Oriented Assembled Prussian Blue Analogue Framework for Confined Catalytic Decomposition of Ammonium Perchlorate.

The design of highly dispersed active sites of hollow materials and unique contact behavior with the components to be catalyzed provide infinite possibilities for exploring the limits of catalyst capacity. In this study, the synthesis strategy of highly open 3-dimensional frame structure Prussian blue analogues (CoFe-PBA) was explored through structure self-transformation, which was jointly guided by template mediated epitaxial growth, restricted assembly and directional assembly. Additionally, good application prospect of CoFe-PBA as combustion catalyst was discussed. The results show that unexpected thermal decomposition behavior can be achieved by limiting AP(ammonium perchlorate) to the framework of CoFe-PBA. The high temperature decomposition stage of AP can be advanced to 283.6 °C and the weight loss rate can reach 399.0%. In-situ monitoring shows that CoFe-PBA can accelerate the formation of NO and NO2 . The calculation of reaction kinetics proved that catalytic process was realized by increasing the nucleation factor. On this basis, the catalytic mechanism of CoFe-PBA on the thermal decomposition of AP was discussed, and the possible interaction process between AP and CoFe-PBA during heating was proposed. At the same time, another interesting functional behavior to prevent AP from caking was discussed.

Extraction, separation and kinetics of phenylethanosides from Plantago asiatica L. by an innovative extraction technology-deep eutectic solvent-based ultrasound-assisted extraction.

In this paper, the total phenylethanosides (TPS) were extracted efficiently by an innovative extraction technology--deep eutectic solvent-based ultrasound-assisted extraction (DES-UAE) from Plantago asiatica L. Ten diverse types of DESs were synthesized as alternative extraction solutions. The extraction efficiency of DES-3 (constituted by choline chloride and lactic acid) was much higher than those of other DESs. On the basis of single factor tests and Box-Behnken design (BBD), the optimum processing parameters of DES-UAE as follow: DES-3 with molar ratio of 1:3, extraction temperature 51 °C, solid/liquid 22.5 mg/ml, water content 30%, ultrasonic power 65 W, extraction time 23 min. The extraction efficiency of TPS from Plantago asiatica L. was 8.395 mg/ml, which was more superior than those of organic solvents (water, methanol, 50% methanol, ethanol, 50% ethanol). The extraction kinetics experiment results showed that water content had a significant influence upon the extraction efficiency of TPS. At the same time, AB-8 macroporous resin column was used to efficiently isolate TPS from DES extraction with a recovery rate of 88.5%.

Porphyromonas gingivalis bacteremia increases the permeability of the blood-brain barrier via the Mfsd2a/Caveolin-1 mediated transcytosis pathway.

Bacteremia induced by periodontal infection is an important factor for periodontitis to threaten general health. P. gingivalis DNA/virulence factors have been found in the brain tissues from patients with Alzheimer's disease (AD). The blood-brain barrier (BBB) is essential for keeping toxic substances from entering brain tissues. However, the effect of P. gingivalis bacteremia on BBB permeability and its underlying mechanism remains unclear. In the present study, rats were injected by tail vein with P. gingivalis three times a week for eight weeks to induce bacteremia. An in vitro BBB model infected with P. gingivalis was also established. We found that the infiltration of Evans blue dye and Albumin protein deposition in the rat brain tissues were increased in the rat brain tissues with P. gingivalis bacteremia and P. gingivalis could pass through the in vitro BBB model. Caveolae were detected after P. gingivalis infection in BMECs both in vivo and in vitro. Caveolin-1 (Cav-1) expression was enhanced after P. gingivalis infection. Downregulation of Cav-1 rescued P. gingivalis-enhanced BMECs permeability. We further found P. gingivalis-gingipain could be colocalized with Cav-1 and the strong hydrogen bonding between Cav-1 and arg-specific-gingipain (RgpA) were detected. Moreover, P. gingivalis significantly inhibited the major facilitator superfamily domain containing 2a (Mfsd2a) expression. Mfsd2a overexpression reversed P. gingivalis-increased BMECs permeability and Cav-1 expression. These results revealed that Mfsd2a/Cav-1 mediated transcytosis is a key pathway governing BBB BMECs permeability induced by P. gingivalis, which may contribute to P. gingivalis/virulence factors entrance and the subsequent neurological impairments.

Advanced MRI features of intraventricular inflammatory myofibroblastic tumor: a case report.

BACKGROUND: Inflammatory myofibroblastic tumor (IMT) is a rare central nervous system (CNS) tumor. We first report a rare case of IMT in the lateral ventricle and describe the magnetic resonance imaging (MRI) findings of the tumor with an emphasis on the advanced MRI features. CASE PRESENTATION: A 49-year-old female patient with headaches and blurred vision for 2 months. Brain MRI revealed a well-circumscribed, lobulated mass occupying the left lateral ventricle trigone, with marked perilesional brain edema. The tumor showed heterogeneous significant hyperintensity on T2-weighted imaging (T2WI) and hypointensity on T1-weighted imaging (T1WI). After the administration of gadolinium, the mass exhibited marked contrast enhancement and the halo sign was observed. On advanced MRI, the lesion showed decreased perfusion on perfusion MRI and reduced diffusion on diffusion-weighted imaging (DWI). On susceptibility-weighted imaging (SWI), there was a punctate low signal intensity in the tumor. The patient underwent surgical resection of the mass and a pathological examination confirmed the lesion to be an inflammatory myofibroblastic tumor with negative expression of anaplastic lymphoma kinase (ALK). This patient had remained healthy without evidence of recurrence during a 20-month follow-up. CONCLUSIONS: On MRI, marked perilesional brain edema, significant hyperintensity on T2WI, hypoperfusion on perfusion MRI but with an obvious enhancement, no diffusion restriction on DWI, and halo sign may be the characteristic findings of intraventricular IMT. The advanced MRI characteristics could provide abundant information to reflect the histological features and physiological metabolic characteristics of the tumor.

Polyetherimide- and folic acid-modified Fe3 O4 nanospheres for enhanced magnetic hyperthermia performance.

Recent studies have highlighted the development prospects of magnetic hyperthermia in cancer therapy. A few studies on the application of Fe3 O4 nanospheres for the magnetic hyperthermia of gynecological malignancies have achieved certain efficacy, but there was no visible progress currently. In this work, Fe3 O4 nanospheres modified with polyetherimide (PEI) and folic acid (FA) were synthesized using a hydrothermal method for possible utility in biocompatible and active tumor-targeting magnetic induction hyperthermia. The PEI- and FA-coated Fe3 O4 nanospheres showed high crystallinity, well-dispersed spherical structures and ideal Ms value. As a result, the designed Fe3 O4 @ PEI@FA nanospheres achieved higher specific absorption rate (SAR) values at 360 kHz and 308 Oe, as well as excellent biocompatibility in Hela, SKOV3, HEC-1-A and NIH3T3 cells. These nanospheres can be used as an optimal heating agent for the magnetic hyperthermia treatment of gynecological cancers.

In Situ Cutting of Ammonium Perchlorate Particles by Co-Bipy "scalpel" for High Efficiency Thermal Decomposition.

Burning rate of solid propellants can be effectively improved by adding catalysts and using smaller size ammonium perchlorate (AP). Although few reports, the exploration of changing the size of AP primary particles by catalysts is of great significance for improving combustion performance. Here, taking Co-bipy as an example, the potential advantages of such materials as AP decomposition catalysts are reported. Due to the existence of NO3 - combined with oxygen rich environment provided by AP, the structural self-transformation from micronrods to nanoparticles can be quickly realized during the heating process. More importantly, when Co-bipy decomposes, it can play the role of "scalpel" and in situ cut AP particles. Results show that high-temperature decomposition of Co-bipy/AP occurs at 305.8 °C, which is 137.5 °C lower than that of pure AP. Catalytic mechanism is discussed by in situ IR and TG-IR, CoO can effectively increase the content of reactive oxygen species and weaken the N-H bond, realizing the rapid oxidation of NH3 . Eventually, the behavior of Co-bipy cutting AP particles is tested. This interesting catalyst structure self-transformation behavior can not only realize the influence on AP, but also perform a positive function in the combustion process of solid propellants, such as opening the adhesive AP interface.

Lyophilizer Leak Rate Testing - An Industry Survey and Best Practice Recommendation.

The vacuum integrity of freeze dryers is critical for attaining adequate process control and maintaining confidence in sterility assurance which is key for the manufacture of sterile pharmaceutical products. Although discussions on the topic have been published, there is no industry standard established that is based on empirical data or that has a justifiable scientific rationale. This article provides a review of the scientific literature in the public domain and most importantly, a perspective from 14 Pharmaceutical companies on the leak rate specifications commonly used in industry. Using this information we recommend a best practice for the lyophilizer leak rate test which includes detailing necessary preparation activities following Steam-In-Place (SIP) sterilization, defining a period of stabilization to eliminate pressure and temperature fluctuations and details of the test conditions and the test period. We conclude that for routine manufacturing practice the operational leak rate should not exceed 20 µbar L/s and we provide additional guidance for large volume and older lyophilisation equipment.

Metformin Combining PD-1 Inhibitor Enhanced Anti-Tumor Efficacy in STK11 Mutant Lung Cancer Through AXIN-1-Dependent Inhibition of STING Ubiquitination.

Background: Non-small-cell lung cancer (NSCLC) with STK11 mutation showed primary resistance to immune checkpoint inhibitors (ICIs). The glucose-lowering drug metformin exerted anti-cancer effect and enhanced efficacy of chemotherapy in NSCLC with KRAS/STK11 co-mutation, yet it is unknown whether metformin may enhance ICI efficacy in STK11 mutant NSCLC. Methods: We studied the impact of metformin on ICI efficacy in STK11 mutant NSCLC in vitro and in vivo using colony formation assay, cell viability assay, Ki67 staining, ELISA, CRISPR/Cas9-mediated knockout, and animal experiments. Results: Through colony formation assay, Ki67 incorporation assay, and CCK-8 assay, we found that metformin significantly enhanced the killing of H460 cells and A549 cells by T cells. In NOD-SCID xenografts, metformin in combination with PD-1 inhibitor pembrolizumab effectively decreased tumor growth and increased infiltration of CD8+ T cells. Metformin enhanced stabilization of STING and activation of its downstream signaling pathway. siRNA-mediated knockdown of STING abolished the effect of metformin on T cell-mediated killing of tumor cells. Next, we found that CRISPR/Cas9-mediated knockout of the scaffold protein AXIN-1 abolished the effect of metformin on T cell-mediated killing and STING stabilization. Immunoprecipitation and confocal macroscopy revealed that metformin enhanced the interaction and colocalization between AXIN-1 and STING. Protein-protein interaction modeling indicated that AXIN-1 may directly bind to STING at its K150 site. Next, we found that metformin decreased K48-linked ubiquitination of STING and inhibited the interaction of E3-ligand RNF5 and STING. Moreover, in AXIN-1 -/- H460 cells, metformin failed to alter the interaction of RNF5 and STING. Conclusion: Metformin combining PD-1 inhibitor enhanced anti-tumor efficacy in STK11 mutant lung cancer through inhibition of RNF5-mediated K48-linked ubiquitination of STING, which was dependent on AXIN-1.

Ibrutinib reverses IL-6-induced osimertinib resistance through inhibition of Laminin α5/FAK signaling.

Osimertinib, a 3rd generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI), is the first-line standard-of-care for EGFR-mutant non-small cell lung cancer (NSCLC) patients, while acquired drug resistance will inevitably occur. Interleukin-6 (IL-6) is a keystone cytokine in inflammation and cancer, while its role in osimertinib efficacy was unknown. Here we show that clinically, plasma IL-6 level predicts osimertinib efficacy in EGFR mutant NSCLC patients. Highly increased IL-6 levels are found in patients with acquired resistance to osimertinib. Addition of IL-6 or exogenous overexpression of IL-6 directly induces osimertinib resistance. Proteomics reveals LAMA5 (Laminin α5) and PTK2, protein tyrosine kinase 2, also called focal adhesion kinase (FAK), are activated in osimertinib-resistant cells, and siRNA knockdown of LAMA5 or PTK2 reverses IL-6-mediated osimertinib resistance. Next, using a large-scale compound screening, we identify ibrutinib as a potent inhibitor of IL-6 and Laminin α5/FAK signaling, which shows synergy with osimertinib in osimertinib-resistant cells with high IL-6 levels, but not in those with low IL-6 levels. In vivo, this combination inhibits tumor growth of xenografts bearing osimertinib-resistant tumors. Taken together, we conclude that Laminin α5/FAK signaling is responsible for IL-6-induced osimertinib resistance, which could be reversed by combination of ibrutinib and osimertinib.

Exploration of a ternary deep eutectic solvent for the efficient extraction of plantamajoside, acteoside, quercetin and kaempferol from Plantago asiatica L.

INTRODUCTION: In the present study, ternary deep eutectic solvent-based ultrasound-assisted extraction was developed for the efficient extraction of plantamajoside, acteoside, quercetin and kaempferol from Plantago asiatica L. METHODOLOGY: Six kinds of choline chloride-based ternary deep eutectic solvents (TDESs) were prepared as potential extraction solutions. In order to obtain optimal extraction efficiency, a series of extraction conditions were investigated by single-factor test and orthogonal test. RESULTS: The extraction efficiency of choline chloride/lactic acid/ethylene glycol (ChCl-LA-EG) was much higher than that of other TDESs. ChCl-LA-EG-11 synthesised with choline chloride, lactic acid and ethylene glycol (1:4:2) was considered to have a higher extraction efficiency. The optimal ultrasound-assisted extraction conditions were as follows: water content in ChCl-LA-EG-11, 50%; extraction temperature, 70°C; ratio of solid/liquid, 20 mg/mL; ultrasonic power, 60 W; extraction time, 35 min; pH of the solution, 8. Under the optimal extraction conditions, the extraction efficiencies of plantamajoside, acteoside, quercetin and kaempferol were 3.83 ± 0.41, 4.23 ± 0.45, 0.56 ± 0.15 and 0.19 ± 0.08 mg/g, respectively. The extraction efficiency of the total target components was 9.21 ± 0.63 mg/g, which was much higher than that of conventional solvents (water, methanol, ethanol, 50% methanol, 50% ethanol). The target components were isolated efficiently from the TDES solution by an AB-8 macroporous resin column with a recovery rate of 95.6%. CONCLUSION: This study demonstrated that TDESs possessed excellent physical and chemical properties and had enormous potential for active component extraction of traditional Chinese medicinal materials.

Calcitriol-enhanced autophagy in gingival epithelium attenuates periodontal inflammation in rats with type 2 diabetes mellitus.

Type 2 diabetes mellitus (T2DM)-associated periodontitis is a common disease with high prevalence, associated with persistent infection and complicated manifestations. Calcitriol (1 alpha, 25-dihydroxyvitamin D3, 1,25D) is the active form of vitamin D that plays a protective role in immune regulation, bone metabolism, and inflammatory response. In this study, we constructed a T2DM model in rats by combining a high-fat diet with low-dose streptozotocin. The periodontitis model in rats was developed by ligation and Porphyromonas gingivalis (ATCC 33277) inoculation. Rats were randomly divided into five groups: non-diabetic blank, diabetic blank, diabetes with calcitriol treatment, diabetes with 3-methyladenine (3-MA) treatment, or diabetes with calcitriol and 3-MA treatment. The diabetic rats exhibited an intense inflammatory response and decreased autophagy compared with the non-diabetic rats. Intraperitoneal injection of calcitriol and autophagy inhibitor (3-MA) allowed us to explore the effect of calcitriol on inflammation in the gingival epithelium and the role of autophagy in this process. Treatment with calcitriol resulted in the decreased expression of NFκB-p65, p62/SQSTM1 and inflammatory response and increased expression of LC3-II/LC3-I. Application of 3-MA significantly suppressed autophagy, which was apparently retrieved by calcitriol. Antibacterial peptide (LL-37) is the only antimicrobial peptide in the cathelicidin family that is found in the human body, and it exhibits a broad spectrum of antibacterial activity and regulates the immune system. In the present study, our findings indicated that calcitriol-enhanced autophagy may attenuated periodontitis and the decrease of LL-37 was rescued by calcitriol treatment in the gingival epithelial cells of T2DM rats. Our study provides evidence for the application of calcitriol as an adjunctive treatment for T2DM-associated periodontitis.

Oral Pathogen Fusobacterium nucleatum Coaggregates With Pseudomonas aeruginosa to Modulate the Inflammatory Cytotoxicity of Pulmonary Epithelial Cells.

Chronic obstructive pulmonary disease (COPD) is the third leading cause of mortality worldwide, and inflammatory damage induced by bacterial infections is an important contributor to the etiology of COPD. Fusobacterium nucleatum, a recognized periodontal pathogen, is considered as a biomarker of lung function deterioration of COPD patients coinfected with Pseudomonas aerugionsa, but the underlying mechanism is still unclear. This study established single- and dual-species infection models, bacterial simultaneous and sequential infection models, and found that F. nucleatum could coaggregate with P. aeruginosa to synergistically invade into pulmonary epithelial cells and transiently resist P. aeruginosa-induced cytotoxic damage to amplify IL-6 and TNF-α associated inflammation in pulmonary epithelial cells simultaneously infected with P. aeruginosa and F. nucleatum. Furthermore, F. nucleatum pretreatment or subsequential infection could maintain or even aggravate P. aeruginosa-induced inflammatory cytotoxicity of pulmonary epithelial cells. These results indicate that oral pathogen F. nucleatum coaggregates with P. aeruginosa to facilitate bacterial invasion and modulates the inflammatory cytotoxicity of pulmonary epithelial cells, which may contribute to lung function deterioration of COPD patients accompanied with P. aeruginosa and F. nucleatum coinfection.

The Effects of Porphyromonas gingivalis on Inflammatory and Immune Responses and Osteogenesis of Mesenchymal Stem Cells.

Mesenchymal stem cells (MSCs) are increasingly used in tissue regeneration, not only because of their multilineage differentiation ability, but also because of their immunomodulatory function, which allows them to play a role in the inflammatory milieu, especially in periodontitis. Porphyromonas gingivalis (P. gingivalis) is an important pathogen associated with the progression of periodontitis. Heterogeneous MSC sources show differences in their inflammatory-immune responsiveness and osteogenesis capabilities when exposed to P. gingivalis and its virulence factors. This article reviews the promoted inflammatory and immune responses of periodontal ligament stem cells, which are potential pitfalls in bone regeneration. MSCs from other sources showed contradictory inflammatory and immune reactions in the few studies on this topic. We also summarize the mechanisms involved in the inflammatory, immune responses and osteogenic potential of MSCs exposed to P. gingivalis and its virulence factors to inform an improved utilization of MSCs in regenerative therapies for periodontitis.

Vitamin D decreases Porphyromonas gingivalis internalized into macrophages by promoting autophagy.

OBJECTIVES: This paper aims to study the effect of the active form of vitamin D (calcitriol) on the internalized Porphyromonas gingivalis in macrophages and to assess the role of autophagy during this process. MATERIALS AND METHODS: Quantitative RT-PCR and bacteria culture were used to quantify live P. gingivalis internalized into U937-derived macrophages. Western blot assays were performed to detect the effect of P. gingivalis and calcitriol on autophagy in macrophages. Transmission electron microscope was used to observe the effect of calcitriol on the status of internalized P. gingivalis. Colocalization of P. gingivalis with the autophagosome and lysosome markers was observed by confocal laser scanning microscopy. RESULTS: Calcitriol caused a dose-dependent decrease in live P. gingivalis numbers and promoted both the endogenous and P. gingivalis-induced autophagy in macrophages. Calcitriol significantly promoted the destruction of P. gingivalis and the colocalization of P. gingivalis with autophagosome and lysosome markers. Conversely, with 3-MA, live P. gingivalis numbers in macrophages increased significantly and inhibition effect of calcitriol on the number of live P. gingivalis was attenuated. CONCLUSION: In U937-derived macrophages, calcitriol may promote colocalization of P. gingivalis with autophagosomes and lysosomes, namely autophagy process, to degrade live P. gingivalis.

LL-37-Induced Autophagy Contributed to the Elimination of Live Porphyromonas gingivalis Internalized in Keratinocytes.

Porphyromonas gingivalis (P. gingivalis), one of the most important pathogens of periodontitis, is closely associated with the aggravation and recurrence of periodontitis and systemic diseases. Antibacterial peptide LL-37, transcribed from the cathelicidin antimicrobial peptide (CAMP) gene, exhibits a broad spectrum of antibacterial activity and regulates the immune system. In this study, we demonstrated that LL-37 reduced the number of live P. gingivalis (ATCC 33277) in HaCaT cells in a dose-dependent manner via an antibiotic-protection assay. LL-37 promoted autophagy of HaCaT cells internalized with P. gingivalis. Inhibition of autophagy with 3-methyladenine (3-MA) weakened the inhibitory effect of LL-37 on the number of intracellular P. gingivalis. A cluster of orthologous groups (COGs) and a gene ontology (GO) functional analysis were used to individually assign 65 (10%) differentially expressed genes (DEGs) to an "Intracellular trafficking, secretion, and vesicular transport" cluster and 306 (47.08%) DEGs to metabolic processes including autophagy. Autophagy-related genes, a tripartite motif-containing 22 (TRIM22), and lysosomal-associated membrane protein 3 (LAMP3) were identified as potentially involved in LL-37-induced autophagy. Finally, bioinformatics software was utilized to construct and predict the protein-protein interaction (PPI) network of CAMP-TRIM22/LAMP3-Autophagy. The findings indicated that LL-37 can reduce the quantity of live P. gingivalis internalized in HaCaT cells by promoting autophagy in these cells. The transcriptome sequencing and analysis also revealed the potential molecular pathway of LL-37-induced autophagy.

A validated miRNA signature for the diagnosis of osteoporosis related fractures using SVM algorithm classification.

The aim of the present study was to develop a circulating microRNA expression signature for early prediction of osteoporotic fractures and to validate the results using Gene Expression Omnibus (GEO) datasets. The GSE70318 dataset was downloaded from GEO and used to build an osteoporotic fracture prediction model based on the receiver operating characteristic curve and support vector machine (SVM) classification index. The GSE74209 dataset was used as a validation dataset. Additionally, in vitro, alkaline phosphatase (ALP) activity was measured in the presence or absence of microRNA (miRNA/miR) treatments in human osteoblast cells. The expression of two selected genes was detected by western blotting. miR-188-3p, miR-942-3p, miR-576-3p and miR-135a-5p were differentially expressed between controls and osteoporotic patients with fractures. SVM classification using these four miRNAs provided better dichotomization. It was further confirmed that miR-576-3p and 135a-5p in the GSE74209 dataset could also significantly discriminate between the controls and fracture patients, the area under the curve of SVM2 was 0.9722 with 95% CI 0.8885-1.056. Further analysis indicated that the target genes of the two miRNAs participated in the Wingless-related integration site, Hedgehog and transforming growth factor-ß signaling pathways and osteoclast differentiation. miR-576-3p and miR-135-5p transfection decreased ALP activity and ALP activity was increased in the presence of blocking antisense oligonucleotides. Western blotting indicated miR-576-3p and miR-135-5p decreased CSNK1A1L and LRP6 levels, respectively. In conclusion, two miRNA signatures were developed and validated for the prediction of osteoporotic fractures.

Genome-wide analysis of WRKY transcription factors in Aquilaria sinensis (Lour.) Gilg.

The WRKY proteins are a superfamily of transcription factor that regulate diverse developmental and physiological processes in plants. Completion of the whole-genome sequencing of Aquilaria sinensis allowed us to perform a genome-wide investigation for WRKY proteins. Here, we predicted 70 WRKY genes from the A. sinensis genome and undertaken a comprehensive bioinformatic analysis. Due to their diverse structural features, the 70 AsWRKY genes are classified into three main groups (group I-III), with five subgroups (IIa-IIe) in group II, except two belong to none of them. Distinct expression profiles of AsWRKYs with RNA sequencing data revealed their diverse expression patterns among different tissues and in the process of whole-tree-inducing agarwood formation. Based on the expression characteristics, we predict some AsWRKYs are pseudogenes, and some may be involved in the biosynthesis of agarwood sesquiterpenes as activators or repressors. Among the tested genes treated with MeJA and H2O2, most of them are induced by H2O2, but downregulated by MeJA, implying the complexity of their involvement in signal transduction regulation. Our results not only provide a basic platform for functional identification of WRKYs in A. sinensis but important clues for further analysis their regulation role in agarwood formation.

WRKY44 represses expression of the wound-induced sesquiterpene biosynthetic gene ASS1 in Aquilaria sinensis.

Agarwood is derived from wounds in Aquilaria trees and is widely used in traditional medicine, incense, and perfume. Sesquiterpenes are one of the main active components in agarwood and are known to be induced by wounding or injury; However, the molecular mechanisms by which wounding leads to sesquiterpene formation remain largely unknown. Agarwood sesquiterpene synthase 1 (ASS1) is one of key enzymes responsible for the biosynthesis of sesquiterpenes and is a crucial jasmonate (JA)-responsive wound-inducible synthase. However, it is not known why ASS1 is not expressed in healthy trees and how its expression is induced as a result of wounding. Here, we report that ASS1 is a wound-induced gene with a promoter in which a 242-bp region (-973 to -731bp) is identified as the core sequence for responding to wound signals. AsWRKY44 binds directly to this region and represses ASS1 promoter activity. Down-regulation or disruption of AsWRKY44 can relieve the inhibition and activate ASS1 expression. In addition, AsWRKY44 is degraded and the expression of ASS1 is significantly up-regulated in response to exogenous application of methyl jasmonate. Thus, AsWRKY44 is a crucial negative regulator of wound-induced ASS1 transcription, and is central to the mechanism of sesquiterpene biosynthesis in agarwood.

Applicability of multifunctional preprocessing device for simultaneous estimation of spreading of green tea, withering of black tea and shaking of oolong tea.

BACKGROUND: Preprocessing technologies of fresh tea leaves have a great influence on tea quality. A multifunctional preprocessing device for tea raw materials has been designed and utilized as a novel item of equipment to synchronously meet the process needs of spreading of green tea, withering of black tea and shaking of oolong tea. RESULTS: The preprocessing parameters of fresh tea leaves for spreading of green tea, withering of black tea and shaking of oolong tea were optimized by orthogonal experiments. Sensory assessment combined with statistical tools was employed as an analytical method to evaluate the pretreatment effect of processing different sorts of tea. The range analysis and variance analysis of tea sensory evaluation combined with chemical components (total polyphenols, free amino acids and soluble sugar) showed that A3 B2 C3 (70%, 25 °C, 8 h), A1 B3 C1 (60%, 28 °C, 18 h) and A1 B1 C3 (5 min, 20 °C, discontinuous leaf turning c) were considered to be the optimum schemes for the best pretreatment conditions of the above three major processing types of tea, respectively. The verification experiment of the proposed schemes was performed with satisfactory performance. CONCLUSION: This study demonstrated that a multifunctional preprocessing device for fresh tea leaves can be successfully applied to simultaneously estimate spreading of green tea, withering of black tea and shaking of oolong tea. © 2019 Society of Chemical Industry.

Calcitriol decreases live Porphyromonas gingivalis internalized into epithelial cells and monocytes by promoting autophagy.

BACKGROUND: The present study aimed to explore the effects of the active form of vitamin D (calcitriol, 1α,25-dihydroxyvitamin D3 , 1α,25 (OH)2 D3 , 1,25D) on live Porphyromonas gingivalis internalized into KB cells and U937 cells. METHODS: Quantitative real-time polymerase chain reaction method was used to evaluate the number of surviving P. gingivalis internalized into KB cells and U937 cells. Transmission electron microscopy was used to detect P. gingivalis in cells. A western blot analysis was performed to observe LC3 expressions. RESULTS: 1) Treatment with 1,25D decreased the number of live P. gingivalis in KB cells and U937 cells in a dose-dependent manner. 2) Dividing P. gingivalis were found only in KB cells but not in U937 cells. The cell walls of most P. gingivalis in KB cells were intact, while those in U937 cells were disrupted. Treatment with 1,25D promoted the encapsulation of P. gingivalis in autophagosomes in both KB and U937 cells. 3) Both 1,25D treatment and P. gingivalis infection increased the LC3 II/I ratio. Furthermore, 1,25D treatment increased the P. gingivalis-upregulated LC3 II/I ratio. 4) Treatment with 3-methyladenine (3-MA) decreased the number of P. gingivalis by 11.41% in KB cells, while increased that by 121.51% in U937 cells. Under 1,25D treatment conditions, 3-MA treatment increased the number of P. gingivalis by 88.71% in KB cells and by 284.70% in U937 cells. CONCLUSIONS: Autophagy may facilitate P. gingivalis survival in KB cells and eliminate P. gingivalis in U937 cells. Treatment with 1,25D may help decrease the number of live P. gingivalis in KB cells and U937 cells by promoting functional autophagy.