Bacteria-driven nanosonosensitizer delivery system for enhanced breast cancer treatment through sonodynamic therapy-induced immunogenic cell death.

BACKGROUND: Sonodynamic therapy (SDT) has shown promise as a non-invasive cancer treatment due to its local effects and excellent tissue penetration. However, the limited accumulation of sonosensitizers at the tumor site hinders its therapeutic efficacy. Although nanosonosensitizers have improved local tumor accumulation through passive targeting via the enhanced permeability and retention effect (EPR), achieving sufficient accumulation and penetration into tumors remains challenging due to tumor heterogeneity and inaccurate targeting. Bacteria have become a promising biological carrier due to their unique characteristic of active targeting and deeper penetration into the tumor. METHODS: In this study, we developed nanosonosensitizers consisting of sonosensitizer, hematoporphyrin monomethyl ether (HMME), and perfluoro-n-pentane (PFP) loaded poly (lactic-co-glycolic) acid (PLGA) nanodroplets (HPNDs). These HPNDs were covalently conjugated onto the surface of Escherichia coli Nissle 1917 (EcN) using carbodiimine chemistry. EcN acted as an active targeting micromotor for efficient transportation of the nanosonosensitizers to the tumor site in triple-negative breast cancer (TNBC) treatment. Under ultrasound cavitation, the HPNDs were disrupted, releasing HMME and facilitating its uptakes by cancer cells. This process induced reactive oxygen species (ROS)-mediated cell apoptosis and immunogenic cell death (ICD) in vitro and in vivo. RESULTS: Our bacteria-driven nanosonosensitizer delivery system (HPNDs@EcN) achieved superior tumor localization of HMME in vivo compared to the group treated with only nanosonosensitizers. This enhanced local accumulation further improved the therapeutic effect of SDT induced-ICD therapeutic effect and inhibited tumor metastasis under ultrasound stimulation. CONCLUSIONS: Our research demonstrates the potential of this ultrasound-responsive bacteria-driven nanosonosensitizer delivery system for SDT in TNBC. The combination of targeted delivery using bacteria and nanosonosensitizer-based therapy holds promise for achieving improved treatment outcomes by enhancing local tumor accumulation and stimulating ICD.

Chemically heparinized PEEK via a green method to immobilize bone morphogenetic protein-2 (BMP-2) for enhanced osteogenic activity.

Osseointegration remains one of the major challenges in the success of bone-related implants. Recently, polyetheretherketone (PEEK) has emerged as an alternative material in orthopedic and dental applications due to its bone-mimicking mechanical properties. However, its bioinertness resulting in poor osseointegration has limited its potential application. So, the surface modification of PEEK with bone morphogenetic protein-2 (BMP-2) can be a potential approach for improving osseointegration. In this study, we proposed the chemical modification of heparin onto PEEK through an environmentally benign method to exploit the BMP-2 binding affinity of heparin. The heparin was successfully functionalized on the PEEK surface via a combination of ozone and UV treatment without using organic solvents or chemicals. Furthermore, BMP-2 was efficiently immobilized on PEEK and exhibited a sustained release of BMP-2 compared to the pristine PEEK with enhancement of bioactivity in terms of proliferation as well as osteogenic differentiation of MG-63. The significant synergistic effect of BMP-2 and heparin grafting on osteogenic differentiation of MG-63 was observed. Overall, we demonstrated a relatively safe method where no harsh chemical reagent or organic solvent was involved in the process of heparin grafting onto PEEK. The BMP-2 loaded, heparin-grafted PEEK could serve as a potential platform for osseointegration improvement of PEEK-based bone implants.

Advancing burn wound treatment: exploring hydrogel as a transdermal drug delivery system.

Burn injuries are prevalent and life-threatening forms that contribute significantly to mortality rates due to associated wound infections. The management of burn wounds presents substantial challenges. Hydrogel exhibits tremendous potential as an ideal alternative to traditional wound dressings such as gauze. This is primarily attributed to its three-dimensional (3D) crosslinked polymer network, which possesses a high water content, fostering a moist environment that supports effective burn wound healing. Additionally, hydrogel facilitates the penetration of loaded therapeutic agents throughout the wound surface, combating burn wound pathogens through the hydration effect and thereby enhancing the healing process. However, the presence of eschar formation on burn wounds obstructs the passive diffusion of therapeutics, impairing the efficacy of hydrogel as a wound dressing, particularly in cases of severe burns involving deeper tissue damage. This review focuses on exploring the potential of hydrogel as a carrier for transdermal drug delivery in burn wound treatment. Furthermore, strategies aimed at enhancing the transdermal delivery of therapeutic agents from hydrogel to optimize burn wound healing are also discussed.

Enhancing photothermal therapy of tumors with image-guided thermal control of gene-expressing bacteria.

Purpose: Bacteria-mediated tumor therapy has showed promising potential for cancer therapy. However, the efficacy of bacterial monotherapy treatment which can express and release therapeutic proteins in tumors has been found to be unsatisfactory. To date, synergistic therapy has emerged as a promising approach to achieve stronger therapeutic outcomes compared to bacterial monotherapy. It is a challenge to visualize these tumor-homing bacteria in vivo and guide them to express and release in situ therapeutic proteins. Procedure: We have developed a kind of engineered bacteria (named CGB@ICG) genetically incorporating acoustic reporter proteins and thermo-inducible ClyA expression gene circuit and chemically modified with indocyanine green on the bacterial surface. The presence of acoustic reporter proteins and indocyanine green facilitates the visualization of CGB@ICG via contrast-enhanced ultrasound imaging and optical imaging, making it possible to guide the sound wave or laser to irradiate precisely these bacteria for inducing the expression of ClyA protein via acoustic- or photothermal effects. The expression and secretion of ClyA protein in the tumor, combined with photothermal therapy, greatly enhanced the anti-tumor efficacy of the engineered bacteria and improved their biosafety. Results: We successfully performed multimodal imaging of CGB@ICG in vivo resulting in remoting control the expression of ClyA protein in tumor. In vivo experiments showed that bacteria-mediated therapy combined photothermal therapy exhibited a rapid decrease in tumor volume compared to other groups, while the tumor volume of the combination therapy group continued to decrease and even achieved complete healing. Thus, combination therapy not only reduced the rate of tumor growth but also prevented the proliferation of tumor cells for an extended period. Conclusion: Our study demonstrated that CGB@ICG serves as an efficacious imaging agent and delivery vector to combine engineered bacteria with photothermal therapy, holding great promise for tumor treatment.

Synthesis of Superabsorbent Hydrogels with Predefined Geometries and Controlled Swelling Properties for Versatile 3D Cell Culture Scaffolds.

This research presents a simple but general method to prepare water-soluble-polymer-based superabsorbent hydrogels with predefined microscale geometries and controlled swelling properties. Unlike conventional hydrogel preparation methods based on bulk solution-phase cross-linking, poly(vinyl alcohol) is homogeneously mixed with polymer-based cross-linkers in the solution phase and thermally cross-linked in the solid phase after drying; the degree of cross-linking is modulated by controlling the cross-linker concentration, pH, and/or thermal annealing conditions. After the shape definition process, cross-linked films or electrospun nanofibers are treated with sulfuric acid to weaken hydrogen bonds and introduce sulfate functionality in polymer crystallites. The resultant superabsorbent hydrogels exhibit an isotropic expansion of the predefined geometry and tunable swelling properties. Particularly, hydrogel microfibers exhibit excellent optical transparency, good biocompatibility, large porosity, and controlled cell adhesion, leading to versatile 3D cell culture scaffolds that not only support immortalized cell lines and primary neurons but also enable stiffness-modulated cell adhesion studies.

Mesenchymal stem cell-encapsulated cellulose nanofiber microbeads and enhanced biological activities by hyaluronic acid incorporation.

Cell microencapsulation is a process to entrap viable and functional cells within a biocompatible and semi-permeable matrix to provide a favorable microenvironment to the cells. Cellulose nanofiber (CNF), a low-cost and sustainable cellulose-derived natural polymer, has been studied as a matrix for 3D stem cell culture in the form of a bulk hydrogel. Here, the preparation of CNF microbeads for the long-term 3D culture of human adipose-derived stem cells (hADSCs) was demonstrated. Furthermore, hyaluronic acid (HA) was physically incorporated into the stem cell encapsulated CNF microbeads with various molecular weights and concentrations to investigate its potential in enhancing the cellular bioactivities. The beneficial effects of HA incorporation on encapsulated cells were significant compared to CNF microbeads, especially with 700 kDa molecular weight and 0.2% in concentration in terms of cell proliferation (~2 times) and VEGF secretion (~2 times) while maintaining their stemness. All the results demonstrated that the HA-incorporated CNF microbeads could serve as a promising microencapsulation matrix for hADSCs.

Injectable and detachable heparin-based hydrogel micropatches for hepatic differentiation of hADSCs and their liver targeted delivery.

A micropatterned heparin-based hydrogel system that can provide sustained release of multiple growth factors upon one time loading was prepared via photopolymerization and lithography and it was employed as a culture matrix for differentiating hADSCs into hepatic lineage. Mature differentiation of hADSCs into hepatic lineage in terms of gene expression and immunofluorostaining of hepatic markers, and functional characteristics such as glycogen storage ability and production of albumin and urea was observed on the soft hydrogel (∼400 Pa) when the gel elasticity was modulated. This optimal heparin-based hydrogel was used to prepare micropatches containing hepatic-differentiated cells by 1) micropatterning of the gel on a polyelectrolyte multilayer (PEM), 2) seeding of hADSCs and inducing hepatic differentiation, and 3) electrochemical retrieval of cell-attached micropatches. Upon i.v. injection, the retrieved cell micropatches showed a prolonged retention in the liver and promoted function compared to single cell injection in a rat model. In conclusion, this injectable and detachable miropatterned heparin-based hydrogel system could serve as a total platform for the stem cell differentiation under well-controlled microenvironment in vitro and for targeted delivery of the differentiated cells in vivo.

Rapid and Versatile Cell Aggregate Formation Using Lipid-Conjugated Heparin.

Cell aggregates hold significant therapeutic promise for in vitro cell analysis, ex vivo tissue models, and in vivo cell therapy and tissue engineering. Traditional methods of making cell aggregates require long incubation times and can only produce three-dimensional-spheroid-shaped aggregates. We propose a novel method of making cell aggregates of diverse sizes and shapes using lipid-conjugated heparin. Shaking the cell suspension containing a small amount of lipid-conjugated heparin for approximately 30 min produced cell aggregates. This approach can be applied to any cell type, including stem cells, fibroblast cells, and T lymphocytes. The shape of biocompatible templates could modulate the shape of cell aggregates. In addition to layered, multicompartmental cell aggregates on template, template-free, tube-shaped cell aggregates could also be made. The cell aggregates formed were alive and maintained biological activities.

In situ formation of injectable and porous heparin-based hydrogel.

In this study, by combining photopolymerization and particle leaching technique, in situ formation of porous hydrogel with pore interconnectivity was demonstrated in vivo upon subcutaneous injection into the back of mice as well as in vitro. A precursor solution containing thiolated heparin, PEG-diacrylate (PEG-DA), and gelatin microparticles (GMPs) as a fast dissolving porogen were photopolymerized by visible-light-initiated thiol-ene reaction with eosin Y (EY) as a photo initiator and triethanolamine (TEOA) as a co-initiator. Formation of porous structure of the hydrogel after subsequent leaching of GMPs was confirmed in an animal model as well as in a physiological environment. The physical characteristics of the hydrogel were analyzed, and the acute in vivo biocompatibility of this system was characterized.

Epidermal growth factor loaded heparin-based hydrogel sheet for skin wound healing.

A heparin-based hydrogel sheet composed of thiolated heparin and diacrylated poly (ethylene glycol) was prepared via photo polymerization and human epidermal growth factor (hEGF) were loaded into it for the purpose of wound healing. It showed a sustained release profile of hEGF in vitro. In order to evaluate its function on wound healing in vivo, full thickness wounds were created on the dorsal surface of mice. Application of hEGF loaded heparin-based hydrogel sheet accelerated the wound closure compared to the non-treated control group, hEGF solution, and hEGF loaded PEG hydrogel sheet. Histological and immunohistological examinations also demonstrated an advanced granulation tissue formation, capillary formation, and epithelialization in wounds treated by hEGF loaded heparin-based hydrogel compared to other groups, and no biocompatibility issue was observed. In conclusion, the delivery of hEGF using the heparin-based hydrogel could accelerate the skin wound healing process.