Engineering M2-type macrophages with a metal polyphenol network for peripheral artery disease treatment.

Functional cell treatment for critical limb ischemia is limited by cell viability loss and dysfunction resulting from a harmful ischemic microenvironment. Metal-polyphenol networks have emerged as novel cell delivery vehicles for protecting cells from the detrimental ischemic microenvironment and prolonging the survival rate of cells in the ischemic microenvironment. M2 macrophages are closely related to tissue repair, and they secrete anti-inflammatory factors that contribute to lesion repair. However, these cells are easily metabolized in the body with low efficiency. Herein, M2 macrophages were decorated with a metal‒polyphenol network that contains copper ions and epigallocatechin gallate (Cu-EGCG@M2) to increase cell survival and therapeutic potential. Cu-EGCG@M2 synergistically promoted angiogenesis through the inherent angiogenesis effect of M2 macrophages and copper ions. We found that Cu-EGCG@M2 increased in vitro viability and strengthened the in vivo therapeutic effect on the ischemic hindlimbs of mice, which promoted the recovery of blood and muscle regeneration, resulting in superior limb salvage. These therapeutic effects were ascribed to the increased survival rate and therapeutic period of M2 macrophages, as well as the ameliorated microenvironment at the ischemic site. Additionally, Cu-EGCG exhibited antioxidant, anti-inflammatory, and proangiogenic effects. Our findings provide a feasible option for cell-based treatment of CLI.

Synergy of antioxidant and M2 polarization in polyphenol-modified konjac glucomannan dressing for remodeling wound healing microenvironment.

Effective skin wound healing and tissue regeneration remain a challenge. Excessive/chronic inflammation inhibits wound healing, leading to scar formation. Herein, we report a wound dressing composed of KGM-GA based on the natural substances konjac glucomannan (KGM) and gallic acid (GA) that accelerates wound healing without any additional drugs. An in vitro study showed that KGM-GA could not only stimulate macrophage polarization to the anti-inflammatory M2 phenotype but also decrease reactive oxygen species (ROS) levels, indicating excellent anti-inflammatory properties. Moreover, in vivo studies of skin wounds demonstrated that the KGM-GA dressing significantly improved wound healing by accelerating wound closure, collagen deposition, and angiogenesis. In addition, it was observed that KGM-GA regulated M2 polarization, reducing the production of intracellular ROS in the wound microenvironment, which was consistent with the in vitro experiments. Therefore, this study designed a multifunctional biomaterial with biological activity, providing a novel dressing for wound healing.

Highly sensitive H2O2-scavenging nano-bionic system for precise treatment of atherosclerosis.

In atherosclerosis, chronic inflammatory processes in local diseased areas may lead to the accumulation of reactive oxygen species (ROS). In this study, we devised a highly sensitive H2O2-scavenging nano-bionic system loaded with probucol (RPP-PU), to treat atherosclerosis more effectively. The RPP material had high sensitivity to H2O2, and the response sensitivity could be reduced from 40 to 10 µmol/L which was close to the lowest concentration of H2O2 levels of the pathological environment. RPP-PU delayed the release and prolonged the duration of PU in vivo. In Apolipoprotein E deficient (ApoE‒/‒) mice, RPP-PU effectively eliminated pathological ROS, reduced the level of lipids and related metabolic enzymes, and significantly decreased the area of vascular plaques and fibers. Our study demonstrated that the H2O2-scavenging nano-bionic system could scavenge the abundant ROS in the atherosclerosis lesion, thereby reducing the oxidative stress for treating atherosclerosis and thus achieve the therapeutic goals with atherosclerosis more desirably.

Redox Modulatory Cu(II)-Baicalein Microflowers Prepared in One Step Effectively Promote Therapeutic Angiogenesis in Diabetic Mice.

Reactive oxygen species (ROS) have been implicated in multiple cellular processes, and an imbalance in redox homeostasis gives rise to diseases, therefore, reestablishing redox homeostasis is a way to cure. Here, copper-based metal-organic networks (Cu-MON) are generated by one-step reaction using anti-inflammatory and antioxidant baicalein as organic ligand and pro-angiogenic copper as metal ions. Phosphate buffered saline is required for triggering Cu-MON formation, and baicalein regulates the morphology and particle size of Cu-MON. Cu-MON are composed of Cu-baicalein complexes (82.08 wt%) and Cu3 (PO4 )2 ·3H2 O (17.92 wt%), thus exhibit a variable catalase-like activity against different H2 O2 levels due to the reversible change between Cu2+ /Cu1+ /Cu0 species. Intramuscular injection of Cu-MON significantly increases blood flow of ischemic limb in diabetic mice, enhances the relative activities of redox-related enzymes in ischemic muscle, thus effectively ameliorating the oxidative damage. Taken together, through moderate and dynamic "precise homeostasis regulation of cells," Cu-MON can be an efficient therapeutic strategy for peripheral arterial disease with diabetic complications.

Corrigendum to "ROS-responsive nanoparticle-mediated delivery of CYP2J2 gene for therapeutic angiogenesis in severe hindlimb ischemia" [Materials Today Bio 13(2022) 100192].

[This corrects the article DOI: 10.1016/j.mtbio.2021.100192.].

H2O2-responsive VEGF/NGF gene co-delivery nano-system achieves stable vascularization in ischemic hindlimbs.

Peripheral vascular disease (PVD) is a common clinical manifestation of atherosclerosis. Vascular endothelial growth factor (VEGF) gene therapy is a promising approach for PVD treatment. However, due to single-gene therapy limitations and high H2O2 pathological microenvironment, VEGF gene therapy are not as expectations and its clinical application are limited. Synergistic effects of Nerve factors and vascular factors in angiogenesis have attracted attention in recent years. In this study, VEGF and nerve growth factor (NGF) genes co-delivery nanoparticles (VEGF/NGF-NPs) were prepared by using H2O2 responsive 6s-PLGA-Po-PEG as a carrier. 6s-PLGA-Po-PEG could react with H2O2 specifically due to the internal peroxalate bond. Angiogenic effects of VEGF/NGF-NPs has been evaluated in cells and hindlimb ischemia mice model. Results showed that VEGF/NGF-NPs promoted VEGF and NGF co-expression simultaneously, eliminated excessive H2O2, strengthened reactions between SH-SY5Ys and HUVECs, and finally enhanced migration, tube formation, proliferation and H2O2 damage resistance of HUVECs. VEGF/NGF-NPs also recovered blood perfusion, promoted the expression of VEGF, NGF, eNOS and NO, and enhanced vascular coverage of pericytes. Treatment effects of VEGF/NGF-NPs may related to VEGF/eNOS/NO pathway. Altogether, VEGF/NGF-NPs eliminated excessive H2O2 while achieving gene co-delivery, and promoted stable angiogenesis. It's a promising way for PVD treatment by using VEGF/NGF-NPs.

Microscopic Imaging Technology Assisted Dynamic Monitoring and Restoration of Micron-Level Cracks in the Painted Layer of Terracotta Warriors and Horses of the Western Han Dynasty.

Cracks are one of the most common issues affecting colored pottery relics; these can be divided into macroscopic cracks, recognizable by the human eye, and micron cracks, which cannot be observed by the naked eye. The gradual development of micron cracks eventually leads to large-scale cracks and the shedding of the coating layer. The repair of such micron cracks poses a key technical difficulty in restoring painted pottery remnants from the Western Han Dynasty. We attempt to solve this problem by reporting on a method that entails the use of a water-borne fluoropolymer material as the adhesive agent, as well as ultra-depth-of-field, digital microscopic imaging technology to build an operating platform for an optical imaging monitoring system. By making simulated ceramic samples, we systematically investigated the influences of water-borne fluoropolymer on chromaticity, adhesion, contact angle, surface morphology, and thermal stability of the paint layer. The results indicate that the color of the painted layer, when treated with the water-borne fluoropolymer, did not change, and the adhesion and contact angle of the painted layer were improved. Additionally, the outcomes of the SEM analysis show that the adhesion and hydrophobicity of the painted layer were improved because the water-borne fluoropolymer filled up the porous structure of the painted layer and covered the pigment particles. These findings demonstrate that aqueous, water-borne fluoropolymer can be used as an adhesive agent for micron cracks. Meanwhile, via the operating platform of the optical imaging monitoring system, the micron cracks of the painted terracotta warriors and horses from the Western Han Dynasty were successfully repaired using the water-borne fluoropolymer. The results imply that the microstructure, size, and geometric spaces of the cracks can be obtained directly utilizing microscopic imaging technology. The dynamic monitoring and imaging system described above can be employed to assist prosthetists in visualizing micro-repair operations in real time, assist with fine visual operations during the repair process, and realize dynamic video recording of the entire repair process. Our work provides a simple visualization method to repair micron-scale cracks in painted pottery relics by applying modern fluoropolymer and ultra-depth-of-field digital microscopic imaging technology.

ROS-responsive nanoparticle-mediated delivery of CYP2J2 gene for therapeutic angiogenesis in severe hindlimb ischemia.

With critical limb ischemia (CLI) being a multi-factorial disease, it is becoming evident that gene therapy with a multiple bio-functional growth factor could achieve better therapeutic outcomes. Cytochrome P450 epoxygenase-2J2 (CYP2J2) and its catalytic products epoxyeicosatrienoic acids (EETs) exhibit pleiotropic biological activities, including pro-angiogenic, anti-inflammatory and cardiovascular protective effects, which are considerably beneficial for reversing ischemia and restoring local blood flow in CLI. Here, we designed a nanoparticle-based pcDNA3.1-CYP2J2 plasmid DNA (pDNA) delivery system (nanoparticle/pDNA complex) composed of a novel three-arm star block copolymer (3S-PLGA-po-PEG), which was achieved by conjugating three-armed PLGA to PEG via the peroxalate ester bond. Considering the multiple bio-functions of CYP2J2-EETs and the sensitivity of the peroxalate ester bond to H2O2, this nanoparticle-based gene delivery system is expected to exhibit excellent pro-angiogenic effects while improving the high oxidative stress and inflammatory micro-environment in ischemic hindlimb. Our study reports the first application of CYP2J2 in the field of therapeutic angiogenesis for CLI treatment and our findings demonstrated good biocompatibility, stability and sustained release properties of the CYP2J2 nano-delivery system. In addition, this nanoparticle-based gene delivery system showed high transfection efficiency and efficient VEGF expression in vitro and in vivo. Intramuscular injection of nanoparticle/pDNA complexes into mice with hindlimb ischemia resulted in significant rapid blood flow recovery and improved muscle repair compared to mice treated with naked pDNA. In summary, 3S-PLGA-po-PEG/CYP2J2-pDNA complexes have tremendous potential and provide a practical strategy for the treatment of limb ischemia. Moreover, 3S-PLGA-po-PEG nanoparticles might be useful as a potential non-viral carrier for other gene delivery applications.

ROS-responsive EPO nanoparticles ameliorate ionizing radiation-induced hematopoietic injury.

Stimulus-responsive polymer materials have attracted much attention as drug carriers because of the ability to deliver drugs to the active site. Reactive oxygen species (ROS) play crucial roles in cellular signaling and regulation of oxygen homeostasis. However, ROS are present in abnormally high levels in many pathological environments. Based on the above points, three-arm poly(lactic-co-glycolic acid)-PO-poly(ethylene glycol) (3s-PLGA-PO-PEG or simply PP) was synthesized by using peroxalate esters (PO) as hydrogen peroxide-responsive linkages. PP was used to deliver promote hematopoietic recovery drugs erythropoietin (EPO) and EPO nanoparticles (EPO NPs) were prepared. We established a hematopoietic system injury model by ionizing radiation (IR) and unexpectedly found the good therapeutic effect of blank PP. Moreover, the administration of EPO NPs obviously decreased IR-induced ROS in bone marrow cells (BMCs) and reconstituted hematopoietic stem cells in BMCs. This study reveals a novel ROS-responsive polymer material that could be employed to remove excess ROS in the lesion and promote the efficacy of drug therapy.

Significant difference between sirolimus and paclitaxel nanoparticles in anti-proliferation effect in normoxia and hypoxia: The basis of better selection of atherosclerosis treatment.

Compared with paclitaxel, sirolimus has been more used in the treatment of vascular restenosis gradually as an anti-proliferative drug, but few basic studies have elucidated its mechanism. The anti-proliferative effects of sirolimus or paclitaxel have been demonstrated by numerous studies under normoxia, but few studies have been achieved focusing hypoxia. In this study, porcine carotid artery injury model and classical cobalt chloride hypoxia cell model were established. Sirolimus nanoparticles (SRM-NPs), paclitaxel nanoparticles (PTX-NPs) and blank nanoparticles (Blank-NPs) were prepared respectively. The effect of RPM-NPs on the degree of stenosis, proliferative index and the expression of PCNA after 28 days of porcine carotid artery injury model was evaluated. Compared with saline group and SRM groups, SRM-NPs group suppressed vascular stenosis, proliferative index and the expression of PCNA (P < 0.01 and P < 0.05). Endothelial cell (EC) and smooth muscle cell (SMC) were pre-treated with cobaltous chloride, followed by SRM-NPs, PTX-NPs, Blank-NPs or PBS control treating, the effects on cell proliferation, HIF-1 expression and glycolysis were detected. SRM-NPs could inhibit EC and SMC proliferation under hypoxia, while PTX-NPs couldn't (P < 0.001). Significant differences between sirolimus and paclitaxel NPs in anti-proliferation effect under normoxia and hypoxia may due to the different inhibitory effects on HIF-1α expression and glycolysis. In conclusion, these results suggest that sirolimus can inhibit the proliferation of hypoxic cells more effectively than paclitaxel. These observations may provide a basis for understanding clinical vascular stenosis therapeutic differences between rapamycin and paclitaxel.

ROS-responsive capsules engineered from EGCG-Zinc networks improve therapeutic angiogenesis in mouse limb ischemia.

The successful treatment of limb ischemia requires that promote angiogenesis along with microenvironment improvement. Zinc ions have been reported to stimulate angiogenesis, but application was limited to the toxicity concerns. We hypothesized that zinc based metal-EGCG capsule (EGCG/Zn Ps) can achieve sustained release Zn2+ resulting in reduced toxicity and improve angiogenesis as well as the improvement of microenvironment by ROS scavenging of EGCG. The surface morphology, zeta potential, infrared absorbance peaks and zinc ion release profile of the EGCG/Zn Ps were measured. In vitro, EGCG/Zn showed significantly antioxidant, anti-inflammatory and induced cell migration effect. In addition, EGCG/Zn Ps enabled the sustained release of zinc ions, which reduced cytotoxicity and enhanced the secretion of vascular endothelial growth factor (VEGF) in vitro and in vivo. In mouse models of limb ischemia, EGCG/Zn Ps promoted angiogenesis and cell proliferation in ischemic tissues. Moreover, EGCG/Zn Ps group exhibited the most significant recovery of limb ischemic score, limb temperature and blood flow than other groups. In conclusion, EGCG/Zn Ps is a safe and promising approach to combine the merit of Zn2+ and EGCG, thus enabling the direct application to limb ischemia.

Construction and application of therapeutic metal-polyphenol capsule for peripheral artery disease.

Lesions of patients with peripheral artery disease (PAD) are in a harmful microenvironment, which features increased oxidative stress and inflammatory infiltration. Hence, it is essential to improve the microenvironment along with angiogenesis. In this study, metal-polyphenol capsules (Cu-EGCG), which combines the therapeutic anti-inflammatory and antioxidant activities of EGCG and the angiogenic activity of copper ions, were synthesized through coordination between EGCG and copper ions. The sustained release of the copper ions from Cu-EGCG was demonstrated in vitro, and biocompatible Cu-EGCG can scavenge intracellular ROS, reduce cell death in the presence of cytotoxic levels of ROS, and decrease the expression of pro-inflammatory cytokines (TNF-α, IL-6). Moreover, Cu-EGCG induced the secretion of vascular endothelial growth factor (VEGF) in a hindlimb ischaemia model of PAD. More importantly, the upregulated expression of platelet endothelial cell adhesion molecule-1 (CD31) and proliferating cell nuclear antigen (PCNA) in ischaemic tissues indicated the remarkable effect of Cu-EGCG on angiogenesis. In addition, Cu-EGCG showed significant blood recovery in ischaemic hindlimbs. Taking these results together, biocompatible Cu-EGCG with therapeutic functions holds great potential applications for PAD therapy.

Protein delivery nanosystem of six-arm copolymer poly(ε-caprolactone)-poly(ethylene glycol) for long-term sustained release.

BACKGROUND: To address the issue of delivery of proteins, a six-arm copolymer, six-arm poly (ε-caprolactone)-poly(ethylene glycol) (6S-PCL-PEG), was synthesized by a simple two-step reaction. Thereafter, the application of 6S-PCL-PEG as a protein carrier was evaluated. MATERIALS AND METHODS: A six-arm copolymer, six-arm poly(ε-caprolactone) (6S-PCL), was synthesized by ring-opening polymerization, with stannous octoate as a catalyst and inositol as an initiator. Then, poly(ethylene glycol) (PEG) was linked with 6S-PCL by oxalyl chloride to obtain 6S-PCL-PEG. Hydrogen-1 nuclear magnetic resonance spectrum, Fourier-transform infrared spectroscopy, and gel-permeation chromatography were conducted to identify the structure of 6S-PCL-PEG. The biocompatibility of the 6S-PCL-PEG was evaluated by a cell counting kit-8 assay. Polymeric nanoparticles (NPs) were prepared by a water-in-oil-in-water double emulsion (W1/O/W2) solvent evaporation method. The size distribution and zeta potential of NPs were determined by dynamic light scattering. Transmission electron microscopy was used to observe the morphology of NPs. Drug-loading capacity, encapsulation efficiency, and the release behavior of ovalbumin (OVA)-loading NPs were tested by the bicinchoninic acid assay kit. The stability and activity of OVA released from NPs were detected and the uptake of NPs was evaluated by NIH-3T3 cells. RESULTS: All results indicated the successful synthesis of amphiphilic copolymer 6S-PCL-PEG, which possessed excellent biocompatibility and could formulate NPs easily. High drug-loading capacity and encapsulation efficiency of protein NPs were observed. In vitro, OVA was released slowly and the bioactivity of OVA was maintained for over 28 days. CONCLUSION: 6S-PCL-PEG NPs prepared in this study show promising potential for use as a protein carrier.

Improved vaccine-induced immune responses via a ROS-triggered nanoparticle-based antigen delivery system.

Subunit vaccines that are designed based on recombinant antigens or peptides have shown promising potential as viable substitutes for traditional vaccines due to their better safety and specificity. However, the induction of adequate in vivo immune responses with appropriate effectiveness remains a major challenge for vaccine development. More recently, the implementation of a nanoparticle-based antigen delivery system has been considered a promising approach to improve the in vivo efficacy for subunit vaccine development. Thus, we have designed and prepared a nanoparticle-based antigen delivery system composed of three-armed PLGA, which is conjugated to PEG via the peroxalate ester bond (3s-PLGA-PO-PEG) and PEI as a cationic adjuvant (PPO NPs). It is known that during a foreign pathogen attack, NADPH, an oxidase, of the host organism is activated and generates an elevated level of reactive oxygen species, hydrogen peroxide (H2O2) primarily, as a defensive mechanism. Considering the sensitivity of the peroxalate ester bond to H2O2 and the cationic property of PEI for the induction of immune responses, this 3s-PLGA-PO-PEG/PEI antigen delivery system is expected to be both ROS responsive and facilitative in antigen uptake without severe toxicity that has been reported with cationic adjuvants. Indeed, our results demonstrated excellent loading capacity and in vitro stability of the PPO NPs encapsulated with the model antigen, ovalbumin (OVA). Co-culturing of bone marrow dendritic cells with the PPO NPs also led to enhanced dendritic cell maturation, antigen uptake, enhanced lysosomal escape, antigen cross-presentation and in vitro CD8+ T cell activation. In vivo experiments using mice further revealed that the administration of the PPO nanovaccine induced robust OVA-specific antibody production, upregulation of splenic CD4+ and CD8+ T cell proportions as well as an increase in memory T cell generation. In summary, we report here a ROS-triggered nanoparticle-based antigen delivery system that could be employed to promote the in vivo efficacy of vaccine-induced immune responses.

Nanoparticles with CD44 Targeting and ROS Triggering Properties as Effective in Vivo Antigen Delivery System.

Currently, development of subunit vaccine based on recombinant antigens or peptides has gradually become an important alternative option for traditional vaccine. However, induction of potent immune response with desired efficacy remains a major challenge. The nanoparticle-based antigen delivery system has been considered a potential carrier system to improve the efficacy of subunit vaccine. In the present study, we have designed an immune-stimulatory delivery system by conjugating three-armed PLGA to PEG via the peroxalate ester bond which is sensitive to hydrogen peroxide (H2O2), a major reactive oxygen species (ROS). Hyaluronic acid (HA), a ligand for CD44 receptors was also modified onto the outer shell of the 3s-PLGA-PEG nanoparticles to promote immune cell uptake. For in vitro and in vivo immune response assessment, a model antigen ovalbumin (OVA) was enclosed within the core of the 3s-PLGA-PEG nanoparticles to form 3s-PLGA-PO-PEG/HA nanoparticles (PHO NPs). Our results showed that the PHO NPs enhanced dendritic cell maturation, antigen uptake, and antigen presentation in vitro, likely due to enhanced lysosomal escape. In vivo experiments further revealed that the PHO nanovaccine robustly promoted OVA-specific antibody production and T cell response accompanied by modest stimulation of memory T cells. In summary, the ROS-responsive PHO NPs with modified HA may be an effective vehicle antigen delivery system to promote antigen-induced immune response.