Solvent-Dependent Emissions Properties of a Model Aurone Enable Use in Biological Applications.

Fluorophores are powerful visualization tools and the development of novel small organic fluorophores are in great demand. Small organic fluorophores have been derived from the aurone skeleton, 2-benzylidenebenzofuran-3(2H)-one. In this study, we have utilized a model aurone derivative with a methoxy group at the 3' position and a hydroxyl group at the 4' position, termed vanillin aurone, to develop a foundational understanding of structural factors impacting aurone fluorescence properties. The fluorescent behaviors of the model aurone were characterized in solvent environments differing in relative polarity and dielectric constant. These data suggested that hydrogen bonding or electrostatic interactions between excited state aurone and solvent directly impact emissions properties such as peak emission wavelength, emission intensity, and Stokes shift. Time-dependent Density Functional Theory (TD-DFT) model calculations suggest that quenched aurone emissions observed in water are a consequence of stabilization of a twisted excited state conformation that disrupts conjugation. In contrast, the calculations indicate that low polarity solvents such as toluene or acetone stabilize a brightly fluorescent planar state. Based on this, additional experiments were performed to demonstrate use as a turn-on probe in an aqueous environment in response to conditions leading to planar excited state stabilization. Vanillin aurone was observed to bind to a model ATP binding protein, YME1L, leading to enhanced emissions intensities with a dissociation equilibrium constant equal to ~ 30 µM. Separately, the aurone was observed to be cell permeable with significant toxicity at doses exceeding 6.25 µM. Taken together, these results suggest that aurones may be broadly useful as turn-on probes in aqueous environments that promote either a change in relative solvent polarity or through direct stabilization of a planar excited state through macromolecular binding.

Aurone-derived 1,2,3-triazoles as potential fluorescence molecules in vitro.

Aurones are a class of well-studied natural compounds primarily responsible for the yellow pigment in flowering plants and have been shown to have fluorescent properties as well as beneficial biological effects. Traditionally, aurones can be easily synthesized through a Knoevenagel condensation of benzofuranones with arylaldehydes. Recently, Kafle et al. unexpectedly synthesized a new aurone derivative containing a 1,2,3-triazole within its backbone. Since, 1,2,3-triazole containing structures have been shown to be useful as fluorophores with large Stokes shifts, we hypothesized that these new aurone-derived triazole compounds (ATs) could be utilized as potential fluorophores. Here we describe a newly-synthesized fluorescent compound which has potential for use as a live-cell probe, having a large Stokes shift of 118.3 ± 1.01 nm in phosphate-buffered saline with the benefit of increased fluorescence in protic environments, which is uncommon in aurone-derived fluorophores.

The SWI/SNF ATPase BRG1 facilitates multiple pro-tumorigenic gene expression programs in SMARCB1-deficient cancer cells.

Malignant rhabdoid tumor (MRT) is driven by the loss of the SNF5 subunit of the SWI/SNF chromatin remodeling complex and then thought to be maintained by residual SWI/SNF (rSWI/SNF) complexes that remain present in the absence of SNF5. rSWI/SNF subunits colocalize extensively on chromatin with the transcription factor MYC, an oncogene identified as a novel driver of MRT. Currently, the role of rSWI/SNF in modulating MYC activity has neither been delineated nor has a direct link between rSWI/SNF and other oncogenes been uncovered. Here, we expose the connection between rSWI/SNF and oncogenic processes using a well-characterized chemical degrader to deplete the SWI/SNF ATPase, BRG1. Using a combination of gene expression and chromatin accessibility assays we show that rSWI/SNF complexes facilitate MYC target gene expression. We also find that rSWI/SNF maintains open chromatin at sites associated with hallmark cancer genes linked to the AP-1 transcription factor, suggesting that AP-1 may drive oncogenesis in MRT. Interestingly, changes in MYC target gene expression are not overtly connected to the chromatin remodeling function of rSWI/SNF, revealing multiple mechanisms used by rSWI/SNF to control transcription. This work provides an understanding of how residual SWI/SNF complexes may converge on multiple oncogenic processes when normal SWI/SNF function is impaired.

Panax quinquefolius (North American Ginseng) Polysaccharides as Immunomodulators: Current Research Status and Future Directions.

Panax quinquefolius (North American ginseng, NAG) is a popular medicinal plant used widely in traditional medicine. NAG products are currently available in various forms such as roots, extracts, nutraceuticals, dietary supplements, energy drinks, etc. NAG polysaccharides are recognized as one of the major bioactive ingredients. However, most NAG reviews are focused on ginsenosides with little information on polysaccharides. NAG polysaccharides have demonstrated a therapeutic activity in numerous studies, in which many of the bioactivities involve regulation of the immune response. The purpose of this review is to summarize the structural features and the immunomodulatory properties of crude, partially purified, and pure polysaccharides isolated from NAG. Receptors of the innate immune system that potentially bind to NAG polysaccharides and the respective signal transduction pathways initiated by these compounds are discussed. Major challenges, recent innovations, and future directions in NAG polysaccharide research are also summarized.

An acidic polysaccharide (AGC3) isolated from North American ginseng (Panax quinquefolius) suspension culture as a potential immunomodulatory nutraceutical.

Polysaccharides isolated from Panax quinquefolius roots are widely used as nutraceuticals due to their immunomodulatory properties. Despite their popularity, several challenges exist in isolating ginseng root polysaccharides such as batch-to-batch structural inconsistencies and bacterial endotoxin contamination. A plant tissue culture-based platform offers a potential solution to isolate natural polysaccharide fractions with consistent chemical characteristics and reduced endotoxin content. In this study, an acidic polysaccharide fraction (AGC3) with immunomodulatory properties was isolated from Panax quinquefolius suspension cultures. The heterogeneous fraction (molecular weight: 4.81 and 32.14 kDa), purified by anion exchange chromatography, was predominantly composed of galactose (>60%) along with the presence of rhamnose, arabinose, glucose, glucuronic acid and galacturonic acid. The major glycosidic linkages were found to be t-Galp (47.7%), 4-Galp (15.6%), 2,4-Rhap (8.1%), 6-Galp (8.1%) and 4-GalAp (6.8%). Structural analyses indicated the presence of a pectic rhamnogalacturonan I polysaccharide in AGC3. AGC3 significantly (p < 0.05) stimulated RAW 264.7 murine macrophage cells and primary murine splenocytes by enhancing the production of several immunomodulatory mediators such as IL-6, TNF-α, GM-CSF and MCP-1. The results also indicated the putative roles of NF-κB (p65/RelA) and MAPK (p38) signaling pathways in the immunostimulatory response. Additionally, AGC3 induced murine splenocyte proliferation, another major indicator of immunostimulation. Overall, AGC3 has the potential to be used as an immunostimulatory nutraceutical.

Panax quinquefolius (North American ginseng) cell suspension culture as a source of bioactive polysaccharides: Immunostimulatory activity and characterization of a neutral polysaccharide AGC1.

In this study, we propose the use of a plant tissue culture-based system for the production of polysaccharides with consistent chemical characteristics and reduced endotoxin content. Polysaccharides were isolated from suspension cultures of Panax quinquefolius (American ginseng), a widely used medicinal herb. A neutral fraction, AGC1, purified by anion exchange and size exclusion chromatography, displayed immunostimulatory activity in vitro and ex vivo. AGC1 (average molecular weight: 5.2kDa) was predominantly composed of galactose (>60%) along with the presence of several other neutral sugars such as arabinose, xylose, glucose, mannose and rhamnose in minor amounts. The major glycosidic linkages were found to be 3-Galp (48.5%), 3,6-Galp (10.2%), t-Galp (5.2%), 6-Galp (4.4%), 4-Glcp (5.7%), 4-Arap/5-Araf (4.0%) and t-Araf (4.5%). AGC1 significantly (p<0.05) stimulated the expression of a range of proinflammatory mediators in RAW 264.7 murine macrophages such as IL-6, TNF-α, MCP-1 and GM-CSF. Additionally, AGC1 treatment of RAW 264.7 cells stimulated NOS2 gene expression, leading to increased levels of iNOS and downstream NO. Consistent with this, AGC1 was able to act as an immunostimulant in primary murine splenocytes, enhancing cell proliferation, as well as NO and TNF-α production. Our results also indicate the partial role of NF-κB pathway in the immunostimulatory response.

Isolation, structure elucidation, and immunostimulatory activity of polysaccharide fractions from Boswellia carterii frankincense resin.

Frankincense has a long history in religious, cultural, and medicinal use. In this study polysaccharides were extracted from frankincense from Boswellia carterii. The polysaccharides were purified by anion exchange chromatography on a DEAE-Sepharose Fast Flow 16/10 FPLC column. Six fractions were obtained and the three most active immunomodulatory fractions were further purified by size exclusion chromatography on a Superdex-200 column. The composition showed the monosaccharides present were predominantly galactose, arabinose, and glucuronic acid along with small amounts of rhamnose and glucose. The monosaccharide composition and glycosyl linkage analysis revealed the polysaccharides belong to the type II arabinogalactans. Fourier-transform infrared spectroscopy and bicinchoninic acid assay showed that the amount of protein in the samples was <1 wt%. One-dimensional 1H NMR were consistent with high molecular weight compounds. The monosaccharides were primarily in the ß conformation. The three fractions exhibited an immunostimulatory effect on RAW 264.7 murine macrophage cells. The most active immunostimulatory fraction FA2, stimulated a range of pro-inflammatory mediators including iNOS, NO, TNF-α, and IL-6 in RAW 264.7 cells. The fractions were effective in proliferating primary murine splenocytes. The results indicate that the polysaccharides isolated from frankincense have the potential to be used as an immunological stimulant or nutraceutical.

Nodular regenerative hyperplasia of the liver secondary to azathioprine in a patient with inflammatory bowel disease.

A case of dramatic portal hypertension with massive ascites and splenomegaly is described in a patient with inflammatory bowel disease receiving azathioprine therapy. Liver biopsy confirmed the subtle changes of nodular regenerative hyperplasia and the patient recovered following withdrawal of the azathioprine and commencement of spironolactone. Thrombocytopaenia is an early clue to azathioprine-induced nodular regenerative hyperplasia of the liver.