Nanoscale liposomes have been extensively researched and employed clinically for the delivery of biologically active compounds, including chemotherapy drugs and vaccines, offering improved pharmacokinetic behaviour and therapeutic outcomes. Traditional laboratory-scale production methods often suffer from limited control over liposome properties (e.g., size and lamellarity) and rely on laborious multistep procedures, which may limit pre-clinical research developments and innovation in this area. The widespread adoption of alternative, more controllable microfluidic-based methods is often hindered by complexities and costs associated with device manufacturing and operation, as well as the short device lifetime and the relatively low liposome production rates in some cases. In this study, we demonstrated the production of liposomes comprising therapeutically relevant lipid formulations, using a cost-effective 3D-printed reactor-in-a-centrifuge (RIAC) device. By adjusting formulation- and production-related parameters, including the concentration of polyethylene glycol (PEG), temperature, centrifugation time and speed, and lipid concentration, the mean size of the produced liposomes could be tuned in the range of 140 to 200 nm. By combining selected experimental parameters, the method was capable of producing liposomes with a therapeutically relevant mean size of ~174 nm with narrow size distribution (polydispersity index, PDI ~0.1) at a production rate of >8 mg/min. The flow-through method proposed in this study has potential to become an effective and versatile laboratory-scale approach to simplify the synthesis of therapeutic liposomal formulations.

Urinary tract infections (UTI) remain a major global health burden, with high recurrence despite antibiotic treatment. The escalating prevalence of antimicrobial resistance further compromises therapeutic efficacy, contributing to an estimated 260,000 deaths annually. Conventional in vitro susceptibility assays often fail to predict clinical outcomes, underscoring the urgent need for physiologically relevant infection models. Here, we examined how microenvironmental complexity shapes uropathogenic Escherichia coli (UPEC) responses to antibiotics and bacteriophages using: human urine, a three-dimensional urothelial microtissue model (3D-UHU), and a novel mesofluidic system (P-FLO) that introduces physiologically relevant flow dynamics to the 3D-UHU. P-FLO was engineered from cost-effective 3D-printed components compatible with standard Transwell systems. Among the antibiotics tested, nitrofurantoin exhibited the greatest potency in minimum inhibitory concentration assays, but it failed to fully eradicate infection within the more physiological 3D-UHU model. A bacteriophage cocktail (LCPR1) showed markedly reduced activity in urine compared with nutrient-rich media, highlighting the influence of infection-site conditions. In contrast, in 3D-UHU, LCRP1 modulated host responses without reducing bacterial burden. Combination therapy (nitrofurantoin + LCPR1) eliminated planktonic bacteria under static conditions but offered no added benefit against adherent or intracellular populations relative to antibiotic monotherapy. Incorporating flow revealed additional layers of complexity, where shear stress induced bacterial elongation and attachment and altered drug performance, diminishing the efficacy of nitrofurantoin and combination therapy against planktonic populations despite increased drug exposure. Together, these findings demonstrate that the bladder microenvironment and its mechanical forces modulate host-pathogen interactions and profoundly influence UPEC infection dynamics and therapeutic outcomes, emphasizing the need for advanced, physiologically informed models to guide treatment strategies in the post-antibiotic era.

Bone disorders and skeletal defects represent a significant clinical challenge, often requiring transplantation techniques limited by donor site morbidity and insufficient regenerative potential. Tissue engineering and regenerative medicine (TERM) strategies using 3D bioprinting have emerged as promising alternatives, but their efficacy is limited by the difficulty of directing stem cell differentiation in a controlled and reproducible manner. To address this limitation, we are proposing 3D bone printing via ultrasound-mediated osteogenic differentiation of stem cells (referred to here as ‘3DBonUS’). This biofabrication approach integrates low-intensity pulsed ultrasound (LIPUS) with a microfluidic-assisted 3D bioprinting system. This unprecedented approach enables biophysical stimulation of human bone marrow stromal cells (HBMSCs) during the fabrication of scaffolds, promoting osteogenic differentiation without the need for extensive post-fabrication treatments. In addition, the incorporation of microbubbles (MBs) enhanced the effects of LIPUS by amplifying mechanical signals at the cellular level. Our results revealed that the 3DBonUS system significantly upregulated key osteogenic markers (RUNX-2, ALP, COL1A1, BMP-2, OCN and OPN) as confirmed by immunofluorescence and RT-qPCR analysis. Moreover, the LIPUS-treated constructs showed a significant (p<0.05) increase in alkaline phosphatase (ALP) activity and calcium deposition, indicating enhanced mineralisation. The biofabricated constructs maintained high cell viability while exhibiting improved osteogenic differentiation, surpassing traditional 3D bioprinting approaches in both efficiency and efficacy. The 3DBonUS strategy represents a new modality in skeletal TERM, combining biofabrication with targeted mechanical stimulation, with potential for scalability of scaffold manufacturing and clinical application. Future studies will aim to validate the functional skeletal scaffolds in vivo to assess their regenerative potential, with the goal of advancing patient-specific bone implants with enhanced osteogenic properties.