Functional diversity of subgroup 5 R2R3-MYBs promoting proanthocyanidin biosynthesis and their key residues and motifs in tea plant.

The tea plant (Camellia sinensis) is rich in polyphenolic compounds. Particularly, flavan-3-ols and proanthocyanidins (PAs) are essential for the flavor and disease-resistance property of tea leaves. The fifth subgroup of R2R3-MYB transcription factors comprises the primary activators of PA biosynthesis. This study showed that subgroup 5 R2R3-MYBs in tea plants contained at least nine genes belonging to the TT2, MYB5, and MYBPA types. Tannin-rich plants showed an expansion in the number of subgroup 5 R2R3-MYB genes compared with other dicotyledonous and monocot plants. The MYBPA-type genes of tea plant were slightly expanded. qRT-PCR analysis and GUS staining analysis of promoter activity under a series of treatments revealed the differential responses of CsMYB5s to biotic and abiotic stresses. In particular, CsMYB5a, CsMYB5b, and CsMYB5e responded to high-intensity light, high temperature, MeJA, and mechanical wounding, whereas CsMYB5f and CsMYB5g were only induced by wounding. Three genetic transformation systems (C. sinensis, Nicotiana tabacum, and Arabidopsis thaliana) were used to verify the biological function of CsMYB5s. The results show that CsMYB5a, CsMYB5b, and CsMYB5e could promote the gene expression of CsLAR and CsANR. However, CsMYB5f and CsMYB5g could only upregulate the gene expression of CsLAR but not CsANR. A series of site-directed mutation and domain-swapping experiments were used to verify functional domains and key amino acids of CsMYB5s responsible for the regulation of PA biosynthesis. This study aimed to provide insight into the induced expression and functional diversity model of PA biosynthesis regulation in tea plants.

Porous capillary monolithic column coupled with ultrahigh performance liquid chromatography-tandem mass spectrometry for fast and effective separation and determination of estrogens.

In this work, a porous capillary monolithic column was simply prepared by in situ thiol-alkyne click polymerization of dipentaerythritol hexakis (3-mercaptopropionate) and dimethyl dipropargylmalonate in fused-silica capillary. The capillary monolithic column shows excellent permeability, high porosity, and thoiether-rich groups, thereby, a high-efficient capacity for trace estrogens from complex samples are obtained via electron-donor-acceptor π-π interaction and hydrophobic interaction. The highest adsorption efficiency for estrogens is achieved at pH = 7.0 with a flow rate of 0.200 mL min-1. The superior adsorption capacities of the as-prepared capillary column for eight estrogens range from 0.092 mg m-1 to 0.31 mg m-1. A simple, reliable, and sensitive method for the determination of eight estrogens in biological and environmental samples is developed using the monolithic polymer as in-tube solid-phase microextraction coupled with ultrahigh performance liquid chromatography-tandem mass spectrometry (SPME-UPLC-MS/MS), and the total instrumental analysis time for the SPME-UPLC-MS/MS procedures was about 60 min per sample. The developed method shows a wide linear range (0.0500-5.00 µg L-1), and low limits of detection (5.34-9.63 ng L-1) for estrogens. The concentrations of estrogens in serum, urine, and pond water samples are found to be no more than 3.69, 0.741, and 1.04 µg L-1, respectively, and the satisfying recoveries for the eight estrogens range from 80.3% to 113% with relative standard deviations (n = 5) of 1.5-9.4%. The established method is highly potential for extraction and analysis of ultratrace target estrogens in complex matrices, such as biological and environmental samples.

Long noncoding RNA GAS5 alleviates the inflammatory response of human periodontal ligament stem cells by regulating the NF-κB signalling pathway.

OBJECTIVES: This study investigated the role of lncRNA growth arrest-specific transcript 5 (GAS5) in the inflammatory response of periodontal ligament stem cells (PDLSCs) during periodontitis with attempts to its possible mechanisms. MATERIALS AND METHODS: Gingiva samples were collected from healthy people and patients with periodontitis. The ligature-induced periodontitis model was established in mice. Cell transfection was utilized to knock down and overexpress GAS5 in PDLSCs. Quantitative real-time polymerase chain reaction (qRT-PCR) and fluorescence in situ hybridization were performed to detect the GAS5 expression. In combination with high-throughput sequencing technology, qRT-PCR, Western blotting, and immunofluorescence were performed to detect the effects of GAS5 on cytokines and proteins in the NF-κB pathway. RESULTS: GAS5 expression decreased in PDLSCs subjected to compressive force. GAS5 expression was downregulated in the gingiva tissues from patients with periodontitis. Consistent with the results of clinical samples, GAS5 expression decreased in the mouse ligature-induced periodontitis model. GAS5 expression was downregulated in PDLSCs under tumour necrosis factor (TNF)-α stimulation. Knockdown and overexpression of GAS5 increased and decreased the expression of cytokines induced by TNF-α in PDLSCs, respectively. The sequencing results showed that overexpressing GAS5 was related to genes in the NF-κB pathway. Overexpressing GAS5 alleviated p65 phosphorylation and inhibited the entry of p65 into the nucleus in the TNF-α activated NF-κB pathway, whereas GAS5 knockdown resulted in contrasting results. CONCLUSIONS: GAS5 alleviated the expression of cytokines in PDLSCs by inhibiting activation of the TNF-α-mediated NF-κB pathway. These findings provide new insight into the regulation of the PDLSCs inflammation response.

Aptamer functionalized and reduced graphene oxide hybridized porous polymers SPE coupled with LC-MS for adsorption and detection of human α-thrombin.

In this study, reduced graphene oxide (rGO) hybridized high internal phase emulsions were developed and polymerized as porous carriers for aptamer (5'/5AmMC6/-AGT CCG TGG TAG GGC AGG TTG GGG TGA CT-3') modification to enrich human α-thrombin from serum. The structure and properties of the materials were confirmed by scanning electron microscope (SEM), Fourier transform infrared spectroscope (FT-IR), and X-ray photoelectron spectra (XPS). The adsorption ability and selectivity were studied and the thrombin was detected with liquid chromatography-mass spectrometry (LC-MS). The adsorption of thrombin onto the sorbent was achieved within 30 min and the desorption was realized using 5.0 mL of acetonitrile/water (80/20, v/v). The thrombin was quantified by LC-MS according to its characteristic peptide sequence of ELLESYIDGR.

Colorimetric sensing of chloride in sweat based on fluorescence wavelength shift via halide exchange of CsPbBr3 perovskite nanocrystals.

Considering the high importance of the rapid detection of chloride ion (Cl-) in sweat for the diagnosis of fibrotic cysts, we have investigated the heterogeneous halide exchange between CsPbBr3 perovskite nanocrystals (PNCs) in n-hexane and Cl- in aqueous solution. The results show that CsPbBr3 PNCs could achieve fast halide exchange with Cl- in the aqueous phase under magnetic stirring at pH = 1, accompanied by a significant wavelength blue shift and vivid fluorescence color changes from green to blue. Therefore, a fluorescence wavelength shift-based colorimetric sensing of Cl- based on the halide exchange of CsPbBr3 PNCs has been developed to realize the rapid detection of Cl- in sweat. Compared with the conventional fluorescence intensity-based method, this method is of high convenience since the whole procedure could be achieved within 5 min without any sample pretreatment (even no dilution), demonstrating promising application prospects. Graphical Abstract Fluorescence wavelength-shift based colorimetric sensing of chloride in sweat via halide exchange of CsPbBr3 perovskite nanocrystals.

Dual-Mode of Fluorescence Turn-On and Wavelength-Shift for Methylamine Gas Sensing Based on Space-Confined Growth of Methylammonium Lead Tribromide Perovskite Nanocrystals.

The defect-tolerant nature of lead halide perovskites renders outstanding luminescence by simple space-confined growth in nanopores. The fluorescence turn-on and wavelength-shift phenomena could be found in the formation of methylammonium lead tribromide (MAPbBr3) nanocrystals in hollow SiO2 nanospheres triggered by the reaction between methylamine (MA) gas and HPbBr3/PbBr2@SiO2 nanospheres. The enhanced fluorescence intensity is linear with the MA concentration in the range of 1.0-95 ppm with a limit of detection (LOD) of 70 ppb (S/N = 3). In addition, the maximum emission wavelength is consistently red-shifted from 478.7 to 510.6 nm as the MA concentration increases from 1.0 to 95 ppm, imparting the potential for colorimetric sensing. By combining the fluorescence turn-on and colorimetric sensing modes, the flexible method meets the demands for visual discrimination and point-of-care determination with portable devices.

Wavelength-Shift-Based Colorimetric Sensing for Peroxide Number of Edible Oil Using CsPbBr3 Perovskite Nanocrystals.

The peroxide number of edible oil relates to its quality. The classical determination methods for the peroxide number are still unsatisfactory due to their complexity and poor reproducibility in the analytical process and their incapability of field rapid detection. In this study, a novel wavelength-shift-based visual method has been developed for the peroxide number determination of edible oil using halide perovskite nanocrystals (CsPbBr3 NCs). In the analysis, the edible oil sample underwent redox reactions with a part of oleylammonium iodide (OLAM-I) in advance. Then, the halogen exchange occurred between the added CsPbBr3 NCs and the iodide ions from the residual OLAM-I. The resulting wavelength shift of the fluorescence emission reflects the peroxide number in the edible oil sample. Under the ultraviolet light excitation at 365 nm, the apparent color of the photoluminescence could directly be compared with a color chart to determine and qualify the peroxide number. Using the approach, the visual detection of the peroxide number of edible oil samples on site could be realized. The detection process takes only ∼15 min and is convenient and accurate.

Antioxidant activity measurement and potential antioxidant peptides exploration from hydrolysates of novel continuous microwave-assisted enzymolysis of the Scomberomorus niphonius protein.

A novel continuous microwave-assisted enzymatic digestion (cMAED) method is proposed for the digestion of protein from Scomberomorus niphonius to obtain potential antioxidant peptides. In this study, bromelain was found to have a high capacity for the digestion of the Scomberomorus niphonius protein. The following cMAED conditions were investigated: protease species, microwave power, temperature, bromelain content, acidity of the substrate solution, and incubation time. At 400W, 40°C, 1500U·g-1 bromelain, 20% substrate concentration, pH 6.0 and 5min incubation, the degree of hydrolysis and total antioxidant activity of the hydrolysates were 15.86% and 131.49µg·mL-1, respectively. The peptide analyses showed that eight of the potential antioxidant peptide sequences, which ranged from 502.32 to 1080.55Da with 4-10 amino acid residues, had features typical of well-known antioxidant proteins. Thus, the new cMAED method can be useful to obtain potential antioxidant peptides from protein sources, such as Scomberomorus niphonius.