Garry P. Duffy’s research while affiliated with Royal College of Surgeons in Ireland and other places

Extracellular vesicles (EVs) are versatile transporters of genetic cargo with enormous potential in the therapeutic setting. Scalable production of EVs, and routes to overcome rapid clearance are required. Biocompatible hydrogels may support precise, localized delivery of EVs to target sites. This study aimed to establish sustained production of EVs in a scalable 3D dynamic bioreactor and to fabricate hydrogels using tyramine-modified hyaluronic acid (HA-TA) to study EV integration and release patterns. MDA-MB-231 cells transduced with lentiviral GFP fused with CD63, were cultured in a 20kD dynamic hollow fiber bioreactor and GFP-EVs harvested over five weeks. GFP-EVs were characterized by Nanoparticle Tracking Analysis(NTA), Western Blot(WB) and Transmission Electron Microscopy(TEM). Tyramine modified hyaluronic acid(HA-TA) hydrogels were formulated via enzymatic crosslinking using hydrogen peroxide and horseradish peroxidase, to investigate EV release patterns in static and dynamic conditions. Hydrogel swelling was recorded at 1-72 hrs and hydrogels were loaded with GFP-EVs to assess distribution and release by Scanning Electron Microscopy(SEM) and NTA respectively. GFP-EV uptake was assessed by confocal microscopy. Longitudinal GFP expression was demonstrated in transduced cells and released EVs throughout bioreactor culture. TEM and NTA demonstrated successful isolation of EVs of 30-200 nm in size with intact lipid bilayers (average 4x109 EVs/harvest). Initial harvests exhibited subpopulations of larger EVs, which disappeared upon serum withdrawal. WB verified the presence of EV markers CD63, TSG101, and CD81. HA-TA hydrogels were successfully formed and swelling assays revealed the requirement for higher concentrations of HA-TA and crosslinkers for scaffold stability and continued swelling. GFP-EVs were successfully incorporated into the hydrogels with variable release patterns observed over time, depending on EV concentration and hydrogel formulation. EV clusters in hydrogels were visualized by SEM. Investigation of GFP-EV release patterns under static and dynamic conditions highlighted a significant increase in release under fluid flow conditions. Efficient transfer of released EVs to recipient cells was also demonstrated in vitro. The data demonstrate the potential for scalable production of engineered EVs in serum free conditions and subsequent incorporation into HA-TA hydrogels for sustained release. These biocompatible hydrogels hold promise for tuneable delivery of therapeutic EVs in a variety of disease settings.

Changes in the microstructure of the aortic wall precede the progression of various aortic pathologies, including aneurysms and dissection. Current clinical decisions with regards to surgical planning and/or radiological intervention are guided by geometric features, such as aortic diameter, since clinical imaging lacks tissue microstructural information. The aim of this proof‐of‐concept work is to investigate a non‐invasive imaging method, diffusion tensor imaging (DTI), in ex vivo aortic tissue to gain insights into the microstructure. This study examines healthy, aneurysm and a type B chronic dissection aortae, via DTI. DTI‐derived metrics, such as the fractional anisotropy, mean diffusivity, helical angle and tractography, were examined in each morphology. The results from this work highlighted distinct differences in fractional anisotropy (healthy, 0.24 ± 0.008; aneurysmal, 0.19 ± 0.002; dissected, 0.13 ± 0.006) and a larger variation in the helical angle in the dissected aorta compared to healthy (39.28 ± 11.93° vs. 26.12 ± 4.60°, respectively). These differences were validated by histological characterisation. This study demonstrates the sensitivity of DTI to pathological changes in aortic tissue, highlighting the potential of this methodology to provide improved clinical insight.

Therapeutic proteins, the fastest growing class of pharmaceuticals, are subject to rapid proteolytic degradation in vivo, rendering them inactive. Sophisticated drug delivery systems that maintain protein stability, prolong therapeutic effects, and reduce administration frequency are urgently required. Herein, a mechanoresponsive hydrogel is developed contained within a soft robotic drug delivery (SRDD) device. In a step‐change from previously reported systems, pneumatic actuation of this system releases the cationic therapeutic protein Vascular Endothelial Growth Factor (VEGF) in a bioactive form which is required for therapeutic angiogenesis, the growth of new blood vessels, in numerous clinical conditions. The ability of the SRDD device to release bioactive VEGF in a spatiotemporal manner from the hydrogel is tested in diabetic rats – a model in which angiogenesis is difficult to stimulate. Daily actuation of the SRDD device in the diabetic rat model significantly increased cluster of differentiation 31+ (CD31+) blood vessel number (p = 0.0335) and the diameter of alpha‐smooth muscle actin+ (α‐SMA+) blood vessels (p = 0.0025) compared to passive release of VEGF from non‐actuated devices. The SRDD device combined with the mechanoresponsive hydrogel offers the potential to deliver an array of bioactive therapeutics in a spatiotemporal manner to mimic their natural release in vivo.

Transplantation of donor islets of Langerhans is a potential therapeutic approach for patients with diabetes mellitus; however, its success is limited by islet death and dysfunction during the initial hypoxic conditions at the transplantation site. This highlights the need to support the donor islets in the days post‐transplantation until the site is vascularized. It was previously demonstrated that the extracellular matrix (ECM) proteins nidogen‐1 (NID1) and decorin (DCN) improve the functionality and survival of the β‐cell line, EndoC‐βH3, and the viability of human islets post‐isolation. To advance the use of these ECM proteins toward a clinical application and elucidate the mechanisms of action in primary islets, the study assesses the effects of ECM proteins NID1 and DCN on isolated human donor islets cultured in normoxic and hypoxic conditions. NID1‐ and DCN‐treatment restore β‐cell functionality of human donor islets in a hypoxic environment through upregulation of genes involved in glycolytic pathways and reducing DNA fragmentation in hypoxic conditions comparable to normoxic control islets. The results demonstrate that the utilization of NID1 or DCN with islets of Langerhans may have the potential to overcome the hypoxia‐induced cell death observed post‐transplantation and improve transplant outcomes.

Primary lung cancer is the leading cause of cancer‐related death worldwide. Recent statistics estimate 1.8 million new cases and 1.6 million deaths per year. Despite significant advances in surgery, chemotherapy, and radiotherapy, the 5‐year survival rate remains at just 18%. Thermal ablation is an established treatment that is safe and effective. It refers to the local application of controlled heat to induce irreversible cell injury and ultimately tumor apoptosis and necrosis. This approach is particularly suitable for the treatment of small tumors in patients who are not suitable for surgery. However, the lack of suitable benchtop models to assess ablation devices complicates prognostic prediction and the development of effective treatment strategies. In this context, the development of a 3D lung tumor model is presented using a cancer cell‐laden hydrogel composed of 2% hyaluronic acid–tyramine (HA‐TA) and 0.5% agarose, resulting in a tumor mimetic of clinically relevant size. This innovative model provides a valuable platform for testing ablation devices and provides insights that can significantly improve prognostic ability and refine treatment approaches for lung cancer patients.

Our understanding of cardiac remodeling processes due to left ventricular pressure overload derives largely from animal models of aortic banding. However, these studies fail to enable control over both disease progression and reversal, hindering their clinical relevance. Here, we describe a method for progressive and reversible aortic banding based on an implantable expandable actuator that can be finely tuned to modulate aortic banding and debanding in a rat model. Through catheterization, imaging, and histologic studies, we demonstrate that our platform can recapitulate the hemodynamic and structural changes associated with pressure overload in a controllable manner. We leveraged soft robotics to enable noninvasive aortic debanding, demonstrating that these changes can be partly reversed because of cessation of the biomechanical stimulus. By recapitulating longitudinal disease progression and reversibility, this animal model could elucidate fundamental mechanisms of cardiac remodeling and optimize timing of intervention for pressure overload.

Implantable medical devices that can facilitate therapy transport to localized sites are being developed for a number of diverse applications, including the treatment of diseases such as diabetes and cancer,...