Oxolinic Acid Generated Green Fluorescence Based on a Terbium-Functionalized Covalent Organic Framework.

Oxolinic acid (OXO), a classic environmental contaminant, has a terrible detrimental effect on human health. The exploration of efficient strategies to detect and detecting OXO has remarkable significance. Herein, we reported a novel terbium(III)-functionalized covalent organic framework (Bpy-DhBt-COF@Tb3+) by fixing Tb3+ on the bipyridine-connecting COF (Bpy-DhBt-COF) as a turn-on fluorescent switch toward OXO for the first time. In this platform, Tb3+ acts as the specific recognition units for OXO and the response signal, while Bpy-DhBt-COF acts as the safehaven for Tb3+. Once introducing OXO to Bpy-DhBt-COF@Tb3+, OXO can instead water molecules coordinate with Tb3+ and sensitize Tb3+ instantly, thereby producing a significant fluorescence signal. Profiting from the excellent porosity of Bpy-DhBt-COF@Tb3+, it can obtain optimal response toward OXO only within 10 s with an ultrasensitive detection limit of 12.5 nM. Furthermore, Bpy-DhBt-COF@Tb3+ displayed outstanding selectivity toward OXO than other general quinolones. Based on these, a Tb3+-based COF was explored for the first time for the turn-on fluorescence detection of an OXO with rapid response, high sensitivity, and outstanding selectivity. In this work, we not only exhibit the attractive performance of Tb3+-functionalized COF to detect OXO but also propose a prospect strategy for creating other fluorescent sensors for multiple targets.

Cellulose-Based Cryogel Microspheres with Nanoporous and Controllable Wrinkled Morphologies for Rapid Hemostasis.

First-aid hemostatic agents for acute bleeding can save lives in emergency situations. However, rapid hemostasis remains challenging when uncontrolled hemorrhage occurs on lethal noncompressible and irregular wounds. Herein, cellulose-based cryogel microspheres with deliberately customized micromorphologies for ultrafast water transportation and diffusion, including the shark skin riblet-inspired wrinkled surface with low fluid drag and the hydrophilic nanoporous 3D networks, are developed to deal with the acute noncompressible bleeding within seconds. These cryogel microspheres can rapidly absorb a large amount of blood over 6 times their own weight in 10 s and form a robust barrier to seal a bleeding wound without applying pressure. Remarkably, massive bleeding from a cardiac penetrating hole is effectively stopped using the microspheres within 20 s and no blood leakage is observed after 30 min. Additionally, these microspheres could be readily removed without rebleeding and capillary thrombus, which is highly favorable to rapid hemostasis in emergency rescue.

Glycyrrhizin micelle as a genistein nanocarrier: Synergistically promoting corneal epithelial wound healing through blockage of the HMGB1 signaling pathway in diabetic mice.

The purpose of this study was to explore the feasibility of targeting the HMGB1 signaling pathway to treat diabetic keratopathy with a dipotassium glycyrrhizinate-based micelle ophthalmic solution encapsulating genistein (DG-Gen), and to evaluate whether these dipotassium glycyrrhizinate (DG) micelles could synergistically enhance the therapeutic effect of encapsulated genistein (Gen). An optimized DG-Gen ophthalmic solution was fabricated with a Gen/DG weight of ratio 1:15, and this formulation featured an encapsulation efficiency of 98.96 ± 0.82%, and an average particle size of 29.50 ± 2.05 nm. The DG-Gen ophthalmic solution was observed to have good in vivo ocular tolerance and excellent in vivo corneal permeation, and to remarkably improve in vitro antioxidant activity. Ocular topical application of the DG-Gen ophthalmic solution significantly prompted corneal re-epithelialization and nerve regeneration in diabetic mice, and this efficacy might be due to the inhibition of HMGB1 signaling through down-regulation of HMGB1 and its receptors RAGE and TLR4, as well as inflammatory factor interleukin (IL)-6 and IL-1ß. In conclusion, these data showed that HMGB1 signaling is a potential regulation target for the treatment of diabetic keratopathy, and novel DG-micelle formulation encapsulating active agents such as Gen could synergistically cause blockage of HMGB1 signaling to prompt diabetic corneal and nerve wound healing.

Expression profiles and potential corneal epithelial wound healing regulation targets of high-mobility group box 1 in diabetic mice.

As a damage-associated molecular pattern molecule, high-mobility group box 1 protein (HMGB1) is involved in diabetes and its complications. However, the role of HMGB1 in diabetic keratopathy is not yet understood. The purpose of this study was to investigate the potential roles of HMGB1 in the development of diabetic keratopathy as well as potential strategies to block HMGB1 in order to prompt epithelial wound healing and nerve regeneration in diabetic corneas. The results demonstrated that diabetic keratopathy developed in mice over the duration of the diabetic condition with typical symptoms, including damaged ocular surfaces and corneal nerves. The diabetic corneas had significantly increased protein expression levels of HMGB1 and its receptors-the receptor for advanced glycation end products (RAGE) and toll-like receptor 4 (TLR4)-compared to the age-matched normal corneas (P < 0.05). Corneal HMGB1 levels significantly increased during the corneal wound healing process of the diabetic mice, peaking on the first day after the wound was created and then decreasing to the unwounded level on the seventh day. Exogenous HMGB1 peptide significantly retarded wound and nerve healing, while glycyrrhizin (an HMGB1 inhibitor) significantly prompted wound and nerve healing. Further, the western blot results confirmed that RAGE and TLR4 were also involved in corneal wound and nerve healing. In conclusion, these data showed that HMGB1 and its related receptors are highly involved in the development of diabetic keratopathy. This finding indicates that the blockage of HMGB1 might serve as a strategy to prompt diabetic corneal and nerve wound healing.

Characterization of Antigenic Relatedness Among GI Norovirus Genotypes Using Serum Samples From Norovirus-Infected Patients and Mouse Sera.

Characterizing diversity and the antigenic relatedness of norovirus remains a primary focus in understanding its biological properties and vaccine designs. The precise antigenic and serological features of GI genotypes have not been studied. The study represented an investigation on a gastroenteritis outbreak related to GI.3 norovirus and the three most detected GI genotypes, GI.2 (belonging to immunotype B), GI.3 and GI.9 (belonging to immunotype C), were selected to characterize their phylogenetic relationship, HBGA binding profiles and antigenic relatedness within (intra-immunotype), and between (inter-immunotypes) genotypes using mouse sera and patient's serum samples from the GI.3 related outbreak. Wide HBGA binding profiles and evolution of binding affinity were observed in the three GI genotypes studied. A low specific blockade antibody to GI.3 in the population generated the pool of susceptible individuals and supported virus spread in the outbreak. We found strong blockade immune response in homologous strains, moderate intra-immunotype blockade but weak inter-immunotypes blockade in humans following GI.3 norovirus infections. These findings further support the immunotypes grouping and will be valuable for optimizing the design of norovirus vaccine.

[Preparation and characterization of monoclonal antibodies against GII.4 norovirus P domain].

Objective To prepare monoclonal antibodies (mAbs) against GII.4 norovirus P domain by multiple antigens in an immunization program. Methods BALB/c mice were immunized with the multiple GII.4 NoV P domain, namely 1996cluster (VA387), 2004cluster, 2006b cluster and 2010 cluster. The spleen cells from the immunized mice were fused with SP2/0 cells and the hybridoma cells were screened by ELISA. The supernatant of the mAbs was collected and purified by the limiting dilution assay. Its subtype was identified, and the specificity and neutralization were analyzed by indirect ELISA and HBGA blocking, respectively. Results We obtained thirteen hybridoma cell lines that stably secreted mAbs against GII.4 NoV P domain. Their titers reached above 10-4 after purification. The subtypes of the mAbs were identified as IgG1. Indirect ELISA showed that all the mAbs specifically bound to all GII.4 norovirus variants. Five mAbs specifically bound to GII.17, GII.3 and GII.6 variants. Three mAbs specifically bound to GII.2 variants and strongly blocked NoV P particle from binding to the histo-blood group antigen (HBGA) receptors. Conclusion The mAbs against GII.4 norovirus P domain have been obtained by combined antigens immunization program. Multi-antigen immunization can enhance immune response significantly and cross-react with other GII.4 norovirus variants. The findings provide a basis for further development of novel GII.4 norovirus vaccines and for the optimization of the immunization programs of combined multi-antigen vaccine candidates.

Novel self-nanomicellizing formulation based on Rebaudioside A: A potential nanoplatform for oral delivery of naringenin.

In the study described here, we strove to develop an orally administered novel self-nanomicellizing formulation based on Rebaudioside A (RA) for delivering naringenin (NAR) with improved bioavailability and therapeutic efficacy. Our research found that RA and naringenin (NAR) could be formulated into self-assembling nanomicelles (RA-NAR) using a simple ethanol dissolution-evaporation method. We found that the RA-NAR self-assemblies comprised ultra-small micelles (5.234 ± 0.311 nm) in a uniform dispersion state (the polydispersity index was 0.243 ± 0.039) with a near-neutral surface charge (-[2.268 ± 0.729] mV). We also found that RA-NAR had a well-storage stability at 4 °C with light protection. In addition, we observed that RA-NAR exhibited enhanced apparent solubility, in-vitro permeability, and antioxidant activity. After we administered RA-NAR to rats orally, we observed an increase in area under the curve (AUC0→t) to 19,500.82 ng/mL/h versus 9324.47 ng/mL/h observed with free NAR and an increase of maximum concentration (Cmax) to 27,326.10 ng/mL from the free-NAR Cmax level of 2549.04 ng/mL. The tissue distribution assessments further demonstrated that RA-NAR could effectively increase the NAR concentration in all tested intestinal segments. Our mouse model results showed as well that oral administration of RA-NAR could efficiently protect against small intestine injuries induced by indomethacin, and the mechanisms by inhibiting proinflammatory cytokines and oxidative stress were involved in its therapeutic effect. Taken together, these findings indicate that a self-nanomicellizing formulation based on RA has great potential as a novel oral nano-drug delivery system for NAR.

Epidemiology and HBGA-susceptibility investigation of a G9P[8] rotavirus outbreak in a school in Lechang, China.

Rotaviruses cause severe gastroenteritis in infants, in which the viruses interact with human histo-blood group antigens (HBGAs) as attachment and host susceptibility factors. While gastroenteritis outbreaks caused by rotaviruses are uncommon in adolescents, we reported here one that occurred in a middle school in China. Rectal swabs and saliva samples were collected from symptomatic and asymptomatic students, and samples were also collected from the environment. Using PCR, followed by DNA sequencing, a single G9P[8] rotavirus strain was identified as the causative agent. The attack rate of the outbreak was 13.5% for boarders, which was significantly higher than that of day students (1.8%). Person-to-person transmission was the most plausible transmission mode. The HBGA phenotypes of the individuals in the study were determined by enzyme immunoassay, using saliva samples, while recombinant VP8* protein of the causative rotavirus strain was produced for HBGA binding assays to evaluate the host susceptibility. Our data showed that secretor individuals had a significantly higher risk of infection than nonsecretors. Accordingly, the VP8* protein bound nearly all secretor saliva samples, but not those of nonsecretors, explaining the observed infection of secretor individuals only. This is the first single-outbreak-based investigation showing that P[8] rotavirus infected only secretors. Our investigation also suggests that health education of school students is an important countermeasure against an outbreak of communicable disease.

The ERK-MNK-eIF4F signaling pathway mediates TPDHT-induced A549 cell death in vitro and in vivo.

The eIF4E/eIF4G complex plays a central role in gene expression regulation during the initial stage of translation. This study aimed to determine if the novel small molecule compound, TPDHT, could disrupt the interaction of eIF4E/eIF4G, and if it could exhibit excellent antitumor activity in vivo without causing apparent toxicity effect. This study investigated the antitumor mechanism of TPDHT in vitro and in vivo. TPDHT showed significant anti-proliferative activity on human lung cancer A549 cells, and it induced G0/G1 cycle arrest. Moreover, TPDHT also induced A549 cell apoptosis through the mitochondria-mediated apoptotic pathway. Our results indicate that TPDHT could disrupt the interaction of eIF4E/eIF4G, and the activity of eIF4F plays an important role in TPDHT-induced cell proliferation inhibition and apoptosis. Further research showed that TPDHT could inhibit the Ras/ERK/MNK pathway, and activation of the ERK pathway reversed TPDHT-induced cell proliferation inhibition and apoptosis. Taken together, our study findings indicated that TPDHT exerts an antitumor effect through an intrinsic apoptotic pathway controlled by the ERK/MNK/eIF4F pathway.

Commercial E2 subunit vaccine provides full protection to pigs against lethal challenge with 4 strains of classical swine fever virus genotype 2.

Classical swine fever (CSF) still threatens the swine industry in China, with genotype 2 isolates of CSFV dominating the epizootics. In 2018 the first E2 subunit marker vaccine against CSFV (Tian Wen Jing, TWJ-E2®), containing a baculovirus-expressed E2 glycoprotein of a genotype 1.1 vaccine strain, was officially licensed in China and commercialized. To evaluate the cross-protective efficacy of TWJ-E2 against different virulent genotype 2 Chinese field isolates (2.1b, 2.1c, 2.1 h, and 2.2), 4-week-old pigs were immunized with the TWJ-E2 vaccine according to the manufacturer's instructions and then challenged with genotype 2 strains. A group vaccinated with the conventional C-strain vaccine was included for comparison. TWJ-E2 vaccinated pigs developed higher levels of E2 and neutralizing antibodies than those receiving the commercial C-strain vaccine. All TWJ-E2 and C-strain vaccinated pigs survived challenge without development of fever, clinical signs or pathological lesions. In contrast, all unvaccinated control pigs displayed severe CSF disease with 40-100% mortalities by 24 days post challenge. None of the TWJ-E2 and C-strain vaccinated pigs developed viremia, viral shedding from tonsils, Erns protein in the sera, or viral RNA loads in different tissues after challenge, all of which were detected in the challenged unvaccinated controls. We conclude that vaccination of young pigs with TWJ-E2 provides complete immune protection against genotypically heterologous CSFVs and prevents viral shedding after challenge, with an efficacy at least comparable to that elicited by the conventional C-strain vaccine.

Ultra-small micelles based on polyoxyl 15 hydroxystearate for ocular delivery of myricetin: optimization, in vitro, and in vivo evaluation.

The aim was to develop a nanocarrier based on polyoxyl 15 hydroxystearate (Kolliphor® HS15, HS15) micelles for the solubility, stability, and ocular delivery of myricetin (Myr). An optimized ratio of HS15 and Myr was prepared to fabricate HS15-Myr micelle ophthalmic solution. Myr-encapsulating HS15 micelles (HS15-Myr micelles) were subjected to physicochemical characterizations. The chemical stability of Myr in HS15 micelles and storage stability of HS15-Myr micelle ophthalmic solutions were evaluated. In vitro parallel artificial membrane permeability assay and antioxidant activity of Myr in HS15 micelles were also measured. In vivo ocular tolerance, corneal permeation, and anti-inflammatory efficacy studies were conducted following ocular topical administration. HS15-Myr micelles were successfully prepared and presented transparent appearance with high encapsulation (96.12 ± 0.31%), ultra-small micelle size (a mean diameter of 12.17 ± 0.73 nm), uniform size distribution (polydispersity index [PDI] = 0.137 ± 0.013), and negative surface charge (- [4.28 ± 0.42] mV). Myr in HS15 micelle solution demonstrated higher aqueous stability than the free Myr solution among the accepted pH range for eyedrops. HS15-Myr micelle ophthalmic solution demonstrated high storage stability at 4 °C and 25 °C. HS15 micelles could significantly improve in vitro antioxidant activity and faster membrane permeation of Myr. No irritations or corneal damage were revealed in rabbit eyes after ocular administration of HS15-Myr micelle solution. In vivo corneal permeation study demonstrated that HS15-Myr micelles could penetrate the cornea efficiently in mouse eyes. Further, HS15-Myr micelles also demonstrated significant in vivo anti-inflammatory activity. It can be concluded that HS15 micelles are a potential ophthalmic delivery nanocarrier for poorly soluble drugs such as Myr.

Novel self-nanomicellizing solid dispersion based on rebaudioside A: a potential nanoplatform for oral delivery of curcumin.

PURPOSE: Rebaudioside A (RA) has nanocarrier characteristics that allow it to self-assemble into micelles in aqueous solutions. The purpose of this study was to determine if a self-nanomicellizing solid dispersion based on RA could be utilized as an oral nano-drug delivery system. MATERIALS AND METHODS: Curcumin (Cur) served as a model hydrophobic drug, and a Cur-loaded self-nanomicellizing solid dispersion based on RA (RA-Cur) was formulated. The properties of RA-Cur in the solid state and in aqueous solution were characterized. The antioxidant activity and mechanism of RA-Cur endocytosis were also investigated. The pharmacokinetics, biodistribution in the intestinal tract, and anti-inflammation properties were also evaluated in vivo. RESULTS: RA-Cur could be easily fabricated, and it self-assembled into ultrasmall micelles (particle size ~4 nm) in a homogeneous distribution state (polydispersity index <0.2) when dissolved in water. Cur was readily encapsulated into RA micelles and this improved its water solubility (to 14.34±1.66 mg/mL), as well as its in vitro release and membrane permeability. The antioxidant activities of Cur in RA-Cur were also significantly improved. Biodistribution in the intestinal tract confirmed a significant enhancement of Cur absorption in the duodenum, jejunum, and ileum by encapsulation in RA-Cur, and the absorption of RA-Cur was governed by mixed transcytosis mechanisms. Pharmacokinetic tests of RA-Cur in rats revealed a dramatic 19.06-fold enhancement of oral bioavailability when compared to free Cur. More importantly, oral administration of RA-Cur could efficiently ameliorate ulcerative colitis in a mouse model induced by dextran sodium sulfate. CONCLUSION: Self-nanomicellizing solid dispersions based on RA have great potential as novel oral nano-drug delivery systems for hydrophobic drugs such as Cur.

Evaluation of polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer nanomicelle for trigeminal ganglion neurons delivering with intranasal administration.

Purpose How to deliver enough medical agents to the trigeminal ganglion (TG) neurons conveniently still remains a challenge in pharmaceutics and clinics. The purpose of this study was to reveal that intranasal administration of polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer (PVCL-PVA-PEG) nanomicelle formulation could efficiently deliver agent to TG neurons in mice. Methods Ocular topical or intranasal administration of nanomicelle coumarin-6 was performed in mice, and tissue distribution after administration (0.25, 1, 2, 4, 6, 8, and 10 h) was analyzed. Fluoro-Gold was used as a retrograde tracer to identify corneal and nasal neurons in the TG. Pharmacokinetic profiles after ocular topical or intranasal administration were explored in detail. Results Coumarin-6 levels in the TG neurons were significantly higher in intranasal administration groups than in topical administration groups, and the difference was statistically significant (P < 0.05) at all time points except for 10 h. Interestingly, in cornea, coumarin-6 was detected after intranasal administration. For intranasal administration groups, it was also interestingly found that coumarin-6 levels in the TG neurons were much higher than that in the brain, suggesting that the TG neurons was a target tissue after the intranasal administration of nanomicelle coumarin-6. These levels also indicated the safety of brain tissue after intranasal administration. Using Fluoro-Gold tract tracing techniques, coumarin-6 was detected in TG neurons after either ocular topical or intranasal administration of nanomicelle coumarin-6, indicating the high colocalization of corneal and nasal neurons in the TG. Conclusions Intranasal administration of PVCL-PVA-PEG nanomicelle formulation could efficiently deliver to TG neurons, and it might be a promising therapy for pathological TG neurons.

Anti-complementary constituents of Anchusa italica.

Activity-guided fractionation for complement inhibitors led to the isolation of 24 known compounds from Anchusa italica. Chemical types include eight megastigmane compounds, five triterpenoid compounds, five lignan compounds, three flavonoid compounds, two alkaloid compounds and one phenthyl alcohol compound. Among which, a lignan (medioresinol), an alkaloid (5-hydroxypyrrolidin-2-one) and a flavonoid (5-hydroxyl-3', 4', 6, 7-tetramethoxy flavone) exhibited better anticomplementary effects with CH50 values ranging from 0.07 to 0.82 mM, which are plausible candidates for developing potent anticomplementary agents.

[Transcriptome Analysis of Rabbit Spleen with Hog Cholera Lapinized Virus Infection Based on High-throughput Sequencing Technology].

In the current study, rabbit spleen was analysed at 12 h,24h,30 h,36h,and 48hpost-infection with hog cholera lapinized virus(C strain)using high-throughput sequencing. The rabbit genome was used as a reference to identify differentially expressed genes(DEGs)at different time points post-infection. The top 10 DEGs were filtered based on significance, and we searched for their biological functions through the Uniprot and NCBI databases. The former three time points have 10co-expressing genes, many of which have a relationship to immunity and inflammation. The latter two time points for the top 10 DEGs are identical, and B2 M,RLA-DR-ALPHA,CD74,and IGJ are involved in the antiviral immune response. GO functional annotation revealed that in biological processes at each time point,except for 24hpost-infection,the immune response has the most terms, followed by metabolism and regulation. According to the KEGG database, the DEGs for 24hpost-infection were enriched for the RIG-I-like receptor signaling pathway and the DEGs for 30hpost-infection were found to have a focal adhesion and ECM-receptor interaction pathway. Moreover, the DEGs for 36 hand 48hpost-infection have seven identical pathways, of which were directly or indirectly related to the antiviral response. These pathways included the proteasome, lysosome, ribosome, chemokine signaling pathways, B cell receptor signaling pathway, antigen processing, and presentation pathway, and the Fc gamma R-mediated phagocytosis pathway.These results provide novel insight into the gene expression in rabbit spleens post-infection with C strain, and provide a theoretical basis for further understanding of the molecular mechanisms by which rabbits adapt to infection with C strain.

[Research Progress in the Core Proteins of the Classical Swine Fever Virus].

The core protein (CP) of the classical swine fever virus (CSFV) is one of its structural proteins. Apart from forming the nucleocapsid to protect internal viral genomic RNA, this protein is involved in transcriptional regulation. Also, during viral infection, the CP is involved in interactions with many host proteins. In this review, we combine study of this protein with its disorders, structural/functional characteristics, as well as its interactions with the non-structural proteins NS3, NS5B and host proteins such as SUMO-1, UBC9, OS9 and IQGAP1. We also summarize the important part played by the CP in CSFV pathogenicity, virulence and replication of genomic RNA. We also provide guidelines for further studies in the CP of the CSFV.