Biology subjects

Ferguson, S. J.

Publications and source records attributed to Ferguson, S. J..

6 recordsLinked to original sources

bioRxiv2026

Dynamic Compression Platform for Live Imaging of Scaffold-Transmitted Cellular Mechanoresponses

Mechanical characterization of biomaterial scaffolds is essential to evaluate their capacity to meet the functional demands of target tissues in tissue engineering and regenerative medicine applications. Scaffolds designed to interface with living tissues must support the transmission of mechanical cues to resident cells and stimulate mechanosignaling pathways that are essential to their function. In joints, bone and cartilage cells act as primary mechanosensors, converting mechanical stimuli into biochemical signals that regulate tissue homeostasis and remodelling. Therefore, evaluating cellular mechanoresponses to scaffold-transmitted compression in vitro can inform the development of functional tissue-engineered constructs. For example, poly({epsilon}-caprolactone) (PCL) scaffolds are highly relevant for bone and cartilage tissue engineering due to their biocompatibility, stable mechanical properties and slow degradation. Here, we applied a custom-built device to study compression-induced mechanosignaling in MC3T3-E1 pre-osteoblast cells. The device is composed of a polydimethylsiloxane (PDMS) pillar, a force-sensing load cell, and a piezoelectric linear track. A protocol is described in which MC3T3-E1 cells are repeatedly compressed, while in parallel live tracking of force measurements and live imaging of intracellular calcium dynamics in MC3T3-E1 cells are recorded. PCL scaffolds fabricated by melt electrowriting (MEW) were subsequently integrated into the platform. Scaffold-transmitted compression triggered dynamic increases in cytosolic calcium; in MC3T3-E1 cells located directly under the PCL microfibers, but also in cells located in the interfiber spaces. This device and workflow facilitate in vitro investigations of real-time cellular mechanoresponses to dynamic compression applied with biomaterial scaffolds, and provides a testing platform for evaluating the mechanotransductive properties of scaffolds intended for tissue engineering applications.

bioengineering↗

bioRxiv2026

A humanized Aβ mouse model reveals E4-dependent cognitive impairments, microglial activation, and cerebrovascular dysfunction

Apolipoprotein E4 (E4) increases the risk of Alzheimers disease (AD) by up to 12-fold. However, understanding of the mechanisms underlying this increased risk has been limited by a lack of preclinical models that accurately reflect the effects of E4 in the presence of humanized non-mutant amyloid-{beta} precursor protein (hA{beta}PP). Therefore, we studied novel humanized APOE and hA{beta}PP mice to investigate the contributions of the E4 genotype to cognitive, inflammatory, and vascular dysfunction, specifically comparing male and female E3/hA{beta}PP and E4/hA{beta}PP mice. E4/hA{beta}PP mice exhibited impaired nest-building behavior and novel object recognition compared with E3/hA{beta}PP mice. Microglial content was higher in E4/hA{beta}PP mice, whereas astrocyte content was not different across groups. E4/hA{beta}PP mice had greater carotid and cerebral artery stiffness, and higher collagen I content in cerebral arteries than E3/hA{beta}PP mice. Under static pressure, cerebral artery endothelium-dependent and endothelium-independent vasodilation were similar across genotypes. However, high pulse pressure selectively impaired cerebral artery endothelial function in E4/hA{beta}PP mice, with the greatest impairment observed in females. The E4/hA{beta}PP mice also exhibited higher cortical expression of Nox2 and Sod1 and elevated cerebral artery Il1b expression. As such, E4/hA{beta}PP mice exhibit convergent cognitive, inflammatory, and vascular abnormalities that recapitulate several features of AD. Elevated pulse pressure revealed an E4-dependent vulnerability of the cerebral vasculature, suggesting that vascular stress may be an important contributor to disease risk. Together, our findings support the use of the APOExhA{beta}PP model to investigate the mechanisms by which E4 promotes vascular dysfunction, neuroinflammation, and cognitive impairment in AD.

physiology↗

bioRxiv2023

A self-renewing biomimetic skeletal muscle construct engineered using induced myogenic progenitor cells

Skeletal muscle is a highly organized and regenerative tissue that maintains its homeostasis primarily by activation and differentiation of muscle stem cells. Mimicking an in vitro skeletal muscle differentiation program that contains self-renewing adult muscle stem cells and aligned myotubes has been challenging. Here, we set out to engineer a biomimetic skeletal muscle construct that can self-regenerate and produce aligned myotubes using induced myogenic progenitor cells (iMPCs), a heterogeneous culture consisting of skeletal muscle stem, progenitor and differentiated cells. Utilizing electrospinning, we fabricated polycaprolactone (PCL) substrates that enabled iMPC-differentiation into aligned myotubes by controlling PCL fiber orientation. Newly-conceived constructs contained highly organized multinucleated myotubes in conjunction with self-renewing muscle stem cells, whose differentiation capacity was augmented by Matrigel supplementation. Additionally, we demonstrate using single cell RNA-sequencing (scRNA-seq) that iMPC-derived constructs faithfully recapitulate a step-wise myogenic differentiation program. Notably, when the constructs were subjected to a damaging myonecrotic agent, self-renewing muscle stem cells rapidly differentiated into aligned myotubes, akin to skeletal muscle repair in vivo. Taken together, we report on a novel in vitro system that mirrors myogenic regeneration and muscle fiber alignment, and can serve as a platform to study myogenesis, model muscular dystrophies or perform drug screens.

bioengineering↗

bioRxiv2023

3D-Printed LEGO(R)-inspired Titanium Scaffolds for Patient-Specific Regenerative Medicine

Despite the recent advances in 3D-printing, it is difficult to fabricate implants that optimally fit a defect size/shape. There are some approaches to resolve this issue, such as patient-specific implant based on CT images, however, it is labor-intensive and costly. Especially in developing countries, affordable treatments are required, while still not excluding these patient groups from manufacturing advances. Here, a SLM 3D-printing strategy was used to fabricate a hierarchical, Assemblable Titanium Scaffold(ATS), which can be manually assembled in any shape or size with ease. A surgeon can create a scaffold that would fit to the defect right before the implantation during the surgery. Additionally, the direct inclusion of micro- and macroporous structures via 3D-printing, as well as a double acid-etched surface treatment(ST) in the ATS, ensure improved nutrient flow and cellular activity. Different structures were designed, 3D-printed and then surface treated for the ST groups. Both individual and stacked ATS have sufficient mechanical properties to withstand physiological loading, and the porous groups resulted enhanced cell proliferation, mineralizaton and osteogenesis compared to non-porous group. Furthermore, successful cell attachment and migration between the assembled ATS were observed. Finally, we demonstrate possible medical applications that reveal the potential of the ATS through assembly.

bioengineering↗

bioRxiv2022

Molecular determinants of dynamic protein-protein interactions in the functional cycle of the membrane protein DsbD

Molecular recognition is of central importance in biology. The molecular determinants shaping recognition of one protein domain by another are incompletely understood, especially in the context of the complex function of molecular machines. Here, we combine NMR experiments and molecular dynamics simulations to elucidate the determinants of recognition of the C-terminal (cDsbD) domain of the transmembrane reductant conductor DsbD by its cognate partner, the N-terminal domain of the protein (nDsbD). As part of the natural cycle of this oxidoreductase, which effectively transfers electrons from the cytoplasm to the periplasm of Gram-negative bacteria, cDsbD and nDsbD toggle between oxidised and reduced states, something that modulates the affinity of the domains for each other and prevents otherwise unproductive reactions. We find that the redox state of cDsbD determines the dissociation rate of cDsbD-nDsbD complexes. Molecular dynamics simulations demonstrate how the redox-state of the active site determines the stability of inter-domain hydrogen bonds and thus the dissociation rate. AlphaFold modelling and atomistic molecular dynamics simulations of full-length DsbD in a realistic bacterial membrane again highlights the close proximity of the periplasmic domains and the importance of tuning the strength of the interactions of the periplasmic domains to enable electron transfer to cognate periplasmic partners such as CcmG. Our AlphaFold models are consistent with in vivo functional assays of DsbD mutants, which together help to reveal for the first-time a putative binding site for thioredoxin on the cytoplasmic side of DsbD.

biophysics↗

bioRxiv2019

Local frustration determines loop opening in protein-protein association

Local structural frustration, the existence of mutually exclusive competing interactions, may explain why some proteins are dynamic while others are rigid. Frustration is thought to underpin biomolecular recognition and the flexibility of protein binding sites. Here we show how a small chemical modification, the oxidation of two cysteine thiols to a disulfide bond, during the catalytic cycle of the N-terminal domain of the key bacterial oxidoreductase DsbD (nDsbD), introduces frustration ultimately influencing protein function. In oxidized nDsbD, local frustration disrupts the packing of the protective cap-loop region against the active site allowing loop opening. By contrast, in reduced nDsbD the cap loop is rigid, always protecting the active-site thiols from the oxidizing environment of the periplasm. Our results point towards an intricate coupling between the dynamics of the active-site cysteines and of the cap loop which modulates the association reactions of nDsbD with its partners resulting in optimized protein function.

biophysics↗