This study compares the biofilm efficacy and tissue compatibility of three cold atmospheric plasma devices: J-Plasma (Apyx Medical), Piezo Brush PZ3 (Reylon Plasma GmbH), and Plasma Care (Terraplasma-Medical GmbH). We focused on surface temperature, pH shifts, and ROS/RNS production, analyzing their effects on bacterial biofilms and tissue both ex vivo and in vivo. We measured temperature changes, pH shifts, optical emission spectra, and ROS/RNS levels in the liquid phase. Antimicrobial efficacy was tested against biofilms of and , grown on 3D-printed poly(ε-caprolactone) discs and titanium sheets. Tissue tolerability was assessed on porcine skin using direct counting for bactericidal effectiveness and SEM for validation, with histological analysis for tissue effects. Plasma Care demonstrated significant bactericidal capabilities (4 logs against in 60 seconds on PCL), minimal tissue disruption, and required short treatment times, functioning independently of target conductivity. Both J-Plasma and Piezo Brush PZ3 showed variable outcomes influenced by treatment duration and target surface conductivity, affecting both bactericidal activity and tissue tolerance. The comparative analysis underscores the importance of selecting plasma devices based on specific clinical needs to enhance biofilm management and minimize tissue damage, supporting tailored applications in precision medicine.

Immune cells experience a wide range of modes and magnitudes of mechanical forces as they infiltrate tissues and physically interact with other cells. Biophysical forces influence cell phenotypes through mechanosensing of the cytoskeleton, cell adhesion, catch and slip bonds, and mechanically gated ion channels. As a result, different mechanical environments impact the function and expression of immune cell receptors, which subsequently affects local and systemic immune responses. Mechanical coupling of immune cell receptors can be exploited in immuno-engineering applications such as adoptive cell transfer and artificial antigen-presenting cells through biomaterial systems with tunable mechanical properties that regulate receptor expression and cell activation. This review covers immune cell receptors in the adaptive and innate immune system that respond to mechanical forces and their potential to be applied for advancing current immunotherapies.

Globally we are faced with a significant burden of tuberculosis (TB), which is difficult to eradicate due to patients' non-adherance, and drug-resistant strains that are spreading at an alarming rate. Novel approaches are required to improve diagnosis and treatment. Metallic nanoparticles (MNPs) have demonstrated potential as sensor probes and in combination therapy, which combines MNPs with antimycobacterial drugs to develop new treatment and theranostic approaches. To strengthen the theoretical foundation towards the clinical application of TB nanomedicine, this review focuses on the properties and effectiveness of therapeutically relevant MNPs. We also elaborate on their antimycobacterial mechanisms. This review aims to analyze the body of literature on the topic, pinpoint important empirical findings, and identify knowledge gaps that could provide a basis for future research endeavors and translation of the technologies. Current data suggest that MNPs are potential systems for efficient diagnosis and treatment although additional pre-clinical and clinical research is needed to bring these technologies to the clinic.

Despite multiple treatments for tuberculosis (TB), there are ≈10 million new cases and 1.5 million deaths annually, warranting the need for new therapeutics. Major clinical treatment issues include the length of treatment which is associated with patient non-compliance; and poor cellular drug penetration leading to the generation of drug-resistant strains. This study underscores the potential of glucan-chitosan (CS) poly(lactic co-glycolic) acid (PLGA) nanoparticles as a promising immunostimulatory adjunct for TB treatment. To facilitate drug delivery to alveolar macrophage, a CS-PLGA nanoparticle is developed containing rifampin in the core with glucan as a surface ligand, to stimulate the immune system. Mice are administered a single dose of nanoparticles or free rifampin by oropharyngeal aspiration. Pharmacokinetic investigations reveal sustained release properties of rifampin in vivo, extending over a week. Furthermore, comprehensive analysis indicates stimulation of the innate immune system, as evidenced by cytokine profiling, while concurrently revealing no detrimental effects on the alveolar epithelium, as indicated by histological examination and albumin lung leak assessment. These findings collectively establish a strong foundation for the development of a novel adjuvant immunotherapy approach for TB.

Impaired bladder compliance secondary to congenital or acquired bladder dysfunction can lead to irreversible kidney damage. This is managed with surgical augmentation utilizing intestinal tissue, which can cause stone formation, infections, and malignant transformation. Co-seeded bone marrow mesenchymal stem cell (MSC)/CD34+ hematopoietic stem cell (HSPC) scaffolds (PRS) have been successful in regenerating bladder tissue. However, the acquisition of viable cells is challenging in the clinical setting. Here, the regenerative capacity of human MSC/CD34+ co-cultured total condition media (TCM) is compared to media alone in immune-competent rats augmented with PRS following partial cystectomy. Augmented bladders are instilled with media (control, = 4) or TCM ( = 5) twice a week for 4 weeks. Regenerated tissue is analyzed for smooth muscle, urothelium, vascular, and peripheral nerve regrowth. Urodynamic (UDS) measures are performed pre- and 4 weeks post-augmentation. The results demonstrate that TCM-instilled grafts have greater muscle content, larger average urothelial widths, higher percent vascularization, and more robust neural infiltration post-augmentation. UDS demonstrates greater percent bladder recovery in the TCM group, indicating functional improvement in bladder storage capacity. This study is the first to propose the use of cell-free TCM as an alternative to traditional cell-seeded scaffolds to promote bladder tissue regeneration.

Tissue regeneration involves dynamic dialogue between and among different cells and their surrounding matrices. Bone regeneration is specifically governed by reciprocity between osteoblasts and osteoclasts within the bone microenvironment. Osteoclast-directed resorption and osteoblast-directed formation of bone are essential to bone remodeling, and the crosstalk between these cells is vital to curating a sequence of events that culminate in the creation of bone tissue. Among bone biomaterial strategies, many have investigated the use of different material cues to direct the development and activity of osteoblasts. However, less attention has been given to exploring features that similarly target osteoclast formation and activity, with even fewer strategies demonstrating or integrating biomaterial-directed modulation of osteoblast-osteoclast coupling. This review aims to describe various biomaterial cues demonstrated to influence osteoclastogenesis and osteoclast function, emphasizing those that enhance a material construct's ability to achieve bone healing and regeneration. Additionally discussed are approaches that influence the communication between osteoclasts and osteoblasts, particularly in a manner that takes advantage of their coupling. Deepening our understanding of how biomaterial cues may dictate osteoclast differentiation, function, and influence on the microenvironment may enable the realization of bone-replacement interventions with enhanced integrative and regenerative capacities.

Nucleic acid vaccines play important roles in prevention and treatment of diseases. However, limited immunogenicity remains a major obstacle for DNA vaccine applications in the clinic. To address the issue, the present study investigates a cocktail approach to DNA vaccination. In this proof-of-the-concept study, the cocktail consists of two DNAs encoding viral hemagglutinin (HA) and granulocyte-macrophage colony stimulatory factor (GM-CSF), respectively. Data from the study demonstrate that recruitment and activation of antigen-presenting cells (APCs) can be substantially improved by spatiotemporal regulation of GM-CSF and HA expressions at the site of vaccination. The types of recruited APCs and their phenotypes are also controllable by adjusting the cocktail compositions. Compared to mono-ingredient vaccine, the optimized cocktail vaccine is able to enhance the anti-viral humoral and T cell immune responses. No significant systemic inflammation has been detected after either prime or boost immunization using the cocktail vaccine. Data in the study suggest that the DNA cocktail is a safe, effective, and controllable platform for improving vaccine efficacy.

Despite decades of intense research, glioma remains a disease for which no adequate clinical treatment exists. Given the ongoing therapeutic failures of conventional treatment approaches, nanomedicine may offer alternative options because it can increase the bioavailability of drugs and alter their pharmacokinetics. Here, a new type of synthetic protein nanoparticles (SPNPs) is reported that allow for effective loading and controlled release of the potent cancer drug, paclitaxel (PTX) - a drug that so far has been unsuccessful in glioma treatment due to hydrophobicity, low solubility, and associated delivery challenges. SPNPs are prepared by electrohydrodynamic (EHD) jetting of dilute solutions of PTX-loaded albumin made by high-pressure homogenization. After EHD jetting, PTX SPNPs possess a dry diameter of 165 ± 44 nm, hydrated diameter of 297 ± 102 nm, and a zeta potential of -19 ± 8 mV in water. For the SPNP formulation with a total PTX loading of 9.4%, the loading efficiency is 94%, and controlled release of PTX is observed over two weeks (6% burst release). PTX SPNPs are more potent (68% lethality) than free PTX (45% lethality using 0.2% dimethyl sulfoxide). PTX SPNPs in combination with IR show a significant survival benefit in glioma-bearing mouse models, avoid adverse liver toxicity, and maintain a normal brain architecture. Immunohistochemistry reveals a dramatic tumor size reduction including 40% long-term survivors without discernible signs of tumor. Using flexibly engineered SPNPs, this work outlines an efficient strategy for the delivery of hydrophobic drugs that are otherwise notoriously hard to deliver.

Despite extensive advances in cancer research, glioblastoma (GBM) still remains a very locally invasive and thus challenging tumor to treat, with a poor median survival. Tumor cells remodel their microenvironment and utilize extracellular matrix to promote invasion and therapeutic resistance. We aim here to determine how GBM cells exploit hyaluronan (HA) to maintain proliferation using ligand-receptor dependent and ligand-receptor independent signaling. We use tissue engineering approaches to recreate the three-dimensional tumor microenvironment in vitro, then analyze shifts in metabolism, hyaluronan secretion, HA molecular weight distribution, as well as hyaluronan synthetic enzymes (HAS) and hyaluronidases (HYAL) activity in an array of patient derived xenograft GBM cells. We reveal that endogenous HA plays a role in mitochondrial respiration and cell proliferation in a tumor subtype dependent manner. We propose a tumor specific combination treatment of HYAL and HAS inhibitors to disrupt the HA stabilizing role in GBM cells. Taken together, these data shed light on the dual metabolic and ligand - dependent signaling roles of hyaluronan in glioblastoma.

Glucagon-like peptide-1 receptor agonists (GLP-1 RAs) are one of the most effective treatments for obesity. The current therapy associated with repeated subcutaneous injections to maintain the drug therapeutic effect causes patient compliance issues and raises environmental concerns (due to sharp biohazard waste from disposed syringes/needles). Herein, we report a programmable scheduled release microneedles (PSR-MNs) system for delivering Semaglutide (a GLP-1 RA agent with a half-life of ~ 7 days) to manage and treat obesity. A single skin administration of a PSR-MNs patch (2 cm × 2 cm) which contains 4 programmable core-shell MNs patches (1 cm each, so-called pixels) enables the repeated release of Semaglutide every 7 days and sustains the drug efficacy for an unprecedented one-month period, simulating the effect of using four bolus injections spaced 7 days apart. Our PSR-MNs system provides an advanced injection-free platform to significantly enhance the current treatment of obesity with GLP-1RAs, addressing concerns related to pain, needle phobia, high cost and the need of medical facilities/personnel in traditional injections to administer the drug.

Most treatments to alleviate major depression work by either inhibiting human monoamine transporters, vital for the reuptake of monoamine neurotransmitters, or by inhibiting monoamine oxidases, which are vital for their degradation. The analysis of the experimental 3D structures of those antidepressants in their drug formulation state is key to precision drug design and development. In this study, we apply microcrystal electron diffraction (MicroED) to reveal the atomic 3D structures for the first time of five of the most prevalent antidepressants (reboxetine, pipofezine, ansofaxine, phenelzine, bifemelane) directly from the commercially available powder of the active ingredients. Their modes of binding are investigated by molecular docking, revealing the essential contacts and conformational changes into the biologically active state. This study underscores the combined use of MicroED and molecular docking to uncover elusive drug structures and mechanisms to aid in further drug development pipelines.

Liposomal J-Aggregates of Indocyanine Green (L-JA) can serve as a biocompatible and biodegradable nanoparticle for photoacoustic imaging and photothermal therapy. When compared to monomeric IcG, L-JA are characterized by longer circulation, improved photostability, elevated absorption at longer wavelengths, and increased photoacoustic signal generation. However, the documented methods for production of L-JA vary widely. We developed an approach to efficiently form IcG J-aggregates (IcG-JA) directly in liposomes at elevated temperatures. Aggregating within fully formed liposomes ensures particle uniformity and allows for control of J-aggregate size. L-JA have unique properties compared to IcG. L-JA provide significant contrast enhancement in photoacoustic images for up to 24 hours after injection, while IcG and unencapsulated IcG-JA are cleared within an hour. L-JA allow for more accurate photoacoustic-based sO estimation and particle tracking compared to IcG. Furthermore, photothermal heating of L-JA with an 852nm laser is demonstrated to be more effective at lower laser powers than conventional 808nm lasers for the first time. The presented technique offers an avenue for formulating a multi-faceted contrast agent for photoacoustic imaging and photothermal therapy that offers significant advantages over other conventional agents.

The paucity of targeted therapies for triple-negative breast cancer (TNBC) causes patients with this aggressive disease to suffer a poor clinical prognosis. A promising target for therapeutic intervention is the Wnt signaling pathway, which is activated in TNBC cells when extracellular Wnt ligands bind overexpressed Frizzled7 (FZD7) transmembrane receptors. This stabilizes intracellular β-catenin proteins that in turn promote transcription of oncogenes that drive tumor growth and metastasis. To suppress Wnt signaling in TNBC cells, we developed therapeutic nanoparticles (NPs) functionalized with FZD7 antibodies and β-catenin small interfering RNAs (siRNAs). The antibodies enable TNBC cell-specific binding and inhibit Wnt signaling by locking FZD7 receptors in a ligand unresponsive state, while the siRNAs suppress β-catenin through RNA interference. Compared to NPs coated with antibodies or siRNAs individually, NPs coated with both agents more potently reduce the expression of several Wnt related genes in TNBC cells, leading to greater inhibition of cell proliferation, migration, and spheroid formation. In two murine models of metastatic TNBC, the dual antibody/siRNA nanocarriers outperformed controls in terms of inhibiting tumor growth, metastasis, and recurrence. These findings demonstrate suppressing Wnt signaling at both the receptor and mRNA levels via antibody/siRNA nanocarriers is a promising approach to combat TNBC.

The gold standard of auricular reconstruction involves manual graft assembly from autologous costal cartilage. The intervention may require multiple surgical procedures and lead to donor-site morbidity, while the outcome is highly dependent on individual surgical skills. A tissue engineering approach provides the means to produce cartilage grafts of a defined shape from autologous chondrocytes. The use of autologous cells minimizes the risk of host immune response; however, factors such as biomaterial compatibility and in vitro maturation of the tissue-engineered (TE) cartilage may influence the engraftment and shape-stability of TE implants. Here, this work tests the biocompatibility of bioprinted autologous cartilage constructs in a rabbit model. The TE cartilage is produced by embedding autologous auricular chondrocytes into hyaluronan transglutaminase (HATG) based bioink, previously shown to support chondrogenesis in human auricular chondrocytes in vitro and in immunocompromised xenotransplantation models in vivo. A drastic softening and loss of cartilage markers, such as sulfated glycosaminoglycans (GAGs) and collagen type II are observed. Furthermore, fibrous encapsulation and partial degradation of the transplanted constructs are indicative of a strong host immune response to the autologous TE cartilage. The current study thus illustrates the crucial importance of immunocompetent autologous animal models for the evaluation of TE cartilage function and compatibility.

Antimicrobial resistance remains a critical global health concern, necessitating the investigation of alternative therapeutic approaches. With the diminished efficacy of conventional small molecule drugs due to the emergence of highly resilient bacterial strains, there is growing interest in the potential for alternative therapeutic modalities. As naturally occurring viruses of bacteria, bacteriophage (or phage) are being re-envisioned as a platform to engineer properties that can be tailored to target specific bacterial strains and employ diverse antibacterial mechanisms. However, limited understanding of key pharmacological properties of phage is a major challenge to translating its use from preclinical to clinical settings. Here, we review modern advancements in phage-based antimicrobial therapy and discuss the in vivo pharmacokinetics and biodistribution of phage, addressing critical challenges in their application that must be overcome for successful clinical implementation.

Multiple therapies have been studied to ameliorate the neuroinhibitory cues present after traumatic injury to the central nervous system. Two previous in vitro studies have demonstrated the efficacy of the FDA-approved cardiovascular therapeutic, protamine (PRM), to overcome neuroinhibitory cues presented by chondroitin sulfates; however, the effect of a wide range of PRM concentrations on neuronal and glial cells has not been evaluated. In this study, we investigate the therapeutic efficacy of PRM with primary cortical neurons, hippocampal neurons, mixed glial cultures, and astrocyte cultures. We show the threshold for PRM toxicity to be at or above 10 μg/ml depending on the cell population, that 10 μg/ml PRM enables neurons to overcome the inhibitory cues presented by chondroitin sulfate type A, and that soluble PRM allows neurons to more effectively overcome inhibition compared to a PRM coating. We also assessed changes in gene expression of reactive astrocytes with soluble PRM and determined that PRM does not increase their neurotoxic phenotype and that PRM may reduce brevican production and serpin transcription in cortical and spinal cord astrocytes. This is the first study to thoroughly assess the toxicity threshold of PRM with neural cells and study astrocyte response after acute exposure to PRM in vitro.

Diabetes Mellitus Type 2 (T2D) is an emerging health burden in the USand worldwide, impacting approximately 15% of Americans. Current front-line therapeutics for T2D patients include sulfonylureas that act to reduce A1C and/or fasting blood glucose levels, or Metformin that antagonizes the action of glucagon to reduce hepatic glucose production. Next generation glucomodulatory therapeutics target members of the high-affinity glucose transporter Sodium-Glucose-Linked-Transporter (SGLT) family. SGLT1 is primarily expressed in intestinal epithelium, whose inhibition reduces dietary glucose uptake, whilst SGLT2 is highly expressed in kidney - regulating glucose reabsorption. A number of SGLT2 inhibitors are FDA approved whilst SGLT1 and dual SGLT1 & 2 inhibitor are currently in clinical trials. Here, we discuss and compare SGLT2, SGLT1, and dual inhibitors' biochemical mechanism and physiological effects.

The immune system plays a crucial role in recognizing and eliminating pathogenic substances and malignant cells in the body. For cancer treatment, immunotherapy is becoming the standard treatment for many types of cancer and is often combined with chemotherapy. Although chemotherapeutic agents are often reported to have adverse effects, including immunosuppression, they can also play a positive role in immunotherapy by directly stimulating the immune system. This has been demonstrated in preclinical and clinical studies in the past decades. Chemotherapeutics can activate immune cells through different immune receptors and signaling pathways depending on their chemical structure and formulation. In this review, we summarize and discuss the direct immunoactivation effects of chemotherapeutics and possible mechanisms behind these effects. Finally, we prospect chemo-immunotherapeutic combinations for the more effective and safer treatment of cancer.

Following a traumatic brain injury (TBI), excess reactive oxygen species (ROS) and lipid peroxidation products (LPOx) are generated and lead to secondary injury beyond the primary insult. A major limitation of current treatments is poor target engagement, which has prevented success in clinical trials. Thus, nanoparticle-based treatments have received recent attention because of their ability to increase accumulation and retention in damaged brain. Theranostic neuroprotective copolymers (NPC3) containing thiol functional groups can neutralize ROS and LPOx. Immediate administration of NPC3 following injury in a controlled cortical impact (CCI) mouse model provides a therapeutic window in reducing ROS levels at 2.08-20.83 mg/kg in males and 5.52-27.62 mg/kg in females. This NPC3-mediated reduction in oxidative stress improves spatial learning and memory in males, while females show minimal improvement. Notably, NPC3-mediated reduction in oxidative stress prevents the bilateral spread of necrosis in male mice, which was not observed in female mice and likely accounts for the sex-based spatial learning and memory differences. Overall, these findings suggest sex-based differences to oxidative stress scavenger nanoparticle treatments, and a possible upper threshold of antioxidant activity that provides therapeutic benefit in injured brain since female mice benefit from NPC3 treatment to a lesser extent than male mice.

Cancer immunotherapy targeting adaptive immune cells has been attracting considerable interest due to its great success in treating multiple cancers. Recently, there is also increasing interest in agents that can stimulate innate immune cell activities. Immune checkpoint inhibitors targeting innate immune cells can block inhibitory interactions ('don't eat me' signals) between tumor cells and phagocytes. CD47 is a transmembrane protein overexpressed in various cancers and acts as a potent 'do not eat me' signal that contributes to the immune evasion of cancer cells. Anti-CD47 peptides that can bind to CD47 and block CD47/SIRPα interaction were discovered using a novel phage display biopanning strategy. Anti-CD47 peptides enhanced the macrophage-mediated phagocytosis of NCI-H82 tumor cells . Unlike anti-CD47 antibodies, these peptides do not induce the agglutination of RBCs. Moreover, anti-CD47 peptides exhibit high specificity for MC-38 cancer cells expressing CD47. CMP-22 peptide showed the ability to increase the antitumor activity of doxorubicin and extends the survival of CT26 tumor-bearing mice. The discovered anti-CD47 peptides can be considered potential candidates for cancer immunotherapy by blocking the CD47/SIRPα interaction, especially in combination with chemotherapy, to elicit synergistic effects.

Protein-based drug delivery systems have gained popularity due to their biocompatibility, straightforward surface modification, and potential for intrinsic therapeutic activity. Among therapeutic proteins, enzymes are particularly attractive because of their specificity, efficient reaction rates, regeneration after substrate turnover, and proven track record in the treatment of diseases ranging from cancer to inherited metabolic and lysosomal storage disorders. Herein, previous work on electrohydrodynamic jetting is expanded upon by developing a novel class of protein nanoparticles that features therapeutic enzymes. In particular, nanoparticles incorporating the antioxidant enzyme, catalase, at weight fractions as high as 50% are reported. Catalase-based synthetic protein nanoparticles (CAT-SPNPs) demonstrate sustained antioxidative activity, retain significantly enhanced enzymatic activity compared to its solute form, and overall demonstrate good structural stability. Moreover, surface functionalization of CAT-SPNPs with targeting antibodies results in ≈12.5-fold increase in uptake over unmodified control particles. Importantly, CAT-SPNPs exert protection from oxidative stress, as indicated by significant increase in viability and reduction in LDH release compared to equivalent amounts of free catalase. Taken together, the work establishes targeted enzyme-based SPNPs as a platform for enhancing the drug-like properties of therapeutic enzymes.