Biology subjects

Fuller, G. G.

Publications and source records attributed to Fuller, G. G..

6 recordsLinked to original sources

bioRxiv2025

Multidirectional alignment of collagen fibers to guide cell orientation in 3D-printed tissue

Natural tissue comprises fibrous proteins with complex fiber alignment patterns. Here, we develop a reproducible method to fabricate biomimetic scaffolds with patterned fiber alignment along multiple orientations. While extrusion-based approaches are commonly used to align fibrous polymers in a single orientation parallel to the direction of flow, we hypothesized that extrusion-based 3D printing could be utilized to achieve more complex patterns of fiber alignment. Specifically, we show control of lateral spreading of a printed filament can induce fiber alignment that is either parallel or perpendicular to the flow direction. Theoretical prediction of the printing parameters that control fiber orientation was experimentally validated using a collagen biomaterial ink. The velocity ratio of the printhead movement relative to the ink extrusion rate was found to dictate collagen fiber alignment, allowing for the informed fabrication of collagen scaffolds with prescribed patterns of fiber alignment. For example, controlled variation of the ink extrusion rate during a single print resulted in scaffolds with specified regions of both parallel and perpendicular collagen fiber alignment. Human corneal mesenchymal stromal cells seeded onto the printed scaffolds adopted a spread morphology that aligned with the underlying collagen fiber patterns. This technique worked well for filaments either printed into air or extruded within a support bath using embedded 3D printing, enabling the fabrication of 3D structures with aligned collagen fibers. Taken together, this work demonstrates a theoretical and experimental framework to achieve the reproducible fabrication of 3D printed structures with controlled collagen fiber patterns that guide cellular alignment.

bioengineering↗

bioRxiv2023

Air-Liquid Interface Induced Epithelial Delamination

Many epithelial tissues reside at air-liquid interfaces, as exemplified by the ocular epithelium, oral mucosa, and alveolar epithelium. The interfacial tension across the epithelial tissues imposes a mechanical challenge to the homeostasis of the tissue. However, the interplay between interfacial properties and homeostasis in biological samples has been overlooked due to a lack of suitable measurement methods and theoretical developments. Here we described a surprising observation in which the surface energy at cell-air interface is sufficient to delaminate a stratified ocular epithelium from its substrate. We demonstrated that the interfacial tension at the epithelium-fluid interfaces can be measured using a modified Schultz method. The measured value is conceptually and numerically distinctive to the tensile modulus measured by deformation-based methods, such as micropippeting and tissue surface tensiometers. Furthermore, a mechanical analysis at the cell-air-liquid triple line during the delamination process revealed a strain hardening behavior of the epithelial layers. Finally, perturbations on different junctional protein complexes revealed that a delicate balance among cortical tension, focal adhesion, and cell-liquid interfacial tension is required for the epithelial tissue mechanical stability.

biophysics↗

bioRxiv2022

Stable High-Concentration Monoclonal Antibody Formulations Enabled by an Amphiphilic Copolymer Excipient

Monoclonal antibodies are a staple in modern pharmacotherapy. Unfortunately, these biopharmaceuticals are limited by their tendency to aggregate in formulation, resulting in poor stability and often requiring low concentration drug formulations. Moreover, existing excipients designed to stabilize these formulations are often limited by their toxicity and tendency to form particles such as micelles. Here, we demonstrate the ability of a simple "drop-in", amphiphilic copolymer excipient to enhance the stability of high concentration formulations of clinically-relevant monoclonal antibodies without altering their pharmacokinetics or injectability. Through interfacial rheology and surface tension measurements, we demonstrate that the copolymer excipient competitively adsorbs to formulation interfaces. Further, through determination of monomeric composition and retained bioactivity through stressed aging, we show that this excipient confers a significant stability benefit to high concentration antibody formulations. Finally, we demonstrate that the excipient behaves as an inactive ingredient, having no significant impact on the pharmacokinetic profile of a clinically relevant antibody in mice. This amphiphilic copolymer excipient demonstrates promise as a simple formulation additive to create stable, high concentration antibody formulations, thereby enabling improved treatment options such as a route-of-administration switch from low concentration intravenous (IV) to high concentration subcutaneous (SC) delivery while reducing dependence on the cold chain.

bioengineering↗

bioRxiv2022

Systematic Characterization of Double Emulsion Droplets for Biological Applications

Double emulsion droplets (DEs) are water/oil/water droplets that can be sorted via Fluorescence-Activated Cell Sorting (FACS), allowing for new opportunities in high-throughput cellular analysis, enzymatic screening, and synthetic biology. These applications require stable, uniform droplets with predictable microreactor volumes. However, predicting DE droplet size, shell thickness, and stability as a function of flow rate has remained challenging for monodisperse single core droplets and those containing biologically-relevant buffers, which influence bulk and interfacial properties. As a result, developing novel DE-based bioassays has typically required extensive initial optimization of flow rates to find conditions that produce stable droplets of the desired size and shell thickness. To address this challenge, we conducted systematic size parameterization quantifying how differences in flow rates and buffer properties (viscosity and interfacial tension at water/oil interfaces) alter droplet size and stability, across 6 inner aqueous buffers used across applications such as cellular lysis, microbial growth, and drug delivery, quantifying the size and shell thickness of >22,000 droplets overall. We restricted our study to stable single core droplets generated in a 2-step dripping-dripping formation regime in a straightforward PDMS device. Using data from 138 unique conditions (flow rates and buffer composition), we also demonstrated that a recent physically-derived size law of Wang et al1 can accurately predict double emulsion shell thickness for >95% of observations. Finally, we validated the utility of this size law by using it to accurately predict droplet sizes for a novel bioassay that requires encapsulating growth media for bacteria in droplets. This work has the potential to enable new screening-based biological applications by simplifying novel DE bioassay development.

bioengineering↗

bioRxiv2021

Engineering Insulin Cold Chain Resilience to Improve Global Access

There are 150 million people with diabetes worldwide who require insulin replacement therapy and the prevalence of diabetes is rising fastest in middle and low-income countries. Current formulations require costly refrigerated transport and storage to prevent loss of insulin integrity. This study shows the development of simple "drop-in" amphiphilic copolymer excipients to maintain formulation integrity, bioactivity, pharmacokinetics and pharmacodynamics for over 6 months when subjected to severe stressed aging conditions that cause current commercial formulation to fail in under 2 weeks. Further, when these copolymers are added to Humulin R (Eli Lilly) in original commercial packaging they prevent insulin aggregation for up to 4 days at 50 {degrees}C compared to less than 1 day for Humulin R alone. These copolymers demonstrate promise as simple formulation additives to increase the cold chain resilience of commercial insulin formulations, thereby expanding global access to these critical drugs for treatment of diabetes.

bioengineering↗

bioRxiv2019

RNA promotes phase separation of glycolysis enzymes into yeast G bodies in hypoxia

In hypoxic stress conditions, glycolysis enzymes assemble into singular cytoplasmic granules called glycolytic (G) bodies. Formation of G bodies in yeast is correlated with increased glucose consumption and cell survival. However, the physical properties and organizing principles that define G body formation are unclear. We demonstrate that glycolysis enzymes are non-canonical RNA binding proteins, sharing many common mRNA substrates that are also integral constituents of G bodies. Tethering a G body component, the beta subunit of the yeast phosphofructokinase, Pfk2, to nonspecific endoribonucleases reveals that RNA nucleates G body formation and subsequent maintenance of G body structural integrity. Consistent with a phase separation mechanism of G body formation, recruitment of glycolysis enzymes to G bodies relies on multivalent homotypic and heterotypic interactions. Furthermore, G bodies can fuse in live cells and are largely insensitive to 1,6-hexanediol treatment, consistent with a hydrogel-like state in its composition. Taken together, our results elucidate the biophysical nature of G bodies and demonstrate that RNA nucleates phase separation of the glycolysis machinery in response to hypoxic stress.

cell biology↗