Drug-loaded mucoadhesive microneedle patch for the treatment of oral submucous fibrosis

Oral submucous fibrosis is a chronic, inflammatory and potentially malignant oral disease. Local delivery of triamcinolone to lesion site is a commonly used therapy. The existing methods for local drug delivery include topical administration and submucosal injection. However, in the wet and dynamic oral microenvironment, these methods have drawbacks such as limited drug delivery efficiency and injection pain. Therefore, it is urgently needed to develop an alternative local drug delivery system with high efficiency and painlessness. Inspired by the structure of band-aid, this study proposed a novel double-layered mucoadhesive microneedle patch for transmucosal drug delivery. The patch consisted of a mucoadhesive silk fibroin/tannic acid top-layer and a silk fibroin microneedle under-layer. When applying the annealing condition for the medium content of β-sheets of silk fibroin, the microneedles in under-layer displayed both superior morphology and mechanical property. The mechanical strength of per needle (0.071N) was sufficient to penetrate the oral mucosa. Sequentially, the gelation efficiency of silk fibroin and tannic acid in top-layer was maximized as the weight ratio of tannic acid to silk fibroin reached 5:1. Moreover, in vitro results demonstrated the double-layered patch possessed undetectable cytotoxicity. The sustained release of triamcinolone was observed from the double-layered patch for at least 7 days. Furthermore, compared with other commercial buccal patches, the double-layered patch exhibited an enhanced wet adhesion strength of 37.74 kPa. In addition, ex vivo mucosal tissue penetration experiment confirmed that the double-layered patch could reach the lamina propria, ensuring effective drug delivery to the lesion site of oral submucous fibrosis. These results illustrate the promising potential of the drug-loaded mucoadhesive microneedle patch for the treatment of oral submucous fibrosis

Multi-Stage Tuberculosis Subunit Vaccine Candidate LT69 Provides High Protection against Mycobacterium tuberculosis Infection in Mice

Effective tuberculosis (TB) vaccine should target tubercle bacilli with various metabolic states and confer long-term protective immunity. In this study, we constructed a novel multi-stage TB subunit vaccine based on fusion protein ESAT6-Ag85B-MPT64(190-198)-Mtb8.4-HspX (LT69 for short) which combined early expressed antigens and latency-associated antigen. The fusion protein was mixed with an adjuvant being composed of N, N’-dimethyl-N, N’-dioctadecylammonium bromide (DDA) and polyriboinosinic polyribocytidylic acid (PolyI:C) to construct subunit vaccine, whose immunogenicity and protective ability were evaluated in C57BL/6 mice. The results showed that LT69 had strong immunogenicity and high protective effect against Mycobacterium tuberculosis (M. tuberculosis) H37Rv aerosol challenge. Low-dose (2 μg) of LT69 generated long-term immune memory responses and provided effective protection, which was even higher than traditional vaccine BCG did at 30 weeks post the last vaccination. In conclusion, multistage subunit vaccine LT69 showed high and long-term protection against M. tuberculosis infection in mice, whose effect could be enhanced by using a relative low dosage of antigen.National Major Science and Technology Projects (China) (2012ZX10003-008-006)National Natural Science Foundation (China) (31470895)National Natural Science Foundation (China) (81072499)China. Ministry of Education (Doctoral Fund 20120211110038

Dispersive Liquid-Liquid Microextraction Combined with Ultrahigh Performance Liquid Chromatography/Tandem Mass Spectrometry for Determination of Organophosphate Esters in Aqueous Samples

A new technique was established to identify eight organophosphate esters (OPEs) in this work. It utilised dispersive liquid-liquid microextraction in combination with ultrahigh performance liquid chromatography/tandem mass spectrometry. The type and volume of extraction solvents, dispersion agent, and amount of NaCl were optimized. The target analytes were detected in the range of 1.0–200 µg/L with correlation coefficients ranging from 0.9982 to 0.9998, and the detection limits of the analytes were ranged from 0.02 to 0.07 µg/L (S/N=3). The feasibility of this method was demonstrated by identifying OPEs in aqueous samples that exhibited spiked recoveries, which ranged between 48.7% and 58.3% for triethyl phosphate (TEP) as well as between 85.9% and 113% for the other OPEs. The precision was ranged from 3.2% to 9.3% (n=6), and the interprecision was ranged from 2.6% to 12.3% (n=5). Only 2 of the 12 selected samples were tested to be positive for OPEs, and the total concentrations of OPEs in them were 1.1 and 1.6 µg/L, respectively. This method was confirmed to be simple, fast, and accurate for identifying OPEs in aqueous samples

E6 Protein Expressed by High-Risk HPV Activates Super-Enhancers of the EGFR and c-MET Oncogenes by Destabilizing the Histone

The high-risk (HR) human papillomaviruses (HPV) are causative agents of anogenital tract dysplasia and cancers and a fraction of head and neck cancers. The HR HPV E6 oncoprotein possesses canonical oncogenic functions, such as p53 degradation and telomerase activation. It is also capable of stimulating expression of several oncogenes, but the molecular mechanism underlying these events is poorly understood. Here, we provide evidence that HPV16 E6 physically interacts with histone H3K4 demethylase KDM5C, resulting in its degradation in an E3 ligase E6AP- and proteasome-dependent manner. Moreover, we found that HPV16-positive cancer cell lines exhibited lower KDM5C protein levels than HPV-negative cancer cell lines. Restoration of KDM5C significantly suppressed the tumorigenicity of CaSki cells, an HPV16-positive cervical cancer cell line. Whole genome ChIP-seq and RNA-seq results revealed that CaSki cells contained super-enhancers in the proto-oncogenes EGFR and c-MET. Ectopic KDM5C dampened these super-enhancers and reduced the expression of proto-oncogenes. This effect was likely mediated by modulating H3K4me3/H3K4me1 dynamics and decreasing bidirectional enhancer RNA transcription. Depletion of KDM5C or HPV16 E6 expression activated these two super-enhancers. These results illuminate a pivotal relationship between the oncogenic E6 proteins expressed by HR HPV isotypes and epigenetic activation of super-enhancers in the genome that drive expression of key oncogenes like EGFR and c-MET. Significance: This study suggests a novel explanation for why infections with certain HPV isotypes are associated with elevated cancer risk by identifying an epigenetic mechanism through which E6 proteins expressed by those isotypes can drive expression of key oncogenes.</p

Description of a new species of the genus Ameletus Eaton, 1885 (Ephemeroptera, Ameletidae) from Yunnan, China

A new species with primitive characteristics, Ameletus daliensis Tong, sp. nov., is described, based on the morphology of imago, larva and egg with molecular data of the mitochondrial COI from Mount Cangshan, Dali, China. The new species is closely related to one of the most primitive mayflies, Ameletus primitivus Traver, 1939, by sharing persistent mouthparts in the alate stage, but it can be distinguished from the latter by the morphological differences of the mouthpart remains, wings and genitals in the imaginal stage. Both morphological and molecular evidence support that A. daliensis Tong, sp. nov. is a new member of the genus Ameletus. The discovery of the new species could help understand the origin and evolution of the genus Ameletus

In Situ Construction of CNT/CuS Hybrids and Their Application in Photodegradation for Removing Organic Dyes

Herein, a coprecipitation method used to synthesize CuS nanostructures is reported. By varying the reaction time and temperature, the evolution of the CuS morphology between nanoparticles and nanoflakes was investigated. It was found that CuS easily crystallizes into sphere-/ellipsoid-like nanoparticles within a short reaction time (0.5 h) or at a high reaction temperature (120 &deg;C), whereas CuS nanoflakes are readily formed at a low reaction temperature (20 &deg;C) for a long time (12 h). Photodegradation experiments demonstrate that CuS nanoflakes exhibit a higher photodegradation performance than CuS nanoparticles for removing rhodamine B (RhB) from aqueous solution under simulated sunlight irradiation. Carbon nanotubes (CNTs) were further used to modify the photodegradation performance of a CuS photocatalyst. To achieve this aim, CNTs and CuS were integrated to form CNT/CuS hybrid composites via an in situ coprecipitation method. In the in situ constructed CNT/CuS composites, CuS is preferably formed as nanoparticles, but cannot be crystallized into nanoflakes. Compared to bare CuS, the CNT/CuS composites manifest an obviously enhanced photodegradation of RhB; notably, the 3% CNT/CuS composite with CNT content of 3% showed the highest photodegradation performance (&eta; = 89.4% for 120 min reaction, kapp = 0.01782 min&minus;1). To make a comparison, CuS nanoflakes and CNTs were mechanically mixed in absolute alcohol and then dried to obtain the 3% CNT/CuS-MD composite. It was observed that the 3% CNT/CuS-MD composite exhibited a slightly higher photodegradation performance (&eta; = 92.4%, kapp = 0.0208 min&minus;1) than the 3% CNT/CuS composite, which may be attributed to the fact that CuS maintains the morphology of nanoflakes in the 3% CNT/CuS-MD composite. The underlying enhanced photocatalytic mechanism of the CNT/CuS composites was systematically investigated and discussed

Potassium fertilization arrests malate accumulation and alters soluble sugar metabolism in apple fruit

Effects of different potassium (K) levels, which were K0 (no fertilizer), K1 (71.5 g KCl plant−1 year−1), K2 (286.7 g KCl plant−1 year−1), and K3 (434 g KCl plant−1 year−1), were evaluated based on sugar and organic acid metabolism levels from 70–126 days after bloom (DAB) in the developing fruit of potted five-year-old apple (Malus domestica, Borkh.) trees. The results indicate that K fertilization promoted greater fruit mass, higher Ca2+ and soluble solid levels, and lower titratable acid levels, as well as increased pH values at harvest. With the application of different levels of K fertilizer, fructose, sorbitol, glucose and sucrose accumulation rates significantly changed during fruit development. Fruit in the K2 group had higher fructose, sucrose and glucose levels than those in other treatment groups at 126 DAB. These changes in soluble sugar are related to the activity of metabolic enzymes. Sucrose synthase (SS) and sorbitol dehydrogenase (SDH) activity in the K2 treated fruit was significantly higher than those in other treatment groups from 70–126 DAB. Malate levels in K-supplemented fruit were notably lower than those in non K-supplemented fruit, and K3 treated fruit had the lowest malate levels during fruit development. Cytosolic malic enzyme (ME) and phosphoenolpyruvate carboxykinase (PEPCK) activity significantly increased in fruit under the K2 treatment during 112–126 DAB and 98–126 DAB, respectively. In addition, Ca2+ concentration increased with increasing K fertilization levels, which promoted a maximum of 11.72 mg g−1 dry weight in apple fruit. These results show that K levels can alter soluble sugar and malate levels due to the interaction between sugars and acid-metabolic enzymes in fruit

Identification and characterization of a novel trehalose synthase gene derived from saline-alkali soil metagenomes.

A novel trehalose synthase (TreS) gene was identified from a metagenomic library of saline-alkali soil by a simple activity-based screening system. Sequence analysis revealed that TreS encodes a protein of 552 amino acids, with a deduced molecular weight of 63.3 kDa. After being overexpressed in Escherichia coli and purified, the enzymatic properties of TreS were investigated. The recombinant TreS displayed its optimal activity at pH 9.0 and 45 °C, and the addition of most common metal ions (1 or 30 mM) had no inhibition effect on the enzymatic activity evidently, except for the divalent metal ions Zn(2+) and Hg(2+). Kinetic analysis showed that the recombinant TreS had a 4.1-fold higher catalytic efficiency (Kcat/K m ) for maltose than for trehalose. The maximum conversion rate of maltose into trehalose by the TreS was reached more than 78% at a relatively high maltose concentration (30%), making it a good candidate in the large-scale production of trehalsoe after further study. In addition, five amino acid residues, His172, Asp201, Glu251, His318 and Asp319, were shown to be conserved in the TreS, which were also important for glycosyl hydrolase family 13 enzyme catalysis

Effects of Iron Deficiency Stress on Plant Growth and Quality in Flowering Chinese Cabbage and Its Adaptive Response

Iron (Fe) plays an important role in the growth and development of plants. The effects of different Fe concentrations, 1-aminocyclopropane-1-carboxylic acid (ACC), and cobalt chloride (Co2+) treatments on plant growth, quality and the adaptive response to Fe deficiency stress were investigated in flowering Chinese cabbage. The results revealed that Fe deficiency stress inhibited plant growth. The contents of vitamin C, soluble protein, and soluble sugar in leaves and stalks were significantly reduced under Fe deficiency stress, while the content of cellulose and nitrate was increased. Fe deficiency stress clearly reduced the net photosynthetic rate and nitrate reductase activity in the leaves. The balance system of active oxygen metabolism was destroyed due to Fe deficiency, resulting in the decrease in catalase activity, superoxide dismutase activity of roots and leaves, and peroxidase (POD) activity of leaves, while POD activity in roots and malonaldehyde content in roots and leaves were significantly increased. The treatments of Fe deficiency and ACC significantly reduced the pH value of the root medium, promoted the release of ethylene, and increased Fe3+ reductase activity, while Co2+ treatment showed results that were the opposite to those of Fe deficiency and ACC treatments. Thus, Fe deficiency stress affected nitrogen metabolism, photosynthesis, reactive oxygen metabolism, pH of root medium, and Fe3+ reductase activity, which was related to physiological adaptive response and tolerance mechanisms. We also found that ethylene could be involved in regulating the adaptive response to Fe deficiency stress in flowering Chinese cabbage