Fabrication of pH- and Thermoresponsive Three-Layered Micelles via Host-Guest Interactions.

Well-defined ß-cyclodextrin (ß-CD)-terminated double hydrophilic diblock copolymers (DHBCs), ß-CD- poly(di(ethylene glycol) methyl ether methacrylate) (PMEO2 MA)-b-poly(2-(diethylamino)ethyl methacrylate) (PDEA) (BP1) and ß-CD-PDEA-b-PMEO2 MA (BP2), are synthesized via sequential atom transfer radical polymerizations of di(ethylene glycol) methyl ether methacrylate and 2-(diethylamino)ethyl methacrylate using alkynyl-functionalized initiator, followed by click reactions with an excess of mono-azido-substituted ß-cyclodextrin (ß-CD-N3 ). The micellization behavior of these as-prepared DHBCs in aqueous solutions suffers from insufficient colloidal stabilities at basic pH and high temperatures (e.g., pH 9 and 45 °C), resulting in the formation of macroscopic precipitations. However, the stabilities of colloidal nanoparticles can be remarkably enhanced as a result of the convenient formation of three-layered micelles by taking advantage of host-guest interactions of BP1/BP2 and Ad-terminated poly(ethylene glycol). The pH- and thermoresponsive three-layered micelles with enhanced stability may augur promising applications in targeted drug delivery and controlled intelligent release.

Solution-processable graphene quantum dots.

This minireview describes recent progress in solution-processable graphene quantum dots (SGQDs). Advances in the preparation, modification, properties, and applications of SGQDs are highlighted in detail. As one of emerging nanostructured materials, possible ongoing research related to the precise control of the lateral size, edge structure and surface functionality; the manipulation and characterization; the relationship between the properties and structure; and interfaces with biological systems of SGQDs have been speculated upon.

Characterization of the anti-pathogenic, genomic and phenotypic properties of a Lacticaseibacillus rhamnosus VHProbi M14 isolate.

A strain of lactic acid bacteria from cheese was isolated, that showed strong growth inhibitory effects on Streptococcus mutans. The API 50CH system and 16S rDNA sequencing verified that this was a novel strain, and was named Lacticaseibacillus rhamnosus VHProbi M14. The strain inhibited the growth of S. mutans and Fusobacterium nucleatum under mixed culture conditions, coaggregated with S. mutans and F. nucleatum, and reduced the adhesion of S. mutans and F. nucleatum on cultured human primary gingival epithelial (HPGE) cells. The pH, peroxidase and protease sensitivity testing found antibacterial substances of protein- and peptide-like structures in addition to organic acids. The antimicrobial substances were sensitive to hydrolysis with trypsin, papain and pineapple protease and were inactived at temperatures above 100°C. Ammonium sulphate-precipitated proteins from the M14 strain retained the ability to inhibit the growth of S. mutans and F. nucleatum. The M14 strain contained 23 bacteriocin-related genes encoding for metabolites, belonging to class II bacteriocins. The M14 strain also showed inhibitory effects on 8 other pathogenic strains (A. actinomycetemcomitans, C. albicans, E. coli, G. vaginalis, P. acnes, P. gingivalis, S. aureus, S. enteritids), and thus has a broad spectrum of bacterial inhibition. This new isolate has been identified as having potential to be used as a probiotic bacterium in clinical applications.

Long-Term Immunogenicity and In Vitro Prophylactic Protective Efficacy of M. tuberculosis Fusion Protein DR2 Combined with Liposomal Adjuvant DIMQ as a Boosting Vaccine for BCG.

The resuscitation of dormant Mycobacterium tuberculosis is an important cause of adult tuberculosis (TB) transmission. According to the interaction mechanism between M. tuberculosis and the host, the latency antigen Rv0572c and region of difference 9 (RD9) antigen Rv3621c were selected in this study to prepare the fusion protein DR2. Stimulating clinically diagnosed active tuberculosis infections (i.e., TB patients), latent tuberculosis infections, and healthy controls confirmed that T lymphocytes could recognize DR2 protein in the peripheral blood of TB-infected individuals more than subcomponent protein. The DR2 protein was then emulsified in the liposome adjuvant dimethyl dioctadecyl ammonium bromide, and imiquimod (DIMQ) was administered to C57BL/6 mice immunized with Bacillus Calmette-Guérin (BCG) vaccine to evaluate their immunogenicity. Studies have shown that DR2/DIMQ, a booster vaccine for BCG primary immunization, can elicit robust CD4+ Th1 cell immune response and predominant IFN-γ+ CD4+ effector memory T cells (TEM) subsets. Furthermore, the serum antibody level and the expression of related cytokines increased significantly with the extension of immunization time, with IL2+, CD4+, or CD8+ central memory T cells (TCM) subsets predominant in the long term. This immunization strategy showed matched prophylactic protective efficacy by performing in vitro challenge experiment. This result provides robust evidence that the novel subunit vaccine prepared by fusion protein DR2 combined with liposomal adjuvant DIMQ is a promising TB vaccine candidate for further preclinical trials as a booster vaccine for BCG.

Identification of a new probiotic strain, Lactiplantibacillus plantarum VHProbi® V38, and its use as an oral health agent.

Introduction: Probiotics can be used to treat oral diseases such as dental caries, gingivitis, periodontitis, and halitosis. Methods: This study screened for strains capable of inhibiting Streptococcus mutans,one of the primary pathogenic bacteria responsible for dental caries by agar diffusion in different samples. Strain identification was performed by 16S rDNA sequencing and the API 50CH system. The potential functions of the strains in terms of oral health properties were also tested by agglutination assays, growth inhibition assays, adhesion assays, biofilm removal assays and inhibition of adhesion in human primary gingival epithelial (HPGE) cells assays. Results: This study identified a probiotic strain from fermented cabbages that has a strong inhibitory effect on Streptococcus mutans. The API 50CH system and 16S rDNA sequencing verified that this was a new strain and it was given the name, Lactiplantibacillus plantarum VHProbi®V38. Agglutination, growth inhibition and adhesion, and biofilm removal tests indicated that L. plantarum VHProbi® V38 inhibited and reduced S. mutans. This probiotic was shown to have a broad antibacterial spectrum, simultaneously inhibiting the growth of periodontal pathogenic bacteria such as Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, and Fusobacterium nucleatum. After 2 hours of co-cultivation with these pathogens, L. plantarum VHProbi® V38 was able to significantly reduce pathogens adhesion on human primary gingival epithelial (HPGE) cells. Discussion: These findings suggest that L. plantarum VHProbi® V38 could potentially prevent and treat periodontal diseases caused by these pathogenic bacteria. L. plantarum VHProbi® V38 also adheres strongly to HPGE cells and thus has potential as an oral probiotic. This study describes new methods that can be used to aid the screening and identification of oral probiotics.

Enhanced immunogenicity of the tuberculosis subunit Rv0572c vaccine delivered in DMT liposome adjuvant as a BCG-booster.

COVID-19 has affected the progress made in the prevention and treatment of tuberculosis (TB); hence, the mortality of tuberculosis has risen. Different strategies-based novel TB vaccine candidates have been developed. This study identifies strategies to overcome the limitations of Bacille Calmette-Guérin (BCG) in preventing latent infection and reactivation of TB. The latency antigen Rv0572c was selected based on the mechanism of interaction between Mycobacterium tuberculosis and its host. The rRv0572c protein was used to stimulate whole blood samples derived from patients with clinically diagnosed active TB (ATBs) or latent TB infections (LTBIs) and healthy control (HCs) donors, confirming that this protein can be recognized by T cells in patients with TB, especially LTBIs. C57BL/6 mice were used to investigate the immunogenicity of the rRv0572c protein emulsified in the liposome adjuvant dimethyldioctadecylammonium [DDA], monophosphoryl lipid A [MPLA], trehalose-6, 6'-dibehenate [TDB] (DMT). The results demonstrated that rRv0572c/DMT could boost BCG-primed mice to induce antigen-specific CD4+ T cell production and generate functional T cells dominated by antigen-specific CD8+ T cells. The rRv0572c/DMT vaccine could also trigger limited Th2 humoral immune responses. These findings suggest that rRv0572c/DMT is a potential subunit vaccine candidate that can be used as a booster vaccine for BCG.

Broad-Spectrum Bactericidal Activity and Remarkable Selectivity of Main-Chain Sulfonium-Containing Polymers with Alternating Sequences.

Incorporation of cationic groups into polymers represents one of the most widely used strategies to prepare antibacterial materials. Sulfonium, as a typical cationic moiety, displays potent antibacterial efficacy in the form of small molecules, however, has long underperformed in polymeric systems. Herein, we developed a series of alternating polysulfoniums, where the hydrophobicity of each alternating unit can be accurately tuned by altering the monomer precursors. Excellent antibacterial activity against a broad spectrum of clinically relevant bacteria, including Methicillin-resistant Staphylococcus aureus, can be obtained in the optimal compositions with minimum bactericidal concentrations in the range of 1.25-10 µg/mL, as well as negligible hemolytic effect at polymer concentrations even up to 10000 µg/mL. Bacteria do not readily develop resistance to polysulfoniums due to the antibacterial action is possibly the membrane disrupting mechanism. This work demonstrates sulfonium-based polymers with well-defined sequences can function as a promising candidate to combat drug-resistant bacterial infection.

[Clinical observation of Flos magnoliae volatile oil nano-liposome nasal drops in treating pediatric allergic rhinitis].

OBJECTIVE: To observe the clinical efficacy of Flos Magnoliae volatile oil nano-liposome nasal drops (FMO) in treating pediatric allergic rhinitis (PAR). METHODS: Adopting parallel controlled method, the 191 patients with PAR were randomized into two groups. The observation group was treated with FMO, and the control group with Cetirizine. The clinical efficacy, main symptoms, signs, syndromes scores of Chinese medicine, and peripheral eosinophil (EOS) count were observed after 3-week treatment. RESULTS: In the observation group, the total effective rate was 94.84%, which was higher than that in the control group (78.72%); the effective rate on alleviating main symptoms (sneezing, nasal obstruction), signs (nasal mucosa edema, pallor) and the EOS count were significantly lowered, all were better than those in the control group (P <0.05). CONCLUSION: FMO has some positive effects on PAR, it might be realized by lowering the peripheral EOS.

Photo-induced hydrogen-bonding complexes for drug periodic release.

The construction of intelligent supramolecular nanocarriers has received much attention for their potential application in chemotherapy. Herein, we report the successful design and synthesis of a photoreactive monomer, N'-(2-nitrobenzyl)-N-acryloyl glycinamide (NBNAGA). Using a poly(ethylene glycol) (PEG)-based macro-RAFT agent, the amphiphilic diblock copolymer (BCP), PEG-b-PNBNAGA, was prepared through a reversible addition-fragmentation chain-transfer (RAFT) polymerization. Then, photoresponsive polymeric micelles (PMs) were fabricated with the hydrophilic PEG shielding coronas and hydrophobic PNBNAGA inner cores via the self-assembly of PEG-b-PNBNAGA BCPs. Upon 365 nm UV light irradiation, the o-nitrobenzyl groups in the micellar cores were removed with freshly formed amide moieties. 1. Combined with the original amide moieties dual hydrogen bond interactions in the side chains came into being, subsequently changing the PMs' cores from hydrophobic to hydrophilic. Thus, the photo-induced dual hydrogen-bonding complex rather than a hydrophobic interaction assembly with inviable nanostructures was achieved, which gave rise to the first stage of doxorubicin (DOX) release. During the second period, the noncovalent cross-linked PMs underwent further structural disintegration upon heating with dissociation of the dual hydrogen bonds, resulting in the sequential DOX release. In other words, periodic drug release was successfully accomplished via a photoirradiation-induced mechanism modification of micellar cores formation and then by subsequent heating-induced hydrogen-bonding complex disruption, thus indicating its promise for use in therapeutics synergistic delivery in severe disease therapy.

Chain-length dependence of diblock copolymer micellization kinetics studied by stopped-flow pH-jump.

A series of well-defined poly(ethylene oxide)- b-poly(2-(diethylamino)ethyl methacrylate) (PEO- b-PDEA) diblock copolymers containing PEO block of identical chain length and PDEA block with varying degrees of polymerization (DP, in the range of 32-154) were prepared via atom transfer radical polymerization (ATRP) employing a PEO-based macroinitiator (DP = 113). Upon a pH-jump from 3 to 12 under highly efficient stopped-flow mixing conditions, PEO- b-PDEA copolymers spontaneously form spherical micelles of increasing sizes and aggregation numbers ( N agg) with increasing PDEA chain lengths. Stopped-flow light scattering technique was used to probe the pH-induced micellization kinetics of PEO- b-PDEA copolymers, aiming to elucidate the PDEA chain-length effects on the unimer-to-micelle transition process. Upon a stopped-flow pH-jump from 3 to 12, the obtained dynamic traces can be well-fitted with double exponential functions. The calculated fast and slow characteristic relaxation times (tau 1 and tau 2) can be ascribed to the formation of quasi-equilibrium micelles (fast process) and subsequent relaxation into final equilibrium micelles (slow process), respectively. For PEO 113- b-PDEA 32 and PEO 113- b-PDEA 61, tau 2 is almost independent of polymer concentrations, suggesting that the relaxation from quasi-equilibrium micelles into final equilibrium micelles mainly proceeds via insertion/expulsion of unimer chains. Upon increasing the DP of pH-responsive PDEA block to 89, 117, and 154, the obtained slow relaxation time, tau 2, tends to decrease with increasing polymer concentrations, suggesting that the slow process is dominated by the micelle fusion/fission mechanism. The apparent activation energy ( E a) associated with tau 2 has also been determined from temperature-dependent micellization kinetics for five PEO- b-PDEA copolymers. It was found that during micellization, copolymers with longer PDEA blocks exhibit much lower E a compared to those with shorter blocks. Thus, we observed experimentally for the first time that increasing the hydrophobic block length in double hydrophilic block copolymers (DHBCs) can transform the mechanism of the slow process from unimer insertion/expulsion to micelle fusion/fission.

Facile preparation of well-defined AB2 Y-shaped miktoarm star polypeptide copolymer via the combination of ring-opening polymerization and click chemistry.

Well-defined AB2 Y-shaped miktoarm star polypeptide copolymer, PZLL-b-(PBLG)2, was synthesized via a combination of ring-opening polymerization (ROP) of alpha-amino acid N-carboxyanhydride (NCA) and click chemistry, where PZLL is poly(epsilon-benzyloxycarbonyl-L-lysine) and PBLG is poly(gamma-benzyl-L-glutamate). First, two types of primary-amine-containing initiators, N-aminoethyl 3,5-bis(propargyloxyl)-benzamide and 3-azidopropylamine, were synthesized and employed for the ROP of NCA, leading to the formation of dialkynyl-terminated PZLL and azide-terminated PBLG, dialkynyl-PZLL and PBLG-N3, respectively. The subsequent copper(I)-catalyzed cycloaddition reaction between dialkynyl-PZLL and slightly excess PBLG-N3 led to facile preparation of PZLL-b-(PBLG)2 Y-shaped miktoarm star polypeptide copolymer. The excess PBLG-N3 was scavenged off by reacting with alkynyl-functionalized Wang resin. The obtained Y-shaped miktoarm star polypeptide copolymer was characterized by gel permeation chromatograph (GPC), Fourier transform-infrared spectroscopy (FT-IR), and (1)H NMR. Moreover, after the hydrolysis of protecting benzyl and benzyloxycarbonyl groups of PZLL-b-(PBLG)2, water-soluble pH-responsive Y-shaped miktoarm star polypeptide copolymer, PLL-b-(PLGA)2, was obtained, where PLL is poly(L-lysine) and PLGA is poly(L-glutamic acid). It can self-assemble into PLGA-core micelles at acidic pH and PLL-core micelles at alkaline pH, accompanied with the coil-to-helix transition of PLGA and PLL sequences, respectively. The spontaneous pH-responsive supramolecular assembly of PLL-b-(PLGA)2 miktoarm star polypeptide copolymer has been investigated via a combination of (1)H NMR, laser light scattering (LLS), transmission electron microscopy (TEM), and circular dichroism (CD) spectroscopy.

Probing the micellization kinetics of pyrene end-labeled diblock copolymer via a combination of stopped-flow light-scattering and fluorescence techniques.

A pyrene end-labeled double hydrophilic diblock copolymer, poly(2-(diethylamino)ethyl methacrylate)-b-poly(2-(dimethylamino)ethyl methacrylate) (Py-PDEA-b-PDMA), was synthesized by sequential monomer addition via oxyanionic polymerization using a 1-pyrenemethanol-based initiator. This diblock copolymer exhibits reversible pH-responsive micellization behavior in aqueous solution, forming PDEA-core micelles stabilized by the soluble PDMA block at neutral or alkaline pH. Taking advantage of the pyrene probe covalently attached to the end of the PDEA block, the pH-induced micellization kinetics of Py-PDEA-b-PDMA was monitored by stopped-flow light scattering using a fluorescence detector. Upon a pH jump from 4.0 to 9.0, both the scattered light intensity and excimer/monomer fluorescence intensity ratios (IE/IM) increase abruptly initially, followed by a more gradual increase to reach plateau values. Interestingly, the IE/IM ratio increases abruptly within the first 10 ms: a triple exponential function is needed to fit the corresponding dynamic trace, leading to three characteristic relaxation time constants (tau(1,fluo) < tau(2,fluo) < tau(3,fluo)). On the other hand, dynamic traces for the scattered light intensity can be well-fitted by double exponential functions: the resulting time constants tau(1,scat) and tau(2,scat) can be ascribed to formation of the quasi-equilibrium micelles and relaxation into their final equilibrium state, respectively. Most importantly, tau(1,scat) obtained from stopped-flow light scattering is in general agreement with tau(2,fluo) obtained from stopped-flow fluorescence. The fastest process (tau(1,fluo) approximately 4 ms) detected by stopped-flow fluorescence is ascribed to the burst formation of small transient micelles comprising only a few chains, which are too small to be detected by conventional light scattering. These nascent micelles undergo rapid fusion and grow into quasi-equilibrium micelles and then slowly approach their final equilibrium state. The latter two processes can be detected by both techniques.