Immune checkpoint inhibition mediated with liposomal nanomedicine for cancer therapy.

Immune checkpoint blockade (ICB) therapy for cancer has achieved great success both in clinical results and on the market. At the same time, success drives more attention from scientists to improve it. However, only a small portion of patients are responsive to this therapy, and it comes with a unique spectrum of side effects termed immune-related adverse events (irAEs). The use of nanotechnology could improve ICBs' delivery to the tumor, assist them in penetrating deeper into tumor tissues and alleviate their irAEs. Liposomal nanomedicine has been investigated and used for decades, and is well-recognized as the most successful nano-drug delivery system. The successful combination of ICB with liposomal nanomedicine could help improve the efficacy of ICB therapy. In this review, we highlighted recent studies using liposomal nanomedicine (including new emerging exosomes and their inspired nano-vesicles) in associating ICB therapy.

Role of the stress-responsive two-component system CpxAR in regulating fimbriae expression in Klebsiella pneumoniae CG43.

BACKGROUND: CpxAR is a two-component system that allows bacteria to reorganize envelope structures in response to extracellular stimuli. CpxAR negatively affects type 1 fimbriae expression in Klebsiella pneumoniae CG43, a hypervirulent strain. The involvement of CpxAR in the regulation of type 3 fimbriae expression was investigated. METHODS: cpxAR, cpxA, and cpxR gene-specific deletion mutants were generated. The deletion effects on the expression of type 1 and type 3 fimbriae were analyzed via measuring the promoter activity, mannose sensitive yeast agglutination activity, biofilm formation, and the production of the major pilins FimA and MrkA respectively. RNA sequencing analysis of CG43S3, ΔcpxAR, ΔcpxR and Δfur was employed to study the regulatory mechanism influencing the expression of type 3 fimbriae. RESULTS: Deletion of cpxAR increased type 1 and type 3 fimbrial expression. Comparative transcriptomic analysis showed that the expression of oxidative stress-responsive enzymes, type 1 and type 3 fimbriae, and iron acquisition and homeostasis control systems were differentially affected by cpxAR or cpxR deletion. Subsequent analysis revealed that the small RNA RyhB negatively affects the expression of type 3 fimbriae, while CpxAR positively controls ryhB expression. Finally, the site-directed mutation of the predicted interacting sequences of RyhB with the mRNA of MrkA attenuated the RyhB repression of type 3 fimbriae. CONCLUSION: CpxAR negatively regulates the expression of type 3 fimbriae by modulating cellular iron levels thereafter activating the expression of RyhB. The activated RyhB represses the expression of type 3 fimbriae by base-pairing binding to the 5'region of mrkA mRNA.

Freeze-thaw-stable high internal phase emulsions stabilized by soy protein isolate and chitosan complexes at pH 3.0 as promising mayonnaise replacers.

The development of cholesterol-free mayonnaise has attracted increasing interest in the food colloid field, due to the potential health concerns as a result of consumption of cholesterol-rich mayonnaise. One effective strategy in this regard is to substitute or partially substitute egg yolk with other organic emulsifiers and stabilizers, without affecting the quality of the product. In the work, we reported an effective strategy to fabricate high freeze-thaw-stability high internal phase emulsions (HIPEs), using complexes of a heated soy protein isolate (SPI) and chitosan (CS) at pH 3.0 as the emulsifiers and stabilizers. The SPI/CS complexes, formed even at a very low CS-to-SPI ratio, e.g., 1:10, showed a high capacity to stabilize HIPEs with a high freeze-thaw stability. Increasing the CS-to-SPI ratio in the complexes resulted in a progressive strengthening of gel network in the corresponding HIPEs, together with a gradual improvement of emulsification performance. The gel network of the HIPEs stabilized by the SPI/CS complexes was mainly maintained by the inter-droplet noncovalent interactions involving the CS molecules. The presence of CS also progressively increased the percentage of adsorbed proteins at the interface, and decreased the surface coverage of proteins at the interface. The high freeze-thaw stability of such HIPEs might be unrelated to the ice crystal formation during the freezing, and was more likely associated with the strong steric repulsion contributed to the adsorbed CS molecules between different droplets. The results indicated that the complexation of heated SPI and CS could provide an effective strategy to facilely fabricate outstanding freeze-thaw-stability HI PEs as potential mayonnaise replacers.

The role of urease in the acid stress response and fimbriae expression in Klebsiella pneumoniae CG43.

BACKGROUND/PURPOSE: Two urease operons were identified in Klebsiella pneumoniae CG43, ure-1 and ure-2. This study investigates whether a differential regulation of the expression of ure-1 and ure-2 exists and how urease activity influences the acid stress response and expression of type 1 and type 3 fimbriae. METHODS: The ureA1 and ureA2 gene specific deletion mutants were constructed. Promoter activity was assessed using a LacZ reporter system. The sensitivity to acid stress was determined by assessing the survival after pH 2.5 treatment. The influence on type 1 and type 3 fimbriae expression was assessed using western blotting and mannose-sensitive yeast agglutination and biofilm formation assay, respectively. RESULTS: Bacterial growth analysis in mM9-U or modified Stuart broth revealed that ure-1 was the principal urease system, and ure-2 had a negative effect on ure-1 activity. Deletion of the fur or nac gene had no apparent effect on the activity of Pure1, Pure2-1, and Pure2-2. The Pure2-2 activity was enhanced by deletion of the hns gene. ureA1 deletion increased acid stress sensitivity, whereas the deleting effect of ureA2 was notable without hns. Deletion of ureA1 or ureA2 significantly induced the expression of type 1 fimbriae but decreased MrkA production and biofilm formation. CONCLUSION: ure-1 is the primary expression system in K. pneumoniae CG43, while ure-2 is active in the absence of hns. Impairment of urease activity increases the sensitivity to acid stress, and the accumulation of urea induces the expression of type 1 fimbriae but represses type 3 fimbriae expression.

Berberine Protects Secondary Injury in Mice with Traumatic Brain Injury Through Anti-oxidative and Anti-inflammatory Modulation.

Traumatic brain injury (TBI) is one of the major causes of death and disability worldwide. Novel and effective therapy is needed to prevent the secondary spread of damage beyond the initial injury. The aim of this study was to investigate whether berberine has a neuroprotective effect on secondary injury post-TBI, and to explore its potential mechanism in this protection. The mice were randomly divided into Sham-saline, TBI-saline and TBI-Berberine (50 mg/kg). TBI was induced by Feeney's weight-drop technique. Saline or berberine was administered via oral gavage starting 1 h post-TBI and continuously for 21 days. Motor coordination, spatial learning and memory were assessed using beam-walking test and Morris water maze test, respectively. Brain sections were processed for lesion volume assessment, and expression of neuronal nuclei (NeuN), cyclooxygenase 2 (COX-2), inducible nitric oxide synthase (iNOS), 8-hydroxy-2-deoxyguanosine (8-OHdG), ionized calcium-binding adapter molecule 1 (Iba1) and glial fibrillary acidic protein (GFAP) were detected via immunohistochemistry and immunofluorescence. There were statistically significant improvement in motor coordination, spatial learning and memory in the TBI-Berberine group, compared to the TBI-saline group. Treatment with berberine significantly reduced cortical lesion volume, neuronal loss, COX-2, iNOS and 8-OHdG expression in both the cortical lesion border zone (LBZ) and ipsilateral hippocampal CA1 region (CA1), compared to TBI-saline. Berberine treatment also significantly decreased Iba1- and GFAP-positive cell number in both the cortical LBZ and ipsilateral CA1, relative to saline controls. These results indicated that berberine exerted neuroprotective effects on secondary injury in mice with TBI probably through anti-oxidative and anti-inflammatory properties.

Clinicopathological Features of Idiopathic Membranous Nephropathy in 33 Adolescents.

Objective To investigate the clinicopathological features and prognosis of idiopathic membranous nephropathy(IMN)in adolescents. Methods This was a retrospective study on IMN patients hospitalized between June 2012 and December 2014,and a total of 33 IMN patients aged between 13 and 24 years old were enrolled in the study.Meanwhile,33 IMN patients aged more than 24 years old were selected randomly as control group during the same period.Diagnosis was confirmed by renal biopsy,and the secondary causes of membranous nephropathy were ruled out.Data collected from medical record and biopsy were analyzed. Results In the adolescent IMN group,the mean age at renal biopsy was(20±3)years old,and the male/female ratio was 22/11.Twenty-three cases presented as nephrotic syndrome.Systolic and diastolic pressures were(127±13)mmHg and(77±9)mmHg,respectively.The median 24-hour urine protein was 5.14(3.39,9.31)g/d,and the median serum creatinine was 62(52,73)µmol/L.The positive rate of serum anti-phospholipase A2 receptor in adolescent group was 54%.Compared with control group,the adolescent patients had lower incidence of hypertension and higher baseline estimated glomerular filtration rate level [15.2% vs.39.3%,χ2=4.889,P=0.03;125 ml/(min·1.73m2)vs.100 ml/(min·1.73m2),U=137.5,P<0.001].According to IMN staging criteria in electron microscopy,adolescent patients were classified as one case in stage I,21 in stage Ⅱ,and 11 in stage Ⅲ or higher.The positive rates of IgG1,IgG2,IgG3 and IgG4 subclass staining in glomeruli were 46.9%,3.1%,56.3%,and 87.5%,respectively.Compared with control group,the adolescent patients had lower incidence of renal interstitial fibrosis and arteriolar lesions(6.1% vs.66.7%,χ2=26.19,P<0.001;15.2% vs.66.7%,χ2=18.11,P<0.001).Three patients lost to follow-up while others started steroid combined with cyclosporine A(n=20),cyclophosphamide(n=7),or mycophenolate(n=1)or solely(n=2).After a median follow-up of 18(12,24)months,the median proteinuria decreased to 0.20(0.10,0.42)g/d,whereas serum creatinine level remained stable [69(56.8,81.3)µmol/L].Seventeen patients(56.7%)achieved complete remission(CR),and the remaining 13 patients(43.4%)achieved partial remission(PR).The median time of CR and PR were three and six months,respectively.Only one patient relapsed during the follow-up.Also,21 cases received maintenance therapy including cyclosporine A(n=18),azathioprine(n=2)and mycophenolate(n=1).Conclusions The immunofluorescence IgG subclass in glomeruli and distribution of serum anti-phospholipase A2 receptor in adolescent IMN patients are similar to those in older IMN patients.IMN patients in adolescents responded well to immunosuppressive therapy.Cyclosporine A in low dose as maintenance therapy is effective after achieving remission,and will not increase risk of nephrotoxicity.

The influence of ionic strength on the characteristics of heat-induced soy protein aggregate nanoparticles and the freeze-thaw stability of the resultant Pickering emulsions.

The improvement of the freeze-thaw stability of emulsions by interfacial engineering has attracted increasing attention in recent years. The present work investigated the potential of using soy protein isolate (SPI) aggregate nanoparticles as the Pickering stabilizers to improve the freeze-thaw stability of the resultant emulsions. SPI nanoparticles with different particle sizes and surface properties were fabricated through heating the SPI solutions (at a constant protein concentration of 2%, w/v) at 95 °C for 15 min, by varying the ionic strength (I) in the range of 0-500 mM. The nanoparticles fabricated at I values of 100-500 mM exhibited larger particle sizes and higher surface hydrophobicity, but poorer emulsification efficiency than those at I = 0.05 mM. The presence of NaCl during the nanoparticle fabrication resulted in the formation of a kind of gel-like emulsion with a high extent of droplet flocculation. The emulsion stabilized by SPI nanoparticles at I = 0.05 mM was highly susceptible to coalescence, flocculation and creaming upon freeze-thaw treatment, while those in the presence of NaCl exhibited excellent freeze-thaw stability. The much better freeze-thaw stability of the emulsions in the presence of NaCl (relative to that at I = 0.05 mM) was largely attributed to the gel-like network formation, rather than the salt itself. The results indicated that a kind of Pickering emulsion with excellent freeze-thaw stability, stabilized by heat-induced SPI nanoparticles, could be fabricated by heating the SPI solutions at I values of 100-500 mM. The findings would be of great relevance for providing important information about the development of food grade Pickering emulsions stabilized by protein-based particles, with potential applications in frozen food, or functional food formulations.

Local Mechanical Perturbation Provides an Effective Means to Regulate the Growth and Assembly of Functional Peptide Fibrils.

Mucin 1 (MUC1) peptide fused with Q11 (MUC1-Q11) having 35 residues has previously been shown to form amyloid fibrils. Using time-dependent and high-resolution atomic force microscopy (AFM) imaging, it is revealed that the formation of individual MUC1-Q11 fibrils entails nucleation and extension at both ends. This process can be altered by local mechanical perturbations using AFM probes. This work reports two specific perturbations and outcomes. First, by increasing load while maintaining tip-surface contact, the fibrils are cut during the scan due to shearing. Growth of fibrils occurs at the newly exposed termini, following similar mechanism of the MUC1-Q11 nucleation growth. As a result, branched fibrils are seen on the surface whose orientation and length can be controlled by the nuclei orientation and reaction time. In contrast to the "one-time-cut", fibrils can be continuously fragmented by modulation at sufficiently high amplitude. As a result, short and highly branched fibrils accumulate and pile on surfaces. Since the fibril formation and assembly of MUC1-Q11 can be impacted by local mechanical force, this approach offers a nonchemical and label-free means to control the presentation of MUC1 epitopes, and has promising application in MUC1 fibril-based immunotherapy.

Food protein-based phytosterol nanoparticles: fabrication and characterization.

The development of food-grade (nano)particles as a delivery system for poorly water soluble bioactives has recently attracted increasing attention. This work is an attempt to fabricate food protein-based nanoparticles as delivery systems for improving the water dispersion and bioaccessibility of phytosterols (PS) by an emulsification-evaporation method. The fabricated PS nanoparticles were characterized in terms of particle size, encapsulation efficiency (EE%) and loading amount (LA), and ξ-potential. Among all the test proteins, including soy protein isolate (SPI), whey protein concentrate (WPC) and sodium caseinate (SC), SC was confirmed to be the most suitable protein for the PS nano-formulation. Besides the type of protein, the particle size, EE% and LA of PS in the nanoparticles varied with the applied protein concentration in the aqueous phase and organic volume fraction. The freeze-dried PS nanoparticles with SC exhibited good water re-dispersion behavior and low crystallinity of PS. The LA of PS in the nanoparticles decreased upon storage, especially at high temperatures (e.g., >25 °C). The PS in the fabricated nanoparticles exhibited much better bioaccessibility than free PS. The findings would be of relevance for the fabrication of food-grade colloidal phytosterols, with great potential to be applied in functional food formulations.

Three-Dimensional Nanoprinting via Scanning Probe Lithography-Delivered Layer-by-Layer Deposition.

Three-dimensional (3D) printing has been a very active area of research and development due to its capability to produce 3D objects by design. Miniaturization and improvement of spatial resolution are major challenges in current 3D printing technology development. This work reports advances in miniaturizing 3D printing to the nanometer scale using scanning probe microscopy in conjunction with local material delivery. Using polyelectrolyte polymers and complexes, we have demonstrated the concept of layer-by-layer nanoprinting by design. Nanometer precision is achieved in all three dimensions, as well as in interlayer registry. The approach enables production of designed functional 3D materials with nanometer resolution and, as such, creates a platform for conducting scientific research in designed 3D nanoenvironments as well. In doing so, it enables production of nanomaterials and scaffolds for photonics devices, biomedicine, and tissue engineering.

Structure and dynamics of polymyxin-resistance-associated response regulator PmrA in complex with promoter DNA.

PmrA, an OmpR/PhoB family response regulator, manages genes for antibiotic resistance. Phosphorylation of OmpR/PhoB response regulator induces the formation of a symmetric dimer in the N-terminal receiver domain (REC), promoting two C-terminal DNA-binding domains (DBDs) to recognize promoter DNA to elicit adaptive responses. Recently, determination of the KdpE-DNA complex structure revealed an REC-DBD interface in the upstream protomer that may be necessary for transcription activation. Here, we report the 3.2-Å-resolution crystal structure of the PmrA-DNA complex, which reveals a similar yet different REC-DBD interface. However, NMR studies show that in the DNA-bound state, two domains tumble separately and an REC-DBD interaction is transiently populated in solution. Reporter gene analyses of PmrA variants with altered interface residues suggest that the interface is not crucial for supporting gene expression. We propose that REC-DBD interdomain dynamics and the DBD-DBD interface help PmrA interact with RNA polymerase holoenzyme to activate downstream gene transcription.

Particle Lithography Enables Fabrication of Multicomponent Nanostructures.

Multicomponent nanostructures with individual geometries have attracted much attention because of their potential to carry out multiple functions synergistically. The current work reports a simple method using particle lithography to fabricate multicomponent nanostructures of metals, proteins, and organosiloxane molecules, each with its own geometry. Particle lithography is well-known for its capability to produce arrays of triangular-shaped nanostructures with novel optical properties. This paper extends the capability of particle lithography by combining a particle template in conjunction with surface chemistry to produce multicomponent nanostructures. The advantages and limitations of this approach will also be addressed.

Near-field scanning optical microscopy enables direct observation of Moiré effects at the nanometer scale.

This work reports probing the Moiré effect directly at the nanometer scale via near-field scanning optical microscopy (NSOM). Periodic metal nanostructures of Au and Cu have been produced sequentially using particle lithography, and the overlapped regions serve as Moiré patterns at nanometer scale. The Moiré effect in these regions can be directly visualized from NSOM images, from which periodicity and structural details are accurately determined. In addition, the near-field Moiré effect was found to be very sensitive to structural changes, such as lateral displacement and/or rotations of the two basic arrays with respect to each other. Further, nanostructures of Cu exhibited higher photon transmission than Au from NSOM images. Collectively, NSOM enables direct visualization of the Moiré effect at nanoscale levels, from optical read out, and without enhancements or modification of the structures. The results demonstrate the feasibility to extend applications of the Moiré effect-based techniques to nanometer levels.

Improving Hematite's Solar Water Splitting Efficiency by Incorporating Rare-Earth Upconversion Nanomaterials.

Confounded by global energy needs, much research has been devoted to convert solar energy to various usable forms, such as chemical energy in the form of hydrogen via water splitting. Most photoelectrodes, such as hematite, utilize UV and visible radiation, whereas ∼40% infrared (IR) energy remains unconverted. This work represents our initial attempt to utilize IR radiation, that is, adding rare-earth materials to existing photoelectrodes. A simple substrate composed of hematite film and rare-earth nanocrystals (RENs) was prepared and characterized. Spectroscopy evidence indicates that the RENs in the composite absorb IR radiation (980 nm) and emit at 550 and 670 nm. The emitted photons are absorbed by surrounding hematite films, leading to improvement of water splitting efficiency as measured by photocurrent enhancement. This initial work demonstrates the feasibility and concept of using RENs for utilizing more solar radiation, thus improving the efficiency of existing solar materials and devices.