Contractile Function and Drives Desmosome-linked Disease Phenotype

The role mechanical forces play in shaping the structure and function of the heart is critical to understanding heart formation and the etiology of disease but is challenging to study in patients. Engineered heart tissues (EHTs) incorporating human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes have the potential to provide insight into these adaptive and maladaptive changes in the heart. However, most EHT systems are unable to model both preload (stretch during chamber filling) and afterload (pressure the heart must work against to eject blood). Here, we have developed a new dynamic EHT (dyn-EHT) model that enables us to tune preload and have unconstrained fractional shortening of >10%. To do this, 3D EHTs are integrated with an elastic polydimethylsiloxane (PDMS) strip that provides mechanical pre- and afterload to the tissue in addition to enabling contractile force measurements based on strip bending. Our results demonstrate in wild-type EHTs that dynamic loading is beneficial based on the magnitude of the forces, leading to improved alignment, conduction velocity, and contractility. For disease modeling, we use hiPSC-derived cardiomyocytes from a patient with arrhythmogenic cardiomyopathy (ACM) due to mutations in desmoplakin. We demonstrate that manifestation of this desmosome-linked disease state requires the dyn-EHT conditioning and that it cannot be induced using 2D or standard 3D EHT approaches. Thus, dynamic loading strategy is necessary to provoke a disease phenotype (diastolic lengthening, reduction of desmosome counts, and reduced contractility), which are akin to primary endpoints of clinical disease, such as chamber thinning and reduced cardiac output. Single Sentence SummaryDevelopment of a dynamic mechanical loading platform to improve contractile function of engineered heart tissues and study cardiac disease progression.

bioengineering↗

bioRxiv2020

Fibronectin-Based Nanomechanical Biosensors to Map 3D Strains in Live Cells and Tissues

Mechanical forces are integral to a wide range of cellular processes including migration, differentiation and tissue morphogenesis; however, it has proved challenging to directly measure strain at high spatial resolution and with minimal tissue perturbation. Here, we fabricated, calibrated, and tested a fibronectin (FN)-based nanomechanical biosensor (NMBS) that can be applied to cells and tissues to measure the magnitude, direction, and dynamics of strain from subcellular to tissue length-scales. The NMBS is a fluorescently-labeled, ultrathin square lattice FN mesh with spatial resolution tailored by adjusting the width and spacing of the lattice fibers from 2-100 {micro}m. Time-lapse 3D confocal imaging of the NMBS demonstrated strain tracking in 2D and 3D following mechanical deformation of known materials and was validated with finite element modeling. Imaging and 3D analysis of the NMBS applied to single cells, cell monolayers, and Drosophila ovarioles demonstrated the ability to dynamically track microscopic tensile and compressive strains in various biological applications with minimal tissue perturbation. This fabrication and analysis platform serves as a novel tool for studying cells, tissues, and more complex systems where forces guide structure and function.

bioengineering↗