TRANSPARENT TESTA GLABRA1 Regulates High-Intensity Blue Light-Induced Phototropism by Reducing CRYPTOCHROME1 Levels.

The asymmetrical distribution of auxin supports high intensity blue light (HBL)-mediated phototropism. Flavonoids, secondary metabolites induced by blue light and TRANSPARENT TESTA GLABRA1 (TTG1), alter auxin transport. However, the role of TTG1 in HBL-induced phototropism in Arabidopsis (Arabidopsis thaliana) remains unclear. We found that TTG1 regulates HBL-mediated phototropism. HBL-induced degradation of CRYPTOCHROME 1 (CRY1) was repressed in ttg1-1, and depletion of CRY1 rescued the phototropic defects of the ttg1-1 mutant. Moreover, overexpression of CRY1 in a cry1 mutant background led to phototropic defects in response to HBL. These results indicated that CRY1 is involved in the regulation of TTG1-mediated phototropism in response to HBL. Further investigation showed that TTG1 physically interacts with CRY1 via its N-terminus and that the added TTG1 promotes the dimerization of CRY1. The interaction between TTG1 and CRY1 may promote HBL-mediated degradation of CRY1. TTG1 also physically interacted with blue light inhibitor of cryptochrome 1 (BIC1) and Light-Response Bric-a-Brack/Tramtrack/Broad 2 (LRB2), and these interactions either inhibited or promoted their interaction with CRY1. Exogenous gibberellins (GA) and auxins, two key plant hormones that crosstalk with CRY1, may confer the recovery of phototropic defects in the ttg1-1 mutant and CRY1-overexpressing plants. Our results revealed that TTG1 participates in the regulation of HBL-induced phototropism by modulating CRY1 levels, which are coordinated with GA or IAA signaling.

The ethanolic extract of Artemisia anomala exerts anti-inflammatory effects via inhibition of NLRP3 inflammasome.

BACKGROUND: Artemisia anomala S. Moore (Compositae), known as "Nan-Liu-Ji-Nu" in traditional Chinese medicine (TCM), has been used to treat many inflammatory diseases, including enteritis, acute icteric hepatitis, rheumatism, toothache, tonsillitis, and chronic bronchitis, for centuries. Our preliminary studies have demonstrated that the ethanolic extract of A. anomala (EAA) might be with the potential of inhibiting the activation of the NLRP3 inflammasome. However, the anti-inflammatory activity of EAA based on NLRP3 inflammasome inhibition is still unclear. PURPOSE: This work aimed to elucidate the anti-inflammatory mechanism of EAA by inhibiting NLRP3 inflammasome activation. METHODS: Lipopolysaccharide (LPS)-primed bone marrow-derived macrophages (BMDMs) were used to evaluate the inhibitory effects on NLRP3 inflammasome activation. The level of IL-1ß was determined by ELISA. The expression levels of IL-1ß, caspase-1, NLRP3, and ASC were assayed using western blot analysis. ASC oligomerization and speck formation were detected by immunofluorescence microscopy. The measurements of intracellular chloride and potassium were conducted using N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide (MQAE) probe assay and inductively coupled plasma-optical emission spectrometry (ICP-OES), respectively. Mitochondrial reactive oxygen species (mtROS) were examined using the MitoSOX method. Acridine orange (AO) staining was used to detect the permeability of the lysosomal membrane. A DSS-induced ulcerative colitis model was established to evaluate the anti-inflammatory effects of EAA in vivo. Finally, high-performance liquid chromatography (HPLC) was employed to identify and quantify the major constituents of EAA. RESULTS: In BMDMs, EAA significantly inhibited the release of IL-1ß induced by LPS. The mechanistic study revealed that EAA inhibited NLRP3 inflammasome activation by blocking the oligomerization of ASC and suppressed the LPS-induced priming step. Furthermore, EAA protected lysosomes by inhibiting the TAK1-JNK pathway, thereby inhibiting the assembly of downstream NLRP3 inflammasome and the production of IL-1ß. In addition, EAA exerted potent protective effects in an ulcerative colitis model by decreasing the content of colonic IL-1ß and alleviating the process of ulcerative colitis. HPLC analysis identified eight main components of EAA, including isofraxidin (1), quercetin-7-O-ß-D-glucopyranoside (2), apigenin-7-O-ß-D-glucopyranoside (3), 7-methoxycoumarin (4), quercetin (5), luteolin (6), kaempferol (7), and eupatorin (8), Of these compounds, quercetin and kaempferol were found to be the most potent ingredients. CONCLUSION: These findings collectively reveal that EAA exerts anti-inflammatory effects by both suppressing the NLRP3 priming step and protecting lysosomes to inhibit NLRP3 inflammasome activation, suggesting that this traditional herbal medicine might be used to treat NLRP3-driven inflammatory diseases.

Synthesis and evaluation of new compounds bearing 3-(4-aminopiperidin-1-yl)methyl magnolol scaffold as anticancer agents for the treatment of non-small cell lung cancer via targeting autophagy.

Magnolol and honokiol are the two major active ingredients with similar structure and anticancer activity from traditional Chinese medicine Magnolia officinalis, and honokiol is now in a phase I clinical trial (CTR20170822) for advanced non-small cell lung cancer (NSCLC). In search of potent lead compounds with better activity, our previous study has demonstrated that magnolol derivative C2, 3-(4-aminopiperidin-1-yl)methyl magnolol, has better activity than honokiol. Here, based on the core of 3-(4-aminopiperidin-1-yl)methyl magnolol, we synthesized fifty-one magnolol derivatives. Among them, compound 30 exhibited the most potent antiproliferative activities on H460, HCC827, H1975 cell lines with the IC50 values of 0.63-0.93 µM, which were approximately 10- and 100-fold more potent than those of C2 and magnolol, respectively. Besides, oral administration of 30 and C2 on an H460 xenograft model also demonstrated that 30 has better activity than C2. Mechanism study revealed that 30 induced G0/G1 phase cell cycle arrest, apoptosis and autophagy in cancer cells. Moreover, blocking autophagy by the autophagic inhibitor enhanced the anticancer activity of 30in vitro and in vivo, suggesting autophagy played a cytoprotective role on 30-induced cancer cell death. Taken together, our study implied that compound 30 combined with autophagic inhibitor could be another choice for NSCLC treatment in further investigation.

Flavonoids with Inhibitory Effects on NLRP3 Inflammasome Activation from Millettia velutina.

Eight new flavonoids, including two ß-hydroxy/methoxychalcones, velutones A and B (1 and 2), two 1,3-diarylpropan-1-ols, velutols C and D (3 and 4), a dihydroxychalcone, velutone E (5), a chalcone, velutone F (6), a furanoflavanone, velutone G (7), and a furanoflavonol, velutone H (8), and 14 known compounds were isolated from Millettia velutina. Their structures were determined by high-resolution electrospray ionisation mass spectrometry (HR-ESIMS) and spectroscopic data analyses and time-dependent density functional theory electronic circular dichroism (TD-DFT-ECD) calculations. Among the isolated constituents, compound 6 exhibited the most potent inhibitory effect (IC50: 1.3 µM) against nigericin-induced IL-1ß release in THP-1 cells. The initial mechanism of action study revealed that compound 6 suppressed NLRP3 inflammasome activation via blocking ASC oligomerization without affecting the priming step, which subsequently inhibited caspase-1 activation and IL-1ß secretion. Most importantly, compound 6 exerted potent protective effects in the LPS-induced septic shock mice model by improving the survival rate of mice and suppressing serum IL-1ß release. These results demonstrated that compound 6 had the potential to be developed as a broad-spectrum NLRP3 inflammasome inhibitor for the treatment of NLRP3-related disease.

[Effects of α-enolase gene interference expression on proliferation and apoptosis of follicular granulosa cells from Zi geese].

Objective: To study the effects of α-enolase (ENO1) gene interference expression on proliferation, and cell cycle of follicular granulosa cells from Zi geese. Methods: F1 follicular granulosa cells were primary cultured (mixed culture), which were divided into four groups: ENO1 interference expression group (RNAi), unrelated sequence group (NC), culture group (Control), transfection reagent group (Lip). The apoptosis rate and cell cycle phase of the interference group and the control group were detected by the flow cytometry. Results: ENO1 gene interference expression slowed the proliferation of granulosa cells, increased the apoptosis, and increased the proportion of granulosa cells in G2/M phase. Conclusion: ENO1 gene interference expression could cause G2/M phase arrest in primary cultured goose follicular granulosa cells, induce cell apoptosis and inhibit cell proliferation.

Modulation of LPS-induced inflammation in RAW264.7 murine cells by novel isoflavonoids from Millettia pulchra.

Millettia pulchra is a renowned anti-inflammatory herbal medicine in southeast provinces of China. However, the underlying anti-inflammation mechanism remained incompletely understood. Herein, four new isoflavones, pulvones A-D and eleven reported constituents were isolated from the stems of Millettia pulchra with their structures being elucidated by HRMS and NMR analysis. The anti-inflammatory activities of pulvones A and C were further evaluated due to the better inhibitory activity on nitric oxide production in LPS-stimulated RAW264.7 cells and no obvious cytotoxicity to RAW264.7 cells. Western blot showed that pulvones A significantly decreased the levels of iNOS and COX-2 proteins and pulvones C only decreased the level of iNOS protein. ELISA analysis demonstrated that pulvones A inhibited the production of both interleukin-6 (IL-6) and IL-1ß while pulvones C showed better suppression effect on IL-1ß production in LPS-stimulated RAW264.7 cells. Then, their potential inhibitory effects on NF-κB pathway were tested in LPS-stimulated RAW264.7 cells. Immunofluorescence and western blot assay showed that pulvones A and C reduced the nuclear translocation of NF-κB(p65) and interrupted IκB phosphorylation. The ADP-Glo™ kinase assay showed pulvones A and C could directedly inhibit the IKKß kinase activity with the inhibitory rate of 40%, which were also verified by docking study. Collectively, these results suggested that pulvones A and C's anti-inflammatory effects were relevant to the interruption of NF-κB activation by inhibiting IKKß kinase.

Identification, characterization and HPLC quantification of formulation-related impurities of honokiol, an antitumor natural drug candidate in clinical trials.

Natural products and their derivatives have historically been invaluable as a source of therapeutic agents. Honokiol, as a well-known natural product in Chinese herbal medicine Houpu, is finally being studied in a Phase I clinical trial (CTR20170822) in patients with Advanced Non-Small Cell Lung Cancer (NSCLS) in China this year. During the honokiol liposome formulation process, five major impurities were present in the range of 0.05-0.1% based on the HPLC analysis. These five major impurities were obtained from the forced degradation product of honokiol through countercurrent chromatography and prep-HPLC. The structure were elucidated with 1H NMR, 13C NMR, 2D NMR and MS spectral data. The proposed HPLC method was validated for specificity, linearity (concentration range 0.01-1.62, 0.003-0.96, 0.05-7.98, 0.04-6.52, 0.03-5.18 µg/ml for impurities I-V respectively, R2 > 0.9988), accuracy (99.11-100.67%), precision (CV < 1.6%), and sensitivity (LOD 3.3, 0.1, 16.7, 13.3, 10.0 ng/ml for impurities I-V respectively). The validated method was employed in the further study of the honokiol drug substance.

[The research of the progesterone and estrogen secretion level and the FSHR, LHR gene quantitative in granular cells cultured in vitro in different time of laying hens].

OBJECTIVE: To research the hormone secretion levels of progesterone and estrogen and the gene expression levels of two go-nadotropin receptors follicle stimulating hormone receptor (FSHR) and luteinizing hormone receptor (LHR) in granular cells of laying hen, and the effect of culture time on the levels of hormone secretion and expression of related receptor gene in granulosa cells was inferred. METHODS: The experiment using the method of cells culture in vitro, the granular cells supernatants of hens were collected at 0 h, 24 h, 48 h, 72 h, 96 h, the progesterone and estrogen concentrations in cell supernatants were determined by ELISA kits, and detected the expression of FSHR and LHR gene in granular cells by real-time fluorescent quantitative PCR. RESULTS: The results showed that the progesterone and estrogen secretion reduced in the early culture of 0 h~48 h(P < 0.05), with the culture time increases to 72 h, the secretion of two hormones began to in-crease, and reaching the level of the initial level of culture. When the cells were cultured to 96 h, the rogesterone and estrogen secretion was reduced again. The lower levels of FSHR and LHR mRNA expression in granular cells appeared with the increase of culture time, compared with the group of cell culture to 0 h, the mRNA expression levels of each groups reduced obviously(P < 0.05). CONCLUSIONS: The amount of progesterone and estrogen in the cultured follicular granulosa cells decreased with the increase of in vitro culture time, and then increased. This might be related to the growth state of cells cultured in vitro. But on the whole, with the extension of the training time, the secretion of proges-terone and estrogen in the cells decreased. This may be related to the decreased expression of the FSHR and LHR genes in the two go-nadotropin receptors.

Adiabatic circular polarizer based on chiral fiber grating.

Based on the adiabatic coupling principle, a new scheme of a broadband circular polarizer formed by twisting a high-birefringence (Hi-Bi) fiber with a slowly varying twist rate is proposed. The conditions of adiabatic coupling for the adiabatic polarizer are first identified through analytical derivations. These conditions are easily realized by choosing a reasonable variation of the twist rate. Moreover, the bandwidth of the polarizer is able to be directly determined by the twist rates at the two ends. Finally, the broadband characteristics of the polarizer are demonstrated by simulations. It is also shown that the performance of the polarizer can be remarkably improved by accomplishing a multi-mode phase-matching along the grating or by using of the couplings of the core mode to lossy modes.

New insight into quasi leaky mode approximations for unified coupled-mode analysis.

To facilitate the analysis of radiation mode couplings, quasi leaky mode approximations were utilized in coupled-mode analysis. The key to effectively and accurately apply this approach is how to well approximate radiation modes by the quasi leaky modes in an equivalent closed waveguide model. In this paper, the principle, applicability and accuracy of the approximations are demonstrated, and the detailed implementation is also suggested by applying a unified coupled-mode analysis to fiber gratings. First of all, based on a thorough study on the characteristics of the complex modes, for the first time, quasi leaky modes are classified into guided-mode-like inner-cladding and radiation-mode-like outer-cladding leaky modes so as to explicitly establish equivalence relationships between the discrete leaky modes and the continuous radiation modes. With this new insight, the whole analysis process especially for some of the practically tricky issues such as the criteria for developing the proper equivalent waveguide model and the subsequent mode expansion basis are better understood and easier to be dealt with for different problems where radiation modes come into play. Moreover, as essential preconditions to extend the conventional coupled mode analysis to the present unified one, the couplings between the guided core mode and a leaky mode are studied in a systematic and consistent manner. An intuitive and then a deep understanding on the roles of complex modes on mode coupling and power exchanging are thus gained for further simulations. Lastly, the transmission spectra of fiber gratings with different surrounding indices are simulated. The simulated results agree well with those obtained theoretically and experimentally in the literatures, which strongly validate the principle of quasi leaky mode approximations and its implementation on the unified coupled-mode analysis expounded in this paper.