Protein Release Profiles Research Articles

PEARL: Protein Eluting Alginate with Recombinant Lactobacilli.

Engineered living materials (ELMs) made of bacteria in hydrogels have shown considerable promise for therapeutic applications through controlled and sustained release of complex biopharmaceuticals at low costs and with reduced wastage. While most therapeutic ELMs use E. coli due to its large genetic toolbox, most live biotherapeutic bacteria in development are lactic acid bacteria due to native health benefits they offer. Among these, lactobacilli form the largest family of probiotics with therapeutic potential in almost all sites of the body with a microbiome. A major factor limiting the use of lactobacilli in ELMs is their limited genetic toolbox. This study expands on recent work to expand the genetic programmability of probiotic Lactiplantibacillus plantarum WCFS1 for protein secretion and encapsulate it in a simple, cost-effective, and biocompatible core-shell alginate bead to develop an ELM. The controlled release of recombinant proteins is demonstrated, even up to 14 days from this ELM, thereby terming it PEARL - Protein Eluting Alginate with Recombinant Lactobacilli. Notably, lactobacillus encapsulation offered benefits like bacterial containment, protein release profile stabilization, and metabolite-induced cytotoxicity prevention. These findings demonstrate the mutual benefits of combining recombinant lactobacilli with alginate for the controlled and sustained release of proteins.

Scalable Manufacturing Method for Model Protein-Loaded PLGA Nanoparticles: Biocompatibility, Trafficking and Release Properties.

Background and Objectives: Drug delivery systems (DDSs) offer efficient treatment solutions to challenging diseases such as central nervous system (CNS) diseases by bypassing biological barriers such as the blood-brain barrier (BBB). Among DDSs, polymeric nanoparticles (NPs), particularly poly(lactic-co-glycolic acid) (PLGA) NPs, hold an outstanding position due to their biocompatible and biodegradable qualities. Despite their potential, the translation of PLGA NPs from laboratory-scale production to clinical applications remains a significant challenge. This study aims to address these limitations by developing scalable PLGA NPs and evaluating their potential biological applications. Methods: We prepared blank and model-protein-loaded (albumin-FITC and wheat germ agglutinin-488 (WGA-488)) fluorescent PLGA NPs using the traditional double-emulsion method combined with the micro-spray-reactor system, a novel approach that enables fine particle production enabling scale-up applications. We tested the biocompatibility of the NPs in living RPMI 2650 and neuroblastoma cell lines, as well as their trafficking and uptake. Release kinetics of the encapsulated proteins were investigated through confocal microscopy and in vitro release studies, providing insights into the stability and functionality of the released proteins. Results: The formulation demonstrated sustained and prolonged protein release profiles. Importantly, cellular uptake studies revealed that the NPs were not internalized. Furthermore, encapsulated WGA-488 protein retained its functional activity after release, validating the integrity of the encapsulation and release processes. Conclusions: The proof-of-concept study on NP manufacturing and an innovative drug trafficking and release approach can bring new perspectives on scalable preparations of PLGA NPs and their biological applications.

Medicated and multifunctional composite alginate-collagen-hyaluronate based scaffolds prepared using two different crosslinking approaches show potential for healing of chronic wounds.

Chronic wounds present significant challenges with high morbidity and mortality. A cost-effective dressing that can absorb large exudate volumes, is hemostatic and therapeutically active is of current interest. This study compares two crosslinking approaches on composite scaffolds comprising fish collagen (FCOL), hyaluronic acid (HA) and sodium alginate (SA) by respectively targeting HA and SA. Crosslinking involved reacting HA with polyethylene glycol diglycidyl ether (PEGDE)/itaconic acid (IT) (IPC scaffolds) or SA with calcium chloride (CC scaffolds) and the crosslinked gels (with/without BSA) freeze-dried. Selected optimized formulations were loaded with basic fibroblast growth factor (b-FGF) as medicated scaffold dressings. NMR and FTIR spectroscopies (crosslinking/component interactions), SEM (morphology), texture analysis (mechanical strength/adhesion), and exudate handling were used to characterize the physico-chemical properties of the scaffolds. Protein (BSA) release profiles, hemostasis, biocompatibility and wound closure were assessed using HPLC, whole blood and methyl thiazolyl tetrazolium (MTT) and scratch assays respectively. The CC SA:FCOL:HA scaffolds showed improved mechanical strength, porosity, water vapor transmission rate, retained structural integrity after absorbing 50% exudate and promoted cell proliferation. The IPC scaffolds showed enhanced structural integrity, excellent hemostasis, retained three times more exudate than non-crosslinked scaffolds and provided acceptable pore size for cell adhesion and proliferation. The results show potential of CC and IPC SA:FCOL:HA scaffolds as medicated dressings for delivering proteins to chronic wounds. The study's significance lies in their potential use as multifunctional, multi-targeted and therapeutic dressings to overcome challenges with chronic wounds and use as delivery platforms for other therapeutic agents for chronic wound healing.

Synthetic conduits efficacy in neural repair: a comparative study of dip-coated polycaprolactone and electrospun polycaprolactone/polyurethane conduits.

Objectives.Peripheral nerve injuries (PNI) represent the most common type of nervous system injuries, resulting in 5 million injuries per year. Current gold standard, autografts, still carry several limitations, including the inappropriate type, size, and function matches in grafted nerves, lack of autologous donor sites, neuroma formation, and secondary surgery incisions. Polymeric nerve conduits, also known as nerve guides, can help overcome the aforementioned issues that limit nerve recovery and regeneration by reducing tissue fibrosis, misdirection of regenerating axons, and the inability to maintain long- distance axonal growth. Polymer-based double-walled microspheres (DWMSs) are designed to locally and in a sustainable fashion deliver bioactive agents. Lysozyme is a natural antimicrobial protein that shares similar physical and chemical properties to glial cell line-derived neurotrophic factor, making it an ideal surrogate molecule to evaluate the release kinetics of encapsulated bioagent from polymeric biodegradable microspheres embedded in polycaprolactone and polycaprolactone/polyurethane blend nerve conduits.Approach.Lysozyme was encapsulated in poly(lactic-co-glycolic acid)/poly(L-lactide) DWMSs fabricated through a modified water-oil-water emulsion solvent evaporation method. Lysozyme-loaded DWMS were further embedded in PCL and PCL-PU based nerve guides constructed via polymer dip-coating and electrospinning method respectively. Lysozyme DWMS and nerve guides were imaged using scanning electron microscopy (SEM). Released lysozyme concentration was determined by using a colorimetric micro-BCA protein assay and spectrophotometric quantitation. Tensile and suture pull-out tests were utilized to evaluate the mechanical properties of both dip-coated and electrospun nerve guides, embedded and free of lysozyme DWMS.Main results.The study revealed significant distinctions in the lysozyme release profiles, and mechanical properties of the manufactured polymer nerve guides. Both PCL dip-coated and PCL/PU electrospun DWMS-embedded nerve guides revealed biphasic protein release profiles. PCL/PU electrospun and PCL dip-coated nerve guides released 16% and 29% of the total protein concentration within 72 h, plateauing at week 16 and week 8, respectively. SEM analysis of the nerve guides confirmed the homogeneity and integrity of the polymer nerve guides' structures. The electrospun guides were found to be more flexible with a higher extension under stress bending, while the dip-coated PCL nerve guides displayed more rigid behavior.Significance.This study provides useful insights on how to optimize nerve guide design and fabrication to enhance recovery progress of PNI.

Effect of Multiple-Cycle Collections of Conditioned Media from Different Cell Sources towards Fibroblasts in In Vitro Wound Healing Model.

Conditioned media refers to a collection of the used cell culture media. The goal of this study was to evaluate the possible impacts of different conditioned media collected across a number of cycles on the fibroblast proliferation, migration, and profiles of protein release. Human dermal fibroblast (HDF) cells and Wharton jelly mesenchymal stem cells (WJMSC) were cultured and incubated for 3 days prior to being harvested as cycle-1 using the serum-free media F12:DMEM and DMEM, respectively. The procedures were repeatedly carried out until the fifth cycle of conditioned media collection. An in-vitro scratch assay was conducted to measure the effectiveness of wound healing. Collagen hydrogel was combined separately with both the Wharton jelly-conditioned medium (WJCM) and the dermal fibroblast-conditioned medium (DFCM) in order to evaluate the protein release profile. The conditioned medium from many cycles had a lower level of fibroblast attachment than the control (complete medium); however, the growth rate increased from 100 to 250 h-1, when supplemented with a conditioned medium collected from multiple cycles. The wound scratch assay showed that fibroblast cell migration was significantly increased by repeating cycles up to cycle-5 of DFCM, reaching 98.73 ± 1.11%. This was faster than the rate of migration observed in the cycle-5 of the WJCM group, which was 27.45 ± 5.55%. Collagen hydrogel from multiple cycles of DFCM and WJCM had a similar protein release profile. These findings demonstrate the potential for employing repeated cycles of DFCM- and WJCM-released proteins with collagen hydrogel for applications in wound healing.

Engineering Metal-Phenolic Network Nanoparticles via Microfluidics.

Microfluidics opens new avenues for materials engineering, as it enables scalable synthesis and provides highly controllable environments for reactions. Herein, we leverage microfluidics to engineer the properties of (bioactive) metal-phenolic network nanoparticles (MPN NPs), an emerging and highly modular nanoparticle platform for the incorporation and delivery of bioactive cargo. By varying the microfluidics operating parameters (flow rate ratio, total flow rate, temperature) and NP composition, we assemble MPN NPs, which consist of poly(ethylene glycol), biomacromolecules, metal ions, and polyphenols. Compared to MPN NPs prepared via bulk assembly, the microfluidics-assembled MPN NPs possess a broader tunable size range (i.e., ∼40-330 nm vs ∼45-220 nm for bulk-assembled NPs) and a higher (by ∼30%) protein loading. The bulk-assembled MPN NPs show pH-responsive protein release behavior (e.g., ∼50% at pH 7; ∼25% at pH 9; 48 h). Likewise, the MPN NPs prepared via microfluidics at a flow rate ratio of 1:1 display similar pH-responsive protein release behavior. For the microfluidics-assembled MPN NPs, protein release is also dependent on temperature (e.g., 30% at 4 °C, and ∼50% at 20 and 37 °C). Furthermore, assembly at a 1:1 flow rate ratio overall enables greater tunability of protein release profiles than that at higher flow rate ratios. While bulk-assembled NPs display a higher degree of cell association, NPs assembled via both strategies can be internalized by cells after 24 h. These findings provide new insights into engineering the properties of metal-organic materials via microfluidics, which is expected to advance their development and application.

D55. A Tunable Nanoparticle-based Botulinum Toxin Formulation for Extended Duration of Therapeutic Effect

PURPOSE: Botulinum toxins (BoNT) are increasingly popular neuromodulators with an array of aesthetic and neurological indications. However, these agents are limited by their duration of effect, necessitating frequent redosing. We aim to develop a nanoparticle-based delivery system to extend BoNT release and characterize its release profile. METHODS: Botulinum toxin A (BoNTa) neuromodulators were encapsulated within polymeric nanoparticles (NPs) using a kinetically controlled microfluidic assembly method. Dynamic light scattering was used to measure particle size and uniformity. ELISA was used to quantify the protein release profile. RESULTS: Particle sizes were uniform with most particles centered at 100nm and nearly all particles between 50-200nm. A tunable release profile was achieved, in which BoNTa could be released linearly or with an initial bolus followed by sustained release at the desired rate. Reproducibility of NP performance was demonstrated in 3 independent trials. Our early in-vivo and mechanistic studies have demonstrated preservation of toxin bioactivity and extended therapeutic effect in a rat model. CONCLUSION: We developed a novel nanoparticle-based Botox formulation capable of providing long-acting release of neuromodulator with tunable kinetics at designated dosage. This sustained release delivery modality enables superior therapeutic outcomes with less frequent re-dosing.

Electrically responsive release of proteins from conducting polymer hydrogels.

Electrically modulated delivery of proteins provides an avenue to target local tissues specifically and tune the dose to the application. This approach prolongs and enhances activity at the target site whilst reducing off-target effects associated with systemic drug delivery. The work presented here explores an electrically active composite material comprising of a biocompatible hydrogel, gelatin methacryloyl (GelMA) and a conducting polymer, poly(3,4-ethylenedioxythiophene), generating a conducting polymer hydrogel. In this paper, the key characteristics of electroactivity, mechanical properties, and morphology are characterized using electrochemistry techniques, atomic force, and scanning electron microscopy. Cytocompatibility is established through exposure of human cells to the materials. By applying different electrical-stimuli, the short-term release profiles of a model protein can be controlled over 4h, demonstrating tunable delivery patterns. This is followed by extended-release studies over 21 days which reveal a bimodal delivery mechanism influenced by both GelMA degradation and electrical stimulation events. This data demonstrates an electroactive and cytocompatible material suitable for the delivery of protein payloads over 3 weeks. This material is well suited for use as a treatment delivery platform in tissue engineering applications where targeted and spatio-temporal controlled delivery of therapeutic proteins is required. STATEMENT OF SIGNIFICANCE: Growth factor use in tissue engineering typically requires sustained and tunable delivery to generate optimal outcomes. While conducting polymer hydrogels (CPH) have been explored for the electrically responsive release of small bioactives, we report on a CPH capable of releasing a protein payload in response to electrical stimulus. The composite material combines the benefits of soft hydrogels acting as a drug reservoir and redox-active properties from the conducting polymer enabling electrical responsiveness. The CPH is able to sustain protein delivery over 3 weeks, with electrical stimulus used to modulate release. The described material is well suited as a treatment delivery platform to deliver large quantities of proteins in applications where spatio-temporal delivery patterns are paramount.

Silica-collagen nanoformulations with extended human growth hormone release

Growth hormone deficiency has been treated by the daily administration of recombinant human growth hormone (hGH) for decades. Patient compliance to this treatment is generally incomplete due to challenges including dose frequency and lack of perceived benefits. This stimulates the research on new formulations to reduce the number of periodic administrations. In this study silica nanoparticles and silica-collagen nanocomposites were evaluated for hGH loading and release. Bare nanoparticles showed higher hGH adsorption capacity than thiol- and isobutyl-bearing particles of similar diameters. Monitoring of bound protein conformation changes indicated hGH structure retention when adsorbed on bare silica nanoparticles and suggested no alterations on protein activity. Protein-loaded particles incorporated into collagen matrices (silica-collagen nanocomposites) showed a progressive protein release profile different from the observed for hGH-loaded silica nanoparticles and hGH-loaded collagen matrices. While both the collagen and the silica nanoparticle systems reached a 100 % release after 4 and 7 days respectively, silica-collagen nanocomposites showed a bi-phasic prolonged hGH release reaching approximately an 80 % after 15 days. These findings suggest that biocompatible silica-collagen nanocomposites could be used as vehicles for the prolonged delivery of hGH which could lead to a potential reduction in the number of periodic administrations.

Size-dependent bioactivity of electrosprayed core–shell chitosan-alginate particles for protein delivery

Nano-bio interactions are size-dependent. The present study investigates whether core–shell chitosan-alginate particle size governs biological activities as well as protein release profile. A coaxial electrospraying was used to fabricate bovine serum albumin (BSA)-loaded core–shell micro/nanoparticles and were fully characterized. The bio/hemocompatibility of the particles was assessed using MTT and hemolytic assays, respectively, followed by the uptake assessment using flow cytometry. Finally, protein absorption was investigated using SDS-PAGE. The SEM size of the microparticles, the hydrodynamic, and the actual sizes of the nanoparticles were 1.2 μm, 90.49 nm, and 50 nm, respectively. Interactions among two polymers and BSA were observed using DSC analysis. BET analysis showed a more surface area for nanoparticles. A sustained release trend of BSA was observed after 14- and 10-day for microparticles and nanoparticles, respectively. Microparticles exhibited excellent hemocompatibility (< 5% hemolysis) and cell viability (at least > 70%) in all concentrations. However, acceptable hemolytic activity and cell viability were observed for nanoparticles in concentrations below 250 μg/mL. Furthermore, nanoparticles showed greater cellular uptake (~ 4 folds) and protein absorption (~ 1.61 folds) than microparticles. Overall, the developed core–shell chitosan-alginate particles in the micro/nanoscale can be promising candidates for biomedical application and regenerative medicine regarding their effects on above mentioned biological activities.

Development of Chitosan/Gelatin-Based Hydrogels Incorporated with Albumin Particles.

The research subject of this paper are natural polymer-based hydrogels modified with albumin particles. The proteins were obtained via the salt-induced precipitation method, and next characterized using dynamic light scattering (DLS), UV-Vis spectroscopy and FT-IR spectroscopy. The most favorable composition showing monodispersity and particles with a size lower than 40 nm was selected for modification of hydrogels. Such systems were obtained via the photopolymerization performed under the influence of UV radiation using diacrylate poly(ethylene glycol) as a crosslinking agent and 2-hydroxy-2-methylpropiophenone as a photoinitiator. Next, the hydrogels' swelling ability, mechanical properties, wettability and surface morphology were characterized. Moreover, FT-IR spectroscopy, incubation studies in simulated physiological liquids, pro-inflammatory activity analysis and MTT reduction assay with L929 murine fibroblasts were performed. The release profiles of proteins from hydrogels were also verified. Materials modified with proteins showed higher swelling ability, increased flexibility even by 50% and increased surface hydrophilicity. Hydrogels' contact angles were within the range 62-69° while the tensile strength of albumin-containing hydrogels was approx. 0.11 MPa. Furthermore, the possibility of the effective release of protein particles from hydrogels in acidic environment (approximately 70%) was determined. Incubation studies showed hydrogels' stability and lack of their degradation in tested media. The viability of fibroblasts was 89.54% for unmodified hydrogel, and approx. 92.73% for albumin-modified hydrogel, and such an increase indicated the positive impact of the albumin on murine fibroblast proliferation.

Biodegradable particles for protein delivery: Estimation of the release kinetics inside cells.

A methodology to quantify the efficiency of the protein loading and in-vitro delivery for biodegradable capsules with different architectures based on polyelectrolytes (dextran sulfate, poly-L-arginine and polyethylenimine) and SiO2 was developed. The capsules were loaded with model proteins such as ovalbumin and green fluorescent protein (GFP), and the protein release profile inside cells (either macrophages or HeLa cells) after endocytosis was analysed. Both, protein loading and release kinetics were evaluated by analysing confocal laser scanning microscopy images using MatLab and CellProfiler software. Our results indicate that silica capsules showed the most efficient release of proteins as cargo molecules within 48h, as compared to their polymeric counterparts. This developed method for the analysis of the intracellular cargo release kinetics from carrier structures could be used in the future for a better control of drug release profiles.

Microgels Formed by Spontaneous Click Chemistries Utilizing Microfluidic Flow Focusing for Cargo Release in Response to Endogenous or Exogenous Stimuli.

Protein therapeutics have become increasingly popular for the treatment of a variety of diseases owing to their specificity to targets of interest. However, challenges associated with them have limited their use for a range of ailments, including the limited options available for local controlled delivery. To address this challenge, degradable hydrogel microparticles, or microgels, loaded with model biocargoes were created with tunable release profiles or triggered burst release using chemistries responsive to endogenous or exogeneous stimuli, respectively. Specifically, microfluidic flow-focusing was utilized to form homogenous microgels with different spontaneous click chemistries that afforded degradation either in response to redox environments for sustained cargo release or light for on-demand cargo release. The resulting microgels were an appropriate size to remain localized within tissues upon injection and were easily passed through a needle relevant for injection, providing means for localized delivery. Release of a model biopolymer was observed over the course of several weeks for redox-responsive formulations or triggered for immediate release from the light-responsive formulation. Overall, we demonstrate the ability of microgels to be formulated with different materials chemistries to achieve various therapeutic release modalities, providing new tools for creation of more complex protein release profiles to improve therapeutic regimens.

Redox-Responsive Hydrogels for Tunable and "On-Demand" Release of Biomacromolecules.

In recent years, stimuli-responsive degradation has emerged as a desirable design criterion for functional hydrogels to tune the release of encapsulated payload as well as ensure degradation of the gel upon completion of its function. Herein, redox-responsive hydrogels with a well-defined network structure were obtained using a highly efficient thiol-disulfide exchange reaction. In particular, gelation occurred upon combining thiol-terminated tetra-arm polyethylene glycol (PEG) polymers with linear telechelic PEG-based polymers containing pyridyl disulfide units at their chain ends. Rapid gelation proceeds with good conversions (>85%) to yield macroporous hydrogels possessing high water uptake. Furthermore, due to the presence of the disulfide linkages, the thus-obtained hydrogels can self-heal. The obtained hydrogels undergo complete degradation when exposed to environments rich in thiol-containing agents such as dithiothreitol (DTT) and L-glutathione (GSH). Also, the release profile of encapsulated protein, namely, bovine serum albumin, can be tuned by varying the molecular weight of the polymeric precursors. Additionally, it was demonstrated that complete dissolution of the hydrogel to rapidly release the encapsulated protein occurs upon treating these hydrogels with DTT. Cytotoxicity evaluation of the hydrogels and their degradation products indicated the benign nature of these hydrogels. Additionally, the cytocompatible nature of these materials was also evident from a live/dead cell viability assay. One can envision that the facile fabrication and their ability to degrade on-demand and release their payload will make these benign polymeric scaffolds attractive for various biomedical applications.

Fabrication of Multiresponsive Magnetic Nanocomposite Double-Network Hydrogels for Controlled Release Applications.

Nanocomposite double-network hydrogels (ncDN hydrogels) have been demonstrated as promising biomaterials to present several desired properties (e.g., high mechanical strength, stimuli-responsiveness, and local therapy) for biomedicine. Here, a new type of ncDN hydrogels featuring definable microstructures and properties as well as multistimuli responsiveness for controlled release applications is developed. Amine-functionalized iron oxide nanoparticles (IOPs_NH2 ) are used as nanoparticle cross-linkers to simultaneously connect the dual networks of gelatin (Gel) and polydextran aldehyde (PDA) through hydrogen bonding, electrostatic interactions, and dynamic imine bonds. The pH- and temperature-responsive Gel/PDA/IOP_NH2 ncDN hydrogels present a fast release profile of proteins at acidic pH and high temperature. Besides, IOP_NH2 also contributes the magnetic-responsiveness to the ncDN hydrogels, allowing the use of magnetic field to generate heat to facilitate the structural change of hydrogels and the subsequent applications. Taken together, a versatile ncDN hydrogel platform capable of multistimuli responsiveness and local heating for controlled release is developed for advanced biomedical applications.

Amniotic fluid stem cell administration can prevent epithelial injury from necrotizing enterocolitis.

Stem cell therapy has been proven to rescue intestinal injury and stimulate intestinal regeneration in necrotizing enterocolitis (NEC). Specifically, stem cells derived from amniotic fluid (AFSCs) and mesenchymal stem cells (MSCs) derived from bone marrow have shown promising results in the treatment of experimental NEC. This study aims to examine the effects of AFSCs and MSCs on the prevention of intestinal injury during experimental NEC. Supernatants from AFSC and MSC cultures were collected to perform proteomic analysis. Prior to NEC induction, mice received intraperitoneal injections of phosphate-buffered saline (PBS), 2 × 106 AFSCs, or 2 × 106 MSCs. We found that AFSCs grew faster than MSCs. Proteomic analysis indicated that AFSCs are primarily involved in cell development and growth, while MSCs are involved in immune regulation. Administering AFSCs before NEC induction decreased NEC severity and mucosal inflammation. Intestinal proliferation and endogenous stem cell activation were increased after AFSC administration. However, administering MSCs before NEC induction had no beneficial effects. This study demonstrated that AFSCs and MSCs have different protein release profiles. AFSCs can potentially be used as a preventative strategy for neonates at risk of NEC, while MSCs cannot be used. AFSCs and MSCs have distinct protein secretory profiles, and AFSCs are primarily involved in cell development and growth, while MSCs are involved in immune regulation. AFSCs are unique in transiently enhancing healthy intestinal epithelial cell growth, which offers protection against the development of experimental NEC. The prevention of NEC via the administration of AFSCs should be evaluated in infants at great risk of developing NEC or in infants with early signs of NEC.

Antioxidative Composites Based on Multienzyme Systems Encapsulated in Metal-Organic Frameworks.

Skin is exposed to ultraviolet radiation from the sun constantly, which may induce overproduction of reactive oxygen species (ROS) causing oxidative stress to cells and tissues. Enzymes and small molecules work together to maintain the redox homeostasis, among which superoxide dismutase (SOD) and catalase (CAT) are two kinds of most important antioxidants that suffer from the fragile nature of proteins. Moreover, the proportion of two enzymes used in products must be precisely controlled to reduce the damage caused by the toxic intermediate H2O2. Metal-organic frameworks (MOFs) are emerging as promising candidates for multiple enzyme encapsulation due to their high porosity, easy synthesis, and good biocompatibility. Herein, we developed enzyme-MOF composites, SC@ZIF-8, which exhibited an excellent antioxidative activity in vitro. Chemically protective cages formed by MOFs endow the encapsulated enzymes the long-term stability under unnatural conditions in cosmetic and biomedical materials. The pH-dependent protein release profile of SC@ZIF-8 facilitated the successful delivery of enzymes into the cytoplasm to scavenge toxic ROS. The nanocomposites protected human cells from paraquat-induced oxidative stress, paving a new path for the stable and efficient application of antioxidative enzymes in cosmetic and dermatological fields.

Release Profiles of Dyes and Proteins from Calcium Phosphate Microspheres with Different Crystalline Phases

Calcium phosphate is attracting attention as a bone repair material and a controlled-release carrier of various drugs such as bone disease therapeutic agents and anticancer agents. Compared with some bioabsorbable polymers, calcium phosphates have the advantage of preventing a pH decrease in the surrounding body fluid. However, there are few studies comparing the effect of supporting substances with different physicochemical properties on the production of calcium phosphate microspheres with different crystalline phases. In this study, we investigated conditions for obtaining low crystallinity apatite and octacalcium phosphate (OCP) microspheres from calcium carbonate microspheres with different crystalline structures using a simple phosphoric acid treatment. Furthermore, we investigated the adsorption and release behavior of different dyes and proteins from the apatite and OCP microspheres. Overall, the factors governing the adsorption and release behavior are different depending on the molecular size and surface charge of the dye and protein adsorbates.

The Constitutive Extracellular Protein Release by Acute Myeloid Leukemia Cells-A Proteomic Study of Patient Heterogeneity and Its Modulation by Mesenchymal Stromal Cells.

Simple SummaryThe formation of normal blood cells in the bone marrow is supported by a network of non-hematopoietic cells including connective tissue cells, blood vessel cells and bone-forming cells. These cell types support and regulate the growth of acute myeloid leukemia (AML) cells and communicate with leukemic cells through the release of proteins to their common extracellular microenvironment. One of the AML-supporting normal cell types is a subset of connective tissue cells called mesenchymal stem cells. In the present study, we observed that AML cells release a wide range of diverse proteins into their microenvironment, but patients differ both with regard to the number and amount of released proteins. Inhibition of this bidirectional communication through protein release between AML cells and leukemia-supporting normal cells may become a new strategy for cancer treatment.Extracellular protein release is important both for the formation of extracellular matrix and for communication between cells. We investigated the extracellular protein release by in vitro cultured normal mesenchymal stem cells (MSCs) and by primary human acute myeloid leukemia (AML) cells derived from 40 consecutive patients. We observed quantifiable levels of 3082 proteins in our study; for the MSCs, we detected 1446 proteins, whereas the number of released proteins for the AML cells showed wide variation between patients (average number 1699, range 557–2380). The proteins were derived from various cellular compartments (e.g., cell membrane, nucleus, and cytoplasms), several organelles (e.g., cytoskeleton, endoplasmatic reticulum, Golgi apparatus, and mitochondria) and had various functions (e.g., extracellular matrix and exosomal proteins, cytokines, soluble adhesion molecules, protein synthesis, post-transcriptional modulation, RNA binding, and ribonuclear proteins). Thus, AML patients were very heterogeneous both regarding the number of proteins and the nature of their extracellularly released proteins. The protein release profiles of MSCs and primary AML cells show a considerable overlap, but a minority of the proteins are released only or mainly by the MSC, including several extracellular matrix molecules. Taken together, our observations suggest that the protein profile of the extracellular bone marrow microenvironment differs between AML patients, these differences are mainly caused by the protein release by the leukemic cells but this leukemia-associated heterogeneity of the overall extracellular protein profile is modulated by the constitutive protein release by normal MSCs.

Engineered PLGA-PVP/VA based formulations to produce electro-drawn fast\xa0biodegradable microneedles for labile biomolecule delivery

Biodegradable polymer microneedles (MNs) are recognized as non-toxic, safe and stable systems for advanced drug delivery and cutaneous treatments, allowing a direct intradermal delivery and in some cases a controlled release. Most of the microneedles found in the literature are fabricated by micromolding, which is a multistep thus typically costly process. Due to industrial needs, mold-free methods represent a very intriguing approach in microneedle fabrication. Electro-drawing (ED) has been recently proposed as an alternative fast, mild temperature and one-step strategy to the mold-based techniques for the fabrication of poly(lactic-co-glycolic acid) (PLGA) biodegradable MNs. In this work, taking advantage of the flexibility of the ED technology, we engineered microneedle inner microstructure by acting on the water-in-oil (W/O) precursor emulsion formulation to tune drug release profile. Particularly, to promote a faster release of the active pharmaceutical ingredient, we substituted part of PLGA with poly(1-vinylpyrrolidone-co-vinyl acetate) (PVP/VA), as compared to the PLGA alone in the matrix material. Moreover, we introduced lecithin and maltose as emulsion stabilizers. Microneedle inner structural analysis as well as collagenase entrapment efficiency, release and activity of different emulsion formulations were compared to reach an interconnected porosity MN structure, aimed at providing an efficient protein release profile. Furthermore, MN mechanical properties were examined as well as its ability to pierce the stratum corneum on a pig skin model, while the drug diffusion from the MN body was monitored in an in vitro collagen-based dermal model at selected time points.