Bioactive coating provides antimicrobial protection through immunomodulation and phage therapeutics.

Medical implant-associated infections (IAI) is a growing threat to patients undergoing implantation surgery. IAI prevention typically relies on medical implants endowed with bactericidal properties achieved through surface modifications with antibiotics. However, the clinical efficacy of this traditional paradigm remains suboptimal, often necessitating revision surgery and posing potentially lethal consequences for patients. To bolster the existing anti-IAI arsenal, we propose herein a chitosan-based bioactive coating, i.e., ChitoAntibac, which exerts bacteria-inhibitory effects either through immune modulation or phage-directed microbial clearance, without relying on conventional antibiotics. The immuno-stimulating effects and phage-induced bactericidal properties can be tailored by engineering the loading dynamic of macrophage migration inhibitory factor (MIF), which polarizes macrophages towards the proinflammatory subtype (M1) with enhanced bacterial phagocytosis, and Staphylococcal Phage K, resulting in rapid and targeted pathogenic clearance (>99.99%) in less than 8 h. Our innovative antibacterial coating opens a new avenue in the pursuit of effective IAI prevention through immuno-stimulation and phage therapeutics.

Nanoscale Methods for Longitudinal Extraction of Intracellular Contents.

Longitudinal analysis of intracellular contents including gene and protein expression is crucial for deciphering the fundamentally dynamic nature of cells. This offers invaluable insights into complex tissue composition and behavior, and drives progress in disease diagnosis, biomarker discovery, and drug development. Traditional longitudinal analysis workflows, involving the destruction of cells at various timepoints, limit insights to singular moments and fail to account for cellular heterogeneity. Current non-destructive approaches, like temporal modeling with single-cell ribonucleic acid sequencing (RNA-seq) and live-cell fluorescence imaging, either rely on biological assumptions or possess the risk of cellular perturbation. Recent advances in nanoscale technologies for non-destructive intracellular content extraction offer a promising solution to these challenges. These novel methods work at the nanoscale to non-destructively access cellular membranes and can be broadly classified into three mechanisms: tip-facilitated aspiration, membrane-based, and probe-based methods. This perspective focuses on these emerging nanotechnologies for repeated intracellular content extraction. Their potential in longitudinal analysis is discussed, the critical requirements for effective repeated sampling are addressed, and the suitability of each technique for various applications is explored. Furthermore, unresolved challenges in repeated sampling are highlighted to encourage further research in this growing field.

Attenuating Epithelial-to-Mesenchymal Transition in Cancer through Angiopoietin-Like 4 Inhibition in a 3D Tumor Microenvironment Model.

Epithelial-to-mesenchymal transition (EMT) plays a crucial role in metastatic cancer progression, and current research, which relies heavily on 2D monolayer cultures, falls short in recapitulating the complexity of a 3D tumor microenvironment. To address this limitation, a transcriptomic meta-analysis is conducted on diverse cancer types undergoing EMT in 2D and 3D cultures. It is found that mechanotransduction is elevated in 3D cultures and is further intensified during EMT, but not during 2D EMT. This analysis reveals a distinct 3D EMT gene signature, characterized by extracellular matrix remodeling coordinated by angiopoietin-like 4 (Angptl4) along with other canonical EMT regulators. Utilizing hydrogel-based 3D matrices with adjustable mechanical forces, 3D cancer cultures are established at varying physiological stiffness levels. A YAP:EGR-1 mediated up-regulation of Angptl4 expression is observed, accompanied by an upregulation of mesenchymal markers, at higher stiffness during cancer EMT. Suppression of Angptl4 using antisense oligonucleotides or anti-cAngptl4 antibodies leads to a dose-dependent abolishment of EMT-mediated chemoresistance and tumor self-organization in 3D, ultimately resulting in diminished metastatic potential and stunted growth of tumor xenografts. This unique programmable 3D cancer cultures simulate stiffness levels in the tumor microenvironment and unveil Angptl4 as a promising therapeutic target to inhibit EMT and impede cancer progression.

Progress on Electro-Enhancement of Cell Manufacturing.

With the long persistence of complex, chronic diseases in society, there is increasing motivation to develop cells as living medicine to treat diseases ranging from cancer to wounds. While cell therapies can significantly impact healthcare, the shortage of starter cells meant that considerable raw materials must be channeled solely for cell expansion, leading to expensive products with long manufacturing time which can prevent accessibility by patients who either cannot afford the treatment or have highly aggressive diseases and cannot wait that long. Over the last three decades, there has been increasing knowledge on the effects of electrical modulation on proliferation, but to the best of the knowledge, none of these studies went beyond how electro-control of cell proliferation may be extended to enhance industrial scale cell manufacturing. Here, this review is started by discussing the importance of maximizing cell yield during manufacturing before comparing strategies spanning biomolecular/chemical/physical to modulate cell proliferation. Next, the authors describe how factors governing invasive and non-invasive electrical stimulation (ES) including capacitive coupling electric field may be modified to boost cell manufacturing. This review concludes by describing what needs to be urgently performed to bridge the gap between academic investigation of ES to industrial applications.

Automating CAR-T Transfection with Micro and Nano-Technologies.

Cancer poses a significant health challenge, with traditional treatments like surgery, radiotherapy, and chemotherapy often lacking in cell specificity and long-term curative potential. Chimeric antigen receptor T cell (CAR-T) therapy,utilizing genetically engineered T cells to target cancer cells, is a promising alternative. However, its high cost limits widespread application. CAR-T manufacturing process encompasses three stages: cell isolation and activation, transfection, and expansion.While the first and last stages have straightforward, commercially available automation technologies, the transfection stage lags behind. Current automated transfection relies on viral vectors or bulk electroporation, which have drawbacks such as limited cargo capacity and significant cell disturbance. Conversely, micro and nano-tool methods offer higher throughput and cargo flexibility, yet their automation remains underexplored.In this perspective, the progress in micro and nano-engineering tools for CAR-T transfection followed by a discussion to automate them is described. It is anticipated that this work can inspire the community working on micro and nano transfection techniques to examine how their protocols can be automated to align with the growing interest in automating CAR-T manufacturing.

Mechano-Activated Cell Therapy for Accelerated Diabetic Wound Healing.

Chronic diabetic wounds are a significant global healthcare challenge. Current strategies, such as biomaterials, cell therapies, and medical devices, however, only target a few pathological features and have limited efficacy. A powerful platform technology combining magneto-responsive hydrogel, cells, and wireless magneto-induced dynamic mechanical stimulation (MDMS) is developed to accelerate diabetic wound healing. The hydrogel encapsulates U.S. Food and Drug Administration (FDA)-approved fibroblasts and keratinocytes to achieve ∼3-fold better wound closure in a diabetic mouse model. MDMS acts as a nongenetic mechano-rheostat to activate fibroblasts, resulting in ∼240% better proliferation, ∼220% more collagen deposition, and improved keratinocyte paracrine profiles via the Ras/MEK/ERK pathway to boost angiogenesis. The magneto-responsive property also enables on-demand insulin release for spatiotemporal glucose regulation through increasing network deformation and interstitial flow. By mining scRNAseq data, a mechanosensitive fibroblast subpopulation is identified that can be mechanically tuned for enhanced proliferation and collagen production, maximizing therapeutic impact. The "all-in-one" system addresses major pathological factors associated with diabetic wounds in a single platform, with potential applications for other challenging wound types.

Advances in Enantiomer-Dependent Nanotherapeutics.

The nanoscale properties of nanomaterials, especially nanoparticles, including size, shape, and surface charge, have been extensively studied for their impact on nanomedicine. Given the inherent chiral nature of biological systems and their high enantiomeric selectivity, there is rising interest to manipulate the chirality of nanomaterials to enhance their biomolecular interactions and improve nanotherapeutics. Chiral nanostructures are currently more prevalently used in biosensing and diagnostic applications owing to their distinctive physical and optical properties, but they hold great promise for use in nanomedicine. In this Review, we first discuss stereospecific interactions between chiral nanomaterials and biomolecules before comparing the synthesis and characterization methods of chiral nanoparticles and nanoassemblies. Finally, we examine the applications of chiral nanotherapeutics in cancer, immunomodulation, and neurodegenerative diseases and propose plausible mechanisms in which chiral nanomaterials interact with cells for biological manipulation. This Review on chirality is a timely reminder of the arsenal of nanoscale modifications to boost research in nanotherapeutics.

Dynamic Stimulations with Bioengineered Extracellular Matrix-Mimicking Hydrogels for Mechano Cell Reprogramming and Therapy.

Cells interact with their surrounding environment through a combination of static and dynamic mechanical signals that vary over stimulus types, intensity, space, and time. Compared to static mechanical signals such as stiffness, porosity, and topography, the current understanding on the effects of dynamic mechanical stimulations on cells remains limited, attributing to a lack of access to devices, the complexity of experimental set-up, and data interpretation. Yet, in the pursuit of emerging translational applications (e.g., cell manufacturing for clinical treatment), it is crucial to understand how cells respond to a variety of dynamic forces that are omnipresent in vivo so that they can be exploited to enhance manufacturing and therapeutic outcomes. With a rising appreciation of the extracellular matrix (ECM) as a key regulator of biofunctions, researchers have bioengineered a suite of ECM-mimicking hydrogels, which can be fine-tuned with spatiotemporal mechanical cues to model complex static and dynamic mechanical profiles. This review first discusses how mechanical stimuli may impact different cellular components and the various mechanobiology pathways involved. Then, how hydrogels can be designed to incorporate static and dynamic mechanical parameters to influence cell behaviors are described. The Scopus database is also used to analyze the relative strength in evidence, ranging from strong to weak, based on number of published literatures, associated citations, and treatment significance. Additionally, the impacts of static and dynamic mechanical stimulations on clinically relevant cell types including mesenchymal stem cells, fibroblasts, and immune cells, are evaluated. The aim is to draw attention to the paucity of studies on the effects of dynamic mechanical stimuli on cells, as well as to highlight the potential of using a cocktail of various types and intensities of mechanical stimulations to influence cell fates (similar to the concept of biochemical cocktail to direct cell fate). It is envisioned that this progress report will inspire more exciting translational development of mechanoresponsive hydrogels for biomedical applications.

Mechano-responsive hydrogel for direct stem cell manufacturing to therapy.

Bone marrow-derived mesenchymal stem cell (MSC) is one of the most actively studied cell types due to its regenerative potential and immunomodulatory properties. Conventional cell expansion methods using 2D tissue culture plates and 2.5D microcarriers in bioreactors can generate large cell numbers, but they compromise stem cell potency and lack mechanical preconditioning to prepare MSC for physiological loading expected in vivo. To overcome these challenges, in this work, we describe a 3D dynamic hydrogel using magneto-stimulation for direct MSC manufacturing to therapy. With our technology, we found that dynamic mechanical stimulation (DMS) enhanced matrix-integrin ß1 interactions which induced MSCs spreading and proliferation. In addition, DMS could modulate MSC biofunctions including directing MSC differentiation into specific lineages and boosting paracrine activities (e.g., growth factor secretion) through YAP nuclear localization and FAK-ERK pathway. With our magnetic hydrogel, complex procedures from MSC manufacturing to final clinical use, can be integrated into one single platform, and we believe this 'all-in-one' technology could offer a paradigm shift to existing standards in MSC therapy.

Dynamic Magneto-Softening of 3D Hydrogel Reverses Malignant Transformation of Cancer Cells and Enhances Drug Efficacy.

High extracellular matrix stiffness is a prominent feature of malignant tumors associated with poor clinical prognosis. To elucidate mechanistic connections between increased matrix stiffness and tumor progression, a variety of hydrogel scaffolds with dynamic changes in stiffness have been developed. These approaches, however, are not biocompatible at high temperature, strong irradiation, and acidic/basic pH, often lack reversibility (can only stiffen and not soften), and do not allow study on the same cell population longitudinally. In this work, we develop a dynamic 3D magnetic hydrogel whose matrix stiffness can be wirelessly and reversibly stiffened and softened multiple times with different rates of change using an external magnet. With this platform, we found that matrix stiffness increased tumor malignancy including denser cell organization, epithelial-to-mesenchymal transition and hypoxia. More interestingly, these malignant transformations could be halted or reversed with matrix softening (i.e., mechanical rescue), to potentiate drug efficacy attributing to reduced solid stress from matrix and downregulation of cell mechano-transductors including YAP1. We propose that our platform can be used to deepen understanding of the impact of matrix softening on cancer biology, an important but rarely studied phenomenon.