Facile Preparation of Doxorubicin-Loaded and Folic Acid-Conjugated Carbon Nanotubes@Poly(<i>N</i>‑vinyl pyrrole) for Targeted Synergistic Chemo–Photothermal Cancer Treatment

We developed a bifunctional nanoplatform for targeted synergistic chemo–photothermal cancer treatment. The nanoplatform was constructed through a facile method in which poly­(<i>N</i>-vinyl pyrrole) (PVPy) was coated on cut multiwalled carbon nanotubes (c-MWNTs); FA-PEG-SH was then linked by thiol–ene click reaction to improve the active targeting ability, water dispersibility, and biocompatibility and to extend the circulation time in blood. The PVPy shell not only enhanced the photothermal effect of c-MWNTs significantly but also provided a surface that could tailor targeting molecules and drugs. The resulting MWNT@PVPy-S-PEG-FA possessed high drug-loading ratio as well as pH-sensitive unloading capacity for a broad-spectrum anticancer agent, doxorubicin. Owing to its outstanding efficiency in photothermal conversion and ability in targeted drug delivery, the material could potentially be used as an efficient chemo–photothermal therapeutic nanoagent to treat cancer

A Novel Approach to Prepare Protein-proanthocyanidins Nano-complexes by the Reversible Assembly of Ferritin Cage

The application of proanthocyanidins (PCs) in food industry is limited because of its instability and vulnerability against oxidative damage. In this work, by taking advantage of the reversible assembly characteristic of ferritin, apo-red bean ferritin (apoRBF)-PCs composites (FPs) were designed, and results showed a ratio of 1/10 of PCs (PCs/apoRBF, w/w) were encapsulated into a ferritin cage, and the encapsulation ratio was 23.8%. The FPs exhibited a homogeneous form, and maintained a spherical morphology. Further more, the thermal and light stability of PCs in FPs was significantly increased compared to free PCs (p < 0.05). The in vitro digestion indicated that apoRBF could prolong the release of PCs in simulated gastrointestinal tract. Additionally, the antioxidant activity of FPs was partly retained (56.6%) as compared to free PCs. This work offers a novel method to encapsulate and stabilize polyphenols and might be favorable for the application of ferritin in food industry

Urea-Driven Epigallocatechin Gallate (EGCG) Permeation into the Ferritin Cage, an Innovative Method for Fabrication of Protein–Polyphenol Co-assemblies

The 8 nm diameter cavity endows the ferritin cage with a natural space to encapsulate food components. In this work, urea was explored as a novel medium to facilitate the formation of ferritin–polyphenol co-assemblies. Results indicated that urea (20 mM) could expand the 4-fold channel size of apo-red bean ferritin (apoRBF) with an increased initial iron release rate υ₀ (0.22 ± 0.02 μM min^–1) and decreased α-helix content (5.6%). Moreover, urea (20 mM) could facilitate the permeation of EGCG into the apoRBF without destroying the ferritin structure and thus form ferritin–EGCG co-assemblies (FECs) with an encapsulation ratio and loading capacity of 17.6 and 2.1% (w/w), respectively. TEM exhibited that FECs maintained a spherical morphology with a 12 nm diameter in size. Fluorescence analysis showed that urea intervention could improve the binding constant <i>K</i> [(1.22 ± 0.8) × 10⁴ M^–1] of EGCG to apoRBF. Furthermore, the EGCG thermal stability was significantly improved (20–60 °C) compared with free EGCG. Additionally, this urea-involved method was applicable for chlorogenic acid and anthocyanin encapsulation by the apoRBF cage. Thus, urea shows potential as a novel potential medium to encapsulate and stabilize bioactive polyphenols for food usage based on the ferritin protein cage structure

Evaluation of topical methylene blue nanoemulsion for wound healing in diabetic mice

AbstractContext Diabetic wounds (DW) are a complication of diabetes and slow wound healing is the main manifestation. Methylene blue (MB) has been shown to exhibit therapeutic effects on diabetes-related diseases.Objective To investigate the mechanisms of action of MB-nanoemulsion (NE) in the treatment of DW.Materials and methods The concentration of MB-NE used in the in vivo and in vitro experiments was 0.1 mg/mL. Streptozocin-induced diabetic mice were used as models. The mice were separated into nondiabetic, diabetic, MB-NE treated, and NE-treated groups. Intervention of high glucose-induced human umbilical vein endothelial cells using MB-NE. The mechanism by which MB-NE promotes DW healing is investigated by combining histological analysis, immunofluorescence analysis, TUNEL and ROS assays and western blotting.Results In diabetic mice, the MB-NE accelerated DW healing (p < 0.05), promoted the expression of endothelial cell markers (α-SMA, CD31 and VEGF) (p < 0.05), and reduced TUNEL levels. In vitro, MB accelerated the migration rate of cells (p < 0.05); promoted the expression of CD31, VEGF, anti-apoptotic protein Bcl2 (p < 0.05) and decreased the expression of the pro-apoptotic proteins cleaved caspase-3 and Bax (p < 0.05). MB upregulated the expression of Nrf2, catalase, HO-1 and SOD2 (p < 0.05). In addition, MB reduced the immunofluorescence intensity of TUNEL and ROS in cells and reduced apoptosis. The therapeutic effect of MB was attenuated after treatment with an Nrf2 inhibitor (ML385).Discussion and conclusion This study provides a foundation for the application of MB-NE in the treatment of DW

A novel approach to prepare protein-proanthocyanidins nano-complexes by the reversible assembly of ferritin cage

The application of proanthocyanidins (PCs) in food industry is limited because of its instability and vulnerability against oxidative damage. In this work, by taking advantage of the reversible assembly characteristic of ferritin, apo-red bean ferritin (apoRBF)-PCs composites (FPs) were designed, and results showed a ratio of 1/10 of PCs (PCs/apoRBF, w/w) were encapsulated into a ferritin cage, and the encapsulation ratio was 23.8%. The FPs exhibited a homogeneous form, and maintained a spherical morphology. Further more, the thermal and light stability of PCs in FPs was significantly increased compared to free PCs (p < 0.05). The in vitro digestion indicated that apoRBF could prolong the release of PCs in simulated gastrointestinal tract. Additionally, the antioxidant activity of FPs was partly retained (56.6%) as compared to free PCs. This work offers a novel method to encapsulate and stabilize polyphenols and might be favorable for the application of ferritin in food industry

A highly sensitive bead-based flow cytometric competitive binding assay to detect SARS-CoV-2 neutralizing antibody activity

Accurate detection of SARS-CoV-2 neutralizing antibody (nAb) is critical for assessing the immunity levels after virus infection or vaccination. As fast, cost-effective alternatives to viral infection-based assays, competitive binding (CB) assays were developed to quantitate nAb by monitoring the ability of sera to inhibit the binding of viral spike (S) protein to the angiotensin converting enzyme 2 (ACE2) receptor. Herein, we established a bead-based flow cytometric CB assay and tested the detection performance of six combination models, i.e. immobilized ACE2 and soluble Fc-tagged S1 subunit of S protein (iACE2/S1-Fc), immobilized ACE2 and soluble Fc-tagged receptor binding domain (RBD) of S protein (iACE2/RBD-Fc), immobilized S1 and soluble Fc-tagged ACE2 (iS1/ACE2-Fc), immobilized S1 and soluble His-tagged ACE2 (iS1/ACE2-His), immobilized RBD and soluble Fc-tagged ACE2 (iRBD/ACE2-Fc), and immobilized RBD and soluble His-tagged ACE2 (iRBD/ACE2-His). Using SARS-CoV-2 monoclonal antibodies and sera of convalescent COVID-19 patients and vaccinated subjects, the combination models iACE2/RBD-Fc, iACE2/S1-Fc and iS1/ACE2-His were identified to be able to specifically detect SARS-CoV-2 nAb, among which iACE2/RBD-Fc model showed the highest sensitivity, superior to a commercial SARS-CoV-2 surrogate virus neutralization test (sVNT) ELISA kit. Further studies demonstrated that the sensitivity and specificity of CB assays were affected by the tag of ACE2, type of spike and method of measuring binding rate between ACE2 and spike. Moreover, the iACE2/RBD-Fc model showed good performance in detecting kinetic development of nAb against both the prototype SARS-CoV-2 strain and an omicron variant of SARS-CoV-2 in people immunized by an inactivated SARS-CoV-2 vaccine, and the results of iACE2/RBD-Fc model are correlated well with those of live virus-based and pseudovirus-based neutralization tests, demonstrating the potential to be developed into a highly sensitive, specific, versatile and high-throughput method for detecting SARS-CoV-2 nAb in clinical practice

DataSheet_1_A highly sensitive bead-based flow cytometric competitive binding assay to detect SARS-CoV-2 neutralizing antibody activity.docx

Accurate detection of SARS-CoV-2 neutralizing antibody (nAb) is critical for assessing the immunity levels after virus infection or vaccination. As fast, cost-effective alternatives to viral infection-based assays, competitive binding (CB) assays were developed to quantitate nAb by monitoring the ability of sera to inhibit the binding of viral spike (S) protein to the angiotensin converting enzyme 2 (ACE2) receptor. Herein, we established a bead-based flow cytometric CB assay and tested the detection performance of six combination models, i.e. immobilized ACE2 and soluble Fc-tagged S1 subunit of S protein (iACE2/S1-Fc), immobilized ACE2 and soluble Fc-tagged receptor binding domain (RBD) of S protein (iACE2/RBD-Fc), immobilized S1 and soluble Fc-tagged ACE2 (iS1/ACE2-Fc), immobilized S1 and soluble His-tagged ACE2 (iS1/ACE2-His), immobilized RBD and soluble Fc-tagged ACE2 (iRBD/ACE2-Fc), and immobilized RBD and soluble His-tagged ACE2 (iRBD/ACE2-His). Using SARS-CoV-2 monoclonal antibodies and sera of convalescent COVID-19 patients and vaccinated subjects, the combination models iACE2/RBD-Fc, iACE2/S1-Fc and iS1/ACE2-His were identified to be able to specifically detect SARS-CoV-2 nAb, among which iACE2/RBD-Fc model showed the highest sensitivity, superior to a commercial SARS-CoV-2 surrogate virus neutralization test (sVNT) ELISA kit. Further studies demonstrated that the sensitivity and specificity of CB assays were affected by the tag of ACE2, type of spike and method of measuring binding rate between ACE2 and spike. Moreover, the iACE2/RBD-Fc model showed good performance in detecting kinetic development of nAb against both the prototype SARS-CoV-2 strain and an omicron variant of SARS-CoV-2 in people immunized by an inactivated SARS-CoV-2 vaccine, and the results of iACE2/RBD-Fc model are correlated well with those of live virus-based and pseudovirus-based neutralization tests, demonstrating the potential to be developed into a highly sensitive, specific, versatile and high-throughput method for detecting SARS-CoV-2 nAb in clinical practice.</p