Biology subjects

Riley, L.

Publications and source records attributed to Riley, L..

12 recordsLinked to original sources

bioRxiv2026

Physical continuity at biomaterial-ECM interfaces regulates fibroblast activation via NF-κB

Fibrotic responses at biomaterial-tissue interfaces limit implant integration and regenerative healing, yet how the interaction between biomaterials and the extracellular matrix (ECM) regulates fibroblast activation remains poorly understood. Granular hydrogels including microporous annealed particle scaffolds (MAP) reduce fibrosis, while chemically and mechanically matched hydrogels do not, suggesting a dominant role for scaffold architecture. In this model, MAP scaffolds allow collagen infiltration and form physically continuous composites, whereas hydrogels exclude collagen and generate interfacial slip planes. To isolate how biomaterial architecture influences extracellular matrix (ECM) integration and fibroblast activation, we developed a reductionist in vitro model that integrates collagen type I with either microporous annealed particle (MAP) scaffolds or chemically and mechanically matched bulk hydrogels. This physical integration stabilizes collagen architecture, limits fibroblast-mediated matrix compaction, suppresses contractility, and attenuates myofibroblast transition. Fibroblasts in mechanically integrated environments exhibit reduced expression and nuclear localization of NF-{kappa}B and are enriched for quiescent phenotypes. Together, these findings identify biomaterial-ECM physical continuity as a design principle for limiting fibrotic signaling.

bioengineering↗

bioRxiv2025

Metrics for studying the porous void space of packed particles

Characterizing porosity in packed particle assemblies is a complex task that requires advanced analytical tools. We present a visually rich and extensive library of global, pore-based, and other metrics for analyzing features of porosity in such assemblies. Our library includes over 25 descriptors of "3D pores" that are identified using our LOVAMAP software. By applying our metrics to a set of simulated packings that vary by particle size, shape, and stiffness, we reveal predictable relationships between particle and void space characteristics. We identify two fundamental parameters of a monodisperse particle system - particle diameter ({delta}) and void volume fraction ({phi}) - that govern several void space features, such as the total number of bottlenecks (i.e., doors between pores), the median value of the largest enclosed sphere across all pores in a packing, and the fraction of reaction-center "hotspots." Through regression analyses on transformations of{delta} and{phi} , we quantify multiple packing-descriptor relationships, demonstrating, for example, that packing properties scale linearly with the median values of length-based descriptors across assemblies. We further introduce approaches for computing the number of vertices, edges, and faces of 3D pores, allowing for approximation to simpler polyhedra. Additional metrics explore surface entrances into the particle scaffold, traversable paths through the void space, and size-based accessibility. Together, these descriptors, which have been bundled into LOVAMAP, offer new insights into particle-pore architecture and spatial organization.

bioengineering↗

bioRxiv2025

Clustered VEGF Nanoparticles in Microporous Annealed Particle (MAP) Hydrogel Accelerates Functional Recovery and Brain Tissue Repair after Stroke

Ischemic stroke, a blockage in the vasculature of the brain that results in insufficient blood flow, is one of the worlds leading causes of disability. The cascade of inflammation and cell death that occurs immediately following stroke drives vascular and functional loss that does not fully recover over time, and no FDA-approved therapies exist that stimulate regeneration post-stroke. We have previously developed a hydrogel scaffold that delivered heparin nanoparticles with and without VEGF bound to their surface to promote angiogenesis and reduce inflammation, respectively. However, the inclusion of the naked heparin nanoparticles warranted concern over the development of bleeding complications. Here, we explore how microporous annealed particle (MAP) scaffolds functionalized with VEGF coated heparin nanoparticles can both reduce inflammation and promote angiogenesis - without the inclusion of free heparin nanoparticles. We show that our updated design not only successfully promotes de novo tissue formation, including the development of mature vessels and neurite sprouting, but it also leads to functional improvement in a photothrombotic stroke model. In addition, we find increased astrocyte infiltration into the infarct site correlated with mature vessel formation. This work demonstrates how our biomaterial design can enhance endogenous regeneration post-stroke while eliminating the need for excess heparin.

bioengineering↗

bioRxiv2024

Highly porous granular hydrogels reinforced by electrospun hydrogel fibers for long-term stability

Granular hydrogels (GHs) formed from hydrogel microparticles (HMPs) possess inherent, microscale porosity. In GHs including microporous annealed particle (MAP) hydrogels as well as unannealed GHs, increases in pore size and total porosity correlate with enhanced cellular infiltration and growth. However, increased porosity or pore size are typically achieved by decreasing HMP density, reducing interparticle contacts and weakening hydrogel structure. Here, to achieve a highly porous GH without compromising stability, we developed an approach using sacrificial HMPs to achieve high, mesoscale porosity and high-aspect ratio hydrogel fibers to reinforce the scaffold. The use of sacrificial HMPs circumvented the need to decrease packing density, creating GHs with up to 60% porosity. The hydrogel fibers span particles in the porous space, stabilizing the porous structure by enhancing interactions among GH components. Upon 5-10% v/v fiber incorporation, highly porous GHs retained their structure over 28 days of in vivo culture, compared to 4 days in GHs without fibers. These scaffolds supported and promoted endothelial cell growth as porosity increased. Overall, this study presents a stable, highly porous granular scaffold system with high processability and cytocompatibility.

bioengineering↗

bioRxiv2024

Myeloid-instructed CD14+CD4+ T cells within the tumor microenvironment are associated with TNFα signaling and prolonged survival in non-small cell lung cancer

The tumor microenvironment (TME) harbors a diverse array of innate and adaptive immune cells that can either support anti-tumor immunity or facilitate tumor progression. Among these, myeloid cells are key drivers of immunosuppression, yet therapeutic strategies targeting their function have achieved limited clinical success. A deeper understanding of the mechanisms by which immunosuppressive myeloid cells promote tumor progression is needed to guide the development of more effective treatments. Using spatial multi-omics analyses, we identified a population of myeloid-instructed CD14+ T cells present in both the tumor core and adjacent non-malignant lung tissue of patients with non-small cell lung cancer (NSCLC). CD4+ T cells acquired CD14 from myeloid cells by trogocytosis, resulting in an atypical T cell phenotype. High infiltration of CD14+CD4+ T cells in the tumor core was associated with poor patient survival. Spatial transcriptomics profiling revealed that tumors enriched in CD14+CD4+ T cells exhibited increased TNF signaling. Functional assays demonstrated that TNF enhanced trogocytosis, promoting the transfer of myeloid membrane components to CD4{square} T cells and driving the accumulation of CD14{square}CD4{square} T cells. These findings uncover a novel TNF-mediated mechanism of immunosuppression in the TME, whereby TNF promotes aberrant myeloid-T cell interactions that may contribute to tumor progression. This work highlights a previously unrecognized axis of innate-adaptive immune crosstalk in NSCLC and suggests that targeting TNF could disrupt this pathway, restore effective T cell function, and improve therapeutic outcomes.

cancer biology↗

bioRxiv2023

Engineering the microstructure and spatial bioactivity of MAP scaffolds in vitro instructs neovascularization in vivo

In tissues where the vasculature is either lacking or abnormal, biomaterials can be designed to promote vessel formation and enhance tissue repair. In this work, we independently tune the microstructure and bioactivity of microporous annealed particle (MAP) scaffolds to guide cell patterning in 3D and promote de novo assembly of endothelial progenitor-like cells into vessels. We implement both in silico characterization and in vitro experimentation to elucidate an optimal scaffold formulation for vessel formation. We determine that MAP scaffolds with pore volumes on the same order of magnitude as cells facilitate cell growth and vacuole formation. We achieve spatial control over cell spreading by incorporating adhesive microgels in well-mixed, heterogeneous MAP scaffolds. While we demonstrate that integrin engagement is the primary driver of network formation in these materials, introducing adhesive microgels loaded with heparin nanoparticles leads to the formation of vascular tubes after 3 days in culture. We then show in vivo that this unique scaffold formulation enhances vessel maturation in a wound healing model and instructs differential vascular patterning in the tumor microenvironment. Taken together, this work determines the optimal microstructure and ligand presentation within MAP scaffolds that lead to vascular constructs in vitro and facilitate neovascularization in vivo.

bioengineering↗

bioRxiv2023

Exploring the Role of Spatial Confinement in Immune Cell Recruitment and Regeneration of Skin Wounds

Microporous annealed particle (MAP) scaffolds are injectable granular materials comprised of micron sized hydrogel particles (microgels). The diameter of these microgels directly determines the size of the interconnected void space between particles where infiltrating or encapsulated cells reside. This tunable porosity allows us to use MAP scaffolds to study the impact of spatial confinement (SC) on both cellular behaviors and the host response to biomaterials. Despite previous studies showing that pore size and SC influence cellular phenotypes, including mitigating the macrophage inflammatory response, there is still a gap in knowledge regarding how SC within a biomaterial modulates immune cell recruitment in vivo in wounds and implants. Thus, we studied the immune cell profile within confined and unconfined biomaterials using small (40 m), medium (70 m), and large (130 m) diameter spherical microgels, respectively. We discovered that MAP scaffolds imparted regenerative wound healing with an IgG1-biased Th2 response. MAP scaffolds generated from 130 m diameter microgels have a median pore size that can accommodate [~]40 {micro}m diameter spheres induced a more balanced pro-regenerative macrophage response and better wound healing outcomes with more mature collagen regeneration and reduced levels of inflammation.

bioengineering↗

bioRxiv2022

Spatial confinement modulates macrophage response in microporous scaffolds

Macrophages are essential in the initiation, maintenance, and transition of inflammatory processes like foreign body response and wound healing. Mounting evidence suggests that physical factors also modulate macrophage activation in vitro and in vivo. 2D in vitro systems have demonstrated that constraining macrophages to small areas or channels modulates their phenotypes and changes their responses to known inflammatory agents such as lipopolysaccharide. However, how dimensionality and pore size affect macrophage phenotype is less explored. In this work, we studied the change in M1/M2 polarization when macrophages were confined in microporous annealed particle scaffolds (MAP), which are granular hydrogels generated from annealed spherical microgels. We engineered three types of MAP gels comprising 40, 70, and 130 {micro}m diameter particle sizes, respectively. Particles sizes were selected using outputs from software LOVAMAP that analyzes the characteristics of 3-D pores in MAP gels. Since the size of building block particle correlates with pore size inside the final scaffolds, our three scaffold types allowed us to study how the degree of spatial confinement modulated the behavior of embedded macrophages. Spatially confining macrophages in scaffolds with pore size on the scale of cells led to a reduced level of the inflammatory response, which was correlated with a change in cell morphology and motility.

bioengineering↗

bioRxiv2022

Void volume fraction of granular scaffolds

Void volume fraction (VVF) of granular biomaterials is a global measurement frequently used to characterize void space. There is currently no gold standard for measuring the VVF of granular scaffolds made in lab. To help the biomaterials field, we provide a library of different simulated scaffolds with known VVF for easy look-up. We use our simulated data to explore the relationship between microscope magnification and VVF, and we study the accuracy of approximating VVF using 2-D z-slice images, which is the most common approach for computing VVF of real scaffolds. Lastly, we test four different approaches for computing VVF using microscope images of real granular scaffolds and reveal that VVF may be an unreliable descriptor.

bioengineering↗

bioRxiv2021

The skeletal muscle circadian clock regulates sarcomere length homeostasis through titin splicing

Circadian rhythms are maintained by a cell autonomous, transcriptional-translational feedback loop known as the molecular clock. While previous research suggests a role of the molecular clock in regulating skeletal muscle structure and function, no mechanisms have connected the molecular clock to sarcomere filaments. Utilizing inducible, skeletal muscle specific, Bmal1 knockout (iMSBmal1-/-) mice, we showed that knocking out skeletal muscle clock function alters titin isoform expression using RNAseq, LC-MS, and SDS-VAGE. This alteration in titins spring length resulted in sarcomere length heterogeneity. We demonstrate the direct link between altered titin splicing and sarcomere length in vitro using U7 snRNPs that truncate the region of titin altered in iMSBmal1-/- muscle. We identified a mechanism whereby the skeletal muscle clock regulates titin isoform expression through transcriptional regulation of Rbm20, a potent splicing regulator of titin. Lastly, we used an environmental model of circadian rhythm disruption and identified significant down-regulation of Rbm20 expression. Our findings demonstrate the importance of the skeletal muscle circadian clock in maintaining titin isoform through regulation of RBM20 expression. Because circadian rhythm disruption is a feature of many chronic diseases, our results highlight a novel pathway that could be targeted to maintain skeletal muscle structure and function in a range of pathologies.

physiology↗

bioRxiv2020

The SAGE genetic toolkit enables highly efficient, iterative site-specific genome engineering in bacteria

Sustainable enhancements to crop productivity and increased resilience to adverse conditions are critical for modern agriculture, and application of plant growth promoting rhizobacteria (PGPR) is a promising method to achieve these goals. However, many desirable PGPR traits are highly regulated in their native microbe, limited to certain plant rhizospheres, or insufficiently active for agricultural purposes. Synthetic biology can address these limitations, but its application is limited by availability of appropriate tools for sophisticated, high-throughput genome engineering that function in environments where selection for DNA maintenance is impractical. Here we present an orthogonal, Serine-integrase Assisted Genome Engineering (SAGE) system, which enables iterative, site-specific integration of up to 10 different DNA constructs at efficiency on par or better than replicating plasmids. SAGE does not require use of replicating plasmids to deliver recombination machinery, and employs a secondary serine-integrase to excise and recycle selection markers. Furthermore, unlike the widely utilized pBBR1 origin, DNA transformed using SAGE is stable without selection. We highlight SAGEs utility by constructing a 287-member constitutive promoter library with a [~]40,000-fold dynamic range in P. fluorescens SBW25. We show that SAGE functions robustly in diverse - and {gamma}-proteobacteria, thus providing evidence that it will be broadly useful for engineering industrial or environmental bacteria.

synthetic biology↗

bioRxiv2017

Transcriptional profiling reveals extraordinary diversity among skeletal muscle tissues

Skeletal muscle comprises a family of diverse tissues with highly specialized morphology, function, and metabolism. Many acquired diseases - including HIV, COPD, cancer cachexia, critical illness myopathy, and sepsis - affect specific muscles while sparing others. Even monogenic muscular dystrophies tend to selectively affect certain muscle groups, despite their causative genetic mutations being present in all tissues. These observations suggest that factors intrinsic to muscle tissues influence their susceptibility to various disease mechanisms. Nevertheless, most studies have not addressed transcriptional diversity among skeletal muscles. Here we use RNA sequencing (RNA-seq) to profile global mRNA expression in a wide array of skeletal, smooth, and cardiac muscle tissues from mice and rats. Our data set, MuscleDB, reveals extensive transcriptional diversity, with greater than 50% of transcripts differentially expressed among skeletal muscle tissues. This diversity is only partly explained by fiber type composition and developmental history, suggesting that specialized transcriptional profiles establish the functional identity of muscle tissues. We find conservation in the transcriptional profiles across species as well as between males and females, indicating that these data may be useful in predicting gene expression in related species. Notably, thousands of differentially expressed genes in skeletal muscle are associated with human disease, and hundreds of these genes encode targets of drugs on the market today. We detect mRNA expression of hundreds of putative myokines that may underlie the endocrine functions of skeletal muscle. In addition to demonstrating the intrinsic diversity of skeletal muscles, these data provide a resource for generating testable hypotheses regarding the mechanisms that establish differential disease susceptibility in muscle.\n\nSignificance StatementSkeletal muscles are a diverse family of tissues with a common contractile function but divergent morphology, development, and metabolism. One need only reflect on the different functions of limb muscles and the diaphragm to realize the highly specialized nature of these tissues. Nevertheless, every study of global gene expression has analyzed at most one representative skeletal muscle. Here we measure gene expression from 11 different skeletal muscles in mice and rats. We show that there is no such thing as a representative skeletal muscle, as gene expression profiles vary widely among the tissues analyzed. These data are an important resource for pharmacologists, tissue engineers, and investigators studying the mechanisms of cellular specialization.

systems biology↗