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Current treatments for osteoarthritis (OA) often fail to address the underlying pathophysiology and may have systemic side effects, particularly associated with long-term use of non-steroidal anti-inflammatory drugs (NSAIDs). Thus, researchers are currently directing their efforts toward innovative polymer-drug combinations, such as mixtures of hyaluronic acid viscoelastic hydrogels and NSAIDs like diclofenac, to ensure sustained release of the NSAID within the joint following intra-articular injection. However, the progress of novel injectable therapies for OA is hindered by the absence of preclinical models that accurately represent the pathology of the disease. The uBeat® MultiCompress platform is here presented as a novel approach for studying anti-OA injectable therapeutics on human mechanically-damaged OA cartilage microtissues, in a physiologically relevant environment. This platform can accommodate injectable therapeutic formulations and is successfully tested with SYN321, a novel diclofenac-sodium hyaluronate conjugate under development as a treatment for knee OA. Results indicate the platform's effectiveness in evaluating therapeutic potential, showing downregulation of inflammatory markers and reduction in matrix degradation in OA cartilage micro-tissues treated with SYN321. The uBeat® MultiCompress platform thus represents a valuable tool for OA research, offering a bridge between traditional in vitro studies and potential clinical applications, with implications for future drug discovery., (© 2024 The Author(s). Advanced Healthcare Materials published by Wiley‐VCH GmbH.)

A high number of varieties from corn (Zea mays L.) have been consumed for long time all over the world, however pigmented varieties are recently gaining renewed attention due to their beneficial effects and polyphenolic content. The natural lack of gluten makes corn suitable for consumption by celiac population, who need to control their inflammatory state through an appropriate gluten-free diet. The biological effects of polyphenols from pigmented corn are poorly investigated in the context of celiac disease. In this work, we analyzed through HPLC-DAD the phenolic composition of two Italian purple and red varieties ("Scagliolo Rosso" and "Rostrato di Rovetta", respectively) comparing their effects in human intestinal epithelial cells (CaCo-2 cells). The possible impact of gastro-intestinal digestion following oral consumption was assessed as well. The phenolic profile showed the presence of phenolic acids in both varieties, while anthocyanins were identified in Scagliolo Rosso only. After simulated digestion, the level of polyphenols did not significantly change and paralleled with an increased scavenging activity. In CaCo-2 cells, stimulated by a proinflammatory cocktail containing gliadin-derived peptides (IL-1β, IFN-γ, digested gliadin), pigmented corn extracts inhibited the release of CXCL-10 and sICAM-1, with mechanisms partially ascribed to NF-κB impairment. At the same concentration (200 μg/mL), ROS production and catalase depletion were reverted through Nrf-2-independent mechanisms. Our data suggest that polyphenols from pigmented corns might help in controlling the inflammatory and oxidative state of people with celiac disease at intestinal level, at concentrations potentially achievable through a gluten-free diet., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Castanea sativa Mill. ( C. sativa ) processing and pruning generate several by-products, including leaves, burs, and shells (inner and outer teguments), which are considered an important source of high-value phytochemicals. Ellagitannins from C. sativa leaf extracts have been described to impair H. pylori viability and inflammation in gastric cells. Furthermore, chestnut shells showed an important anti-inflammatory effect in gastric epithelial cells. Dietary polyphenols, including tannins, have been reported to interfere with targets of inflammation, including the nuclear factor κB (NF-κB). A promising role as a further therapeutical target for gut disorders has been recently proposed for the regulatory subunit of hypoxia-inducible factor (HIF-1α), as a potential stabilizer of intestinal barrier integrity. Therefore, the main objective of this work is the chemical characterization of several chestnut by-products (bud, spiny bur, wood, pericarp and episperm), together with the exploitation of their anti-inflammatory properties in intestinal cells, scavenging capacity, and stability following gastrointestinal digestion. The chemical characterization confirmed the presence of bioactive polyphenols in the extracts, including ellagitannins. In CaCo-2 cells stimulated by an IL-1β-IFN-γ cocktail, nearly all chestnut by-products (50 µg/mL) inhibited the release of proinflammatory mediators (CXCL-10, IL-8, MCP-1, ICAM), along with the NF-κB-driven transcription, and induced the HRE-driven transcription. The stability of the most promising extracts, identified through PCA and cluster analysis, was addressed by in vitro gastrointestinal digestion. Despite the significant reduction in total polyphenol index of chestnut bud and wood after gastric and intestinal digestion, the activity of these extracts on both scavenging and anti-inflammatory parameters remained promising. These data contribute to exploit the potential of chestnut by-products as sources of dietary polyphenols with anti-inflammatory properties at the intestinal level. Moreover, this study could represent an important step to encourage the recycling and valorization of chestnut by-products, promoting the circular economy and reducing the environmental impact related to the management of agriculture waste.

Vicia faba L. is a leguminous plant with seeds rich in nutritional compounds, such as polyphenols and L-dopa, a dopamine precursor and first-line treatment for Parkinson's symptoms. Recently, its by-products have been revalued as a sustainable source of bioactive compounds. In this study, aqueous extracts of Lucan broad bean pod valves (BPs) were characterized to evaluate their potential use as adjuvants in severe Parkinson's disease. L-dopa content, quantified by LC-UV, was much higher in BPs than in seeds (28.65 mg/g dw compared to 0.76 mg/g dw). In addition, vicine and convicine, the metabolites responsible for favism, were not detected in pods. LC-ESI/LTQ-Orbitrap/MS 2 allowed the identification of the major polyphenolic compounds, including quercetin and catechin equivalents, that could ensure neuroprotection in Parkinson's disease. ESI(±)-FT-ICR MS was used to build 2D van Krevelen diagrams; polyphenolic compounds and carbohydrates were the most representative classes. The neuroprotective activity of the extracts after MPP + -induced neurotoxicity in SH-SY5Y cells was also investigated. BP extracts were more effective than synthetic L-dopa, even at concentrations up to 100 µg/mL, due to the occurrence of antioxidants able to prevent oxidative stress. The stability and antioxidant component of the extracts were then emphasized by using naturally acidic solutions of Punica granatum L., Ribes rubrum L., and gooseberry ( Phyllanthus emblica L.) as extraction solvents.

Sulforaphane is considered the bioactive metabolite of glucoraphanin after dietary consumption of broccoli sprouts. Although both molecules pass through the gut lumen to the large intestine in stable form, their biological impact on the first intestinal tract is poorly described. In celiac patients, the function of the small intestine is affected by celiac disease (CD), whose severe outcomes are controlled by gluten-free dietary protocols. Nevertheless, pathological signs of inflammation and oxidative stress may persist. The aim of this study was to compare the biological activity of sulforaphane with its precursor glucoraphanin in a cellular model of gliadin-induced inflammation. Human intestinal epithelial cells (CaCo-2) were stimulated with a pro-inflammatory combination of cytokines (IFN-γ, IL-1β) and in-vitro-digested gliadin, while oxidative stress was induced by H 2 O 2 . LC-MS/MS analysis confirmed that sulforaphane from broccoli sprouts was stable after simulated gastrointestinal digestion. It inhibited the release of all chemokines selected as inflammatory read-outs, with a more potent effect against MCP-1 (IC 50 = 7.81 µM). On the contrary, glucoraphanin (50 µM) was inactive. The molecules were unable to counteract the oxidative damage to DNA (γ-H2AX) and catalase levels; however, the activity of NF-κB and Nrf-2 was modulated by both molecules. The impact on epithelial permeability (TEER) was also evaluated in a Transwell ® model. In the context of a pro-inflammatory combination including gliadin, TEER values were recovered by neither sulforaphane nor glucoraphanin. Conversely, in the context of co-culture with activated macrophages (THP-1), sulforaphane inhibited the release of MCP-1 (IC 50 = 20.60 µM) and IL-1β (IC 50 = 1.50 µM) only, but both molecules restored epithelial integrity at 50 µM. Our work suggests that glucoraphanin should not merely be considered as just an inert precursor at the small intestine level, thus suggesting a potential interest in the framework of CD. Its biological activity might imply, at least in part, molecular mechanisms different from sulforaphane.

Despite millennia of therapeutic plant use, deliberate exploitation of Cannabis 's diverse biomedical potential has only recently gained attention. Bioactivity studies focus mainly on cannabidiol (CBD) and tetrahydrocannabinol (THC) with limited information about the broader cannabinome's "minor phytocannabinoids". In this context, our research targeted the synthesis of minor cannabinoids containing a lateral chain with 3 or 4 carbon atoms, focusing on cannabigerol (CBG) and cannabichromene (CBC) analogues. Using known and innovative strategies, we achieved the synthesis of 11 C3 and C4 analogues, five of which were inhibitors of skin inflammation, with the CBG-C4 ester derivative emerging as the most potent compound.

Objectives: Neural stem/progenitor cells derived from olfactory neuroepithelium (hereafter olfactory neural stem/progenitor cells, ONSPCs) are emerging as a potential tool in the exploration of psychiatric disorders. The present study intended to assess whether ONSPCs could help discern individuals with schizophrenia (SZ) from non-schizophrenic (NS) subjects by exploring specific cellular and molecular features., Methods: ONSPCs were collected from 19 in-patients diagnosed with SZ and 31 NS individuals and propagated in basal medium. Mitochondrial ATP production, expression of β-catenin and cell proliferation, which are described to be altered in SZ, were examined in freshly isolated or newly thawed ONSPCs after a few culture passages., Results: SZ-ONSPCs exhibited a lower mitochondrial ATP production and insensitivity to agents capable of positively or negatively affecting β-catenin expression with respect to NS-ONSPCs. As to proliferation, it declined in SZ-ONSPCs as the number of culture passages increased compared to a steady level of growth shown by NS-ONSPCs., Conclusions: The ease and safety of sample collection as well as the differences observed between NS- and SZ-ONSPCs, may lay the groundwork for a new approach to obtain biological material from a large number of living individuals and gain a better understanding of the mechanisms underlying SZ pathophysiology.

Objectives: The sweet chestnut Castanea sativa Mill. is the only native Castanea species in Europe, and it is a tree of high economic value that provides appreciated fruits and valuable wood. In this study, we assembled a high-quality nuclear genome of the ancient Italian chestnut variety 'Marrone di Chiusa Pesio' using a combination of Oxford Nanopore Technologies long reads, whole-genome and Omni-C Illumina short reads., Data Description: The genome was assembled into 238 scaffolds with an N50 size of 21.8 Mb and an N80 size of 7.1 Mb for a total assembled sequence of 750 Mb. The BUSCO assessment revealed that 98.6% of the genome matched the embryophyte dataset, highlighting good completeness of the genetic space. After chromosome-level scaffolding, 12 chromosomes with a total length of 715.8 and 713.0 Mb were constructed for haplotype 1 and haplotype 2, respectively. The repetitive elements represented 37.3% and 37.4% of the total assembled genome in haplotype 1 and haplotype 2, respectively. A total of 57,653 and 58,146 genes were predicted in the two haplotypes, and approximately 73% of the genes were functionally annotated using the EggNOG-mapper. The assembled genome will be a valuable resource and reference for future chestnut breeding and genetic improvement., (© 2024. The Author(s).)

Khat leaves, indigenous to eastern Africa, have been chewed for centuries for their stimulant effects, attributed to alkaloids such as cathinone and cathine. Although associated with gastric disorders like gastritis and gastro-oesophageal reflux disease, the underlying molecular mechanisms remain unclear. This study aimed to examine the morpho-anatomy of khat leaves using light microscopy and histochemistry and to assess the effects of leaf extracts and alkaloids on human gastric epithelial cells (GES-1). The study identified specific cells in the palisade-spongy transition zone as storage sites for psychoactive alkaloids. Leaf extracts were prepared by mimicking the chewing process, including a prolonged salivary phase followed by a gastric phase. Cytotoxicity and cell viability were evaluated using LDH and MTT assays, respectively. Additionally, the impact on IL-8 secretion, a key chemokine in gastric inflammation, was analysed under normal and TNF-α-stimulated conditions. The results showed no increase in cytotoxicity up to 250 µg/mL. However, there was a significant decrease in cell metabolism and a reduction in both basal and TNF-α-induced IL-8 secretion, but cathinone and cathine were inactive. These findings suggest that khat may not directly cause the gastric issues reported in the literature, which would rather be attributed to other confounding factors, highlighting the need for further research to clarify its biological impacts.

Acne vulgaris is a prevalent skin disorder affecting many young individuals, marked by keratinization, inflammation, seborrhea, and colonization by Cutibacterium acnes ( C. acnes ). Ellagitannins, known for their antibacterial and anti-inflammatory properties, have not been widely studied for their anti-acne effects. Chestnut ( Castanea sativa Mill., C. sativa ), a rich ellagitannin source, including castalagin whose acne-related bioactivity was previously unexplored, was investigated in this study. The research assessed the effect of C. sativa leaf extract and castalagin on human keratinocytes (HaCaT) infected with C. acnes , finding that both inhibited IL-8 and IL-6 release at concentrations below 25 μg/mL. The action mechanism was linked to NF-κB inhibition, without AP-1 involvement. Furthermore, the extract displayed anti-biofilm properties and reduced CK-10 expression, indicating a potential role in mitigating inflammation, bacterial colonization, and keratosis. Castalagin's bioactivity mirrored the extract's effects, notably in IL-8 inhibition, NF-κB inhibition, and biofilm formation at low μM levels. Other polyphenols, such as flavonol glycosides identified via LC-MS, might also contribute to the extract's biological activities. This study is the first to explore ellagitannins' potential in treating acne, offering insights for developing chestnut-based anti-acne treatments pending future in vivo studies.

(1) Background: Within the framework of the European Interreg Italy-Switzerland B-ICE & Heritage project (2018-2022), this study originated from a three-year ethnobotanical survey in Valmalenco (Sondrio, Italy). Following a preliminary work published by our group, this research further explored the folk therapeutic use of Achillea erba-rotta subsp. moschata (Wulfen) I.Richardson (Asteraceae) for dyspepsia disorders, specifically its anti-inflammatory potential at a gastrointestinal level. (2) Methods: Semi-structured interviews were performed. The bitter taste was investigated through molecular docking software (PLANTS, GOLD), while the anti-inflammatory activity of the hydroethanolic extract, infusion, and decoction was evaluated based on the release of IL-8 and IL-6 after treatment with TNFα or Helicobacter pylori . The minimum inhibitory concentration and bacterial adhesion on the gastric epithelium were evaluated. (3) Results: In total, 401 respondents were interviewed. Molecular docking highlighted di-caffeoylquinic acids as the main compounds responsible for the interaction with bitter taste receptors. The moderate inhibition of IL-6 and IL-8 release was recorded, while, in the co-culture with H. pylori , stronger anti-inflammatory potential was expressed (29-45 μg/mL). The concentration-dependent inhibition of H. pylori growth was recorded (MIC = 100 μg/mL), with a significant anti-adhesive effect. (4) Conclusions: Confirming the folk tradition, the study emphasizes the species' potentiality for dyspepsia disorders. Future studies are needed to identify the components mostly responsible for the biological effects.

Cistus spp. have been traditionally used for inflammatory and infectious disorders, including gastrointestinal ailments, in the Mediterranean area. Among them, Cistus × incanus L. is one of the most frequently cited species in the literature for a variety of biological activities which include inflammatory diseases. Cistus spp. aerial parts are rich in polyphenols such as condensed and hydrolysable tannins, procyanidins, and flavonoids, which show gastroprotective activities. The purpose of the present study is to investigate the biological activities of a hydroalcoholic extract from Cistus × incanus L. aerial parts in gastric epithelial cells (GES-1) infected with H. pylori . The extracts inhibited IL-8 and NF-κB induced by H. pylori and showed antibacterial activity after simulated digestion. Since our previous paper reported interesting results on the ability of Castanea sativa Mill. leaf extract to decrease inflammatory conditions in H. pylori -infected gastric cells, the combination of Castanea sativa and Cistus × incanus extracts was also investigated, showing strong anti-inflammatory activity and inhibition of bacterial adhesion. This association of botanicals is proposed herein as a novel food supplement capable of counteracting gastric inflammatory conditions.

Helicobacter pylori is a leading cause of chronic gastric inflammation, generally associated with gastritis and adenocarcinoma. Activation of the NF-κB pathway mainly contributes to the inflammatory phenotype observed in H. pylori infection in humans and experimental models. Since the gastric epithelium undergoes rapid turnover, inflammation and pathogenicity of H. pylori result from early phase and chronically activated pathways. In the present study we investigated the early host response to H. pylori in non-tumoral human gastric epithelial cells (GES-1). To dissect the pathogen-specific mechanisms we also examined the response to tumor necrosis factor (TNF), a prototypical cytokine. By analyzing the activation state of NF-κB signaling, cytokine expression and secretion, and the transcriptome, we found that the inflammatory response of GES-1 cells to H. pylori and TNF results from activation of multiple pathways and transcription factors, e.g., NF-κB and CCAAT/enhancer-binding proteins (CEBPs). By comparing the transcriptomic profiles, we found that H. pylori infection induces a less potent inflammatory response than TNF but affects gene transcription to a greater extent by specifically inducing transcription factors such as CEBPβ and numerous zinc finger proteins. Our study provides insights on the cellular pathways modulated by H. pylori in non-tumoral human gastric cells unveiling new potential targets.

An analytical method based on ultrasound assisted extraction (UAE) and liquid chromatography coupled to electrospray tandem mass spectrometry (LC-ESI/MS/MS) was validated and applied for determining L-dopa in four ecotypes of Fagioli di Sarconi beans (Phaseolus vulgaris L.), marked with the European label PGI (Protected Geographical Indication). The selectivity of the proposed method was ensured by the specific fragmentation of the analyte. Simple isocratic chromatographic conditions and mass spectrometric detection in multiple reaction monitoring (MRM) acquisition mode were used for sensitive quantification. The LC-ESI/MS/MS method was validated within a linear range of 0.001-5.000 μg/mL. Values of 0.4 and 1.1 ng/mL were obtained for the limits of detection and quantification, respectively. The repeatability, inter-day precision, and recovery values ranges were 0.6%-4.5%, 5.4%-9.9%, and 83%-93%, respectively. Fresh and dried beans, as well as pods, cultivated exclusively with organic methods avoiding any synthetic fertilizers and pesticides were analyzed showing an L-dopa content ranging from 0.020 ± 0.005 to 2.34 ± 0.05 μg/g dry weight., (© 2023 John Wiley & Sons Ltd.)

(1) Background: Filipendula ulmaria (L.) Maxim. (Rosaceae) (meadowsweet) is widely used in phytotherapy against inflammatory diseases. However, its active constituents are not exactly known. Moreover, it contains many constituents, such as flavonoid glycosides, which are not absorbed, but metabolized in the colon by gut microbiota, producing potentially active metabolites that can be absorbed. The aim of this study was to characterize the active constituents or metabolites. (2) Methods: A F. ulmaria extract was processed in an in vitro gastrointestinal biotransformation model, and the metabolites were characterized using UHPLC-ESI-QTOF-MS analysis. In vitro anti-inflammatory activity was evaluated by testing the inhibition of NF-κB activation, COX-1 and COX-2 enzyme inhibition. (3) Results: The simulation of gastrointestinal biotransformation showed a decrease in the relative abundance of glycosylated flavonoids such as rutin, spiraeoside and isoquercitrin in the colon compartment, and an increase in aglycons such as quercetin, apigenin, naringenin and kaempferol. The genuine as well as the metabolized extract showed a better inhibition of the COX-1 enzyme as compared to COX-2. A mix of aglycons present after biotransformation showed a significant inhibition of COX-1. (4) Conclusions: The anti-inflammatory activity of F. ulmaria may be explained by an additive or synergistic effect of genuine constituents and metabolites.

Helicobacter pylori ( H. pylori ) is an etiologic factor of peptic ulcer disease and gastric cancer. Virulent strains of H. pylori are correlated with the severity of gastritis, due to NF-κB activation and IL-8 expression at the epithelial level. Ellagitannins have been documented for antibacterial and anti-inflammatory activities, thus suggesting their potential use in gastritis. Recently, several authors, including our group, demonstrated that tannin-rich extracts from chestnut byproducts, at present considered agricultural waste, display promising biological activities. In this work, we detected high levels of polyphenols in hydroalcoholic extracts from chestnut leaves ( Castanea sativa L.). Among polyphenols, the ellagitannin isomers castalagin and vescalagin (about 1% w/w of dry extract) were identified as potential bioactive compounds. In GES-1 cells infected by H. pylori , leaf extract and pure ellagitannins inhibited IL-8 release (IC 50 ≈ 28 µg/mL and 11 µM, respectively). Mechanistically, the anti-inflammatory activity was partly due to attenuation of NF-κB signaling. Moreover, the extract and pure ellagitannins reduced bacterial growth and cell adhesion. A simulation of the gastric digestion suggested that the bioactivity might be maintained after oral administration. At the transcriptional level, castalagin downregulated genes involved in inflammatory pathways (NF-κB and AP-1) and cell migration (Rho GTPase). To the best of our knowledge, this is the first investigation in which ellagitannins from plant extracts have demonstrated a potential role in the interaction among H. pylori and human gastric epithelium.

Dietary bioactive compounds from natural sources (e.g., herbal medicines, foods) are known to potentially suppress acute or chronic inflammation and promote the effectiveness of treatment to reduce the harmful effects of gastritis alone or in combination. In this regard, we have characterized four Cameroonian spice extracts, namely Aframomum citratum , Dichrostachys glomerata , Tetrapleura tetraptera , and Xylopia parviflora through reverse phase-high-performance liquid chromatography (RP-HPLC), ultra-performance liquid chromatography-electrospray ionization high-resolution mass spectrometry (UPLC-ESI-HRMS/MS), and Fourier transform infrared spectroscopic (FTIR) analyses and investigated their antioxidant and synergistic anti-inflammatory activities in human gastric adenocarcinoma (AGS) and gastric epithelial (GES-1) cells. The extracts showed a high amount of total phenolic (TPC: 150-290 mg gallic acid equivalents (GAE)/g of extract) and flavonoid content (TFC: 35-115 mg catechin equivalents (CE)/g of extract) with antioxidant properties in a cell-free system (1,1-Diphenyl-2-picryl-hydrazyl (DPPH) half maximal inhibitory concentration (IC 50 s) ≤ 45 µg/mL; 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) half maximal inhibitory concentration (IC 50 s) ≤ 29 µg/mL. The extracts in combination (MIX) exert a synergistic beneficial effect (combination index (CIs) < 1 and dose reduction index (DRIs) > 1) on inflammatory markers (interleukin (IL)-8 and -6 release, and nuclear factor kappa B (NF-κB) driven transcription) in human gastric epithelial cells, which may result from the presence of phenolic compounds (phenolic acids, flavonoids) or other compounds (protein, lipid, aromatic, and polysaccharide compounds) tentatively identified in the extracts. The general findings of the present study provide supporting evidence on the chemical composition of four Cameroonian dietary plants and their significant synergistic inhibitory activities on inflammatory markers of gastric epithelial cells.

Phloridzin is the most abundant polyphenolic compound in apple (Malus × domestica Borkh.), which results from the action of a key phloretin-specific UDP-2'-O-glucosyltransferase (MdPGT1). Here, we simultaneously assessed the effects of targeting MdPGT1 by conventional transgenesis and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9)-mediated genome editing. To this end, we conducted transcriptomic and metabolic analyses of MdPGT1 RNA interference knockdown and genome-edited lines. Knockdown lines exhibited characteristic impairment of plant growth and leaf morphology, whereas genome-edited lines exhibited normal growth despite reduced foliar phloridzin. RNA-sequencing analysis identified a common core of regulated genes, involved in phenylpropanoid and flavonoid pathways. However, we identified genes and processes differentially modulated in stunted and genome-edited lines, including key transcription factors and genes involved in phytohormone signalling. Therefore, we conducted a phytohormone profiling to obtain insight into their role in the phenotypes observed. We found that salicylic and jasmonic acid were increased in dwarf lines, whereas auxin and ABA showed no correlation with the growth phenotype. Furthermore, bioactive brassinosteroids were commonly up-regulated, whereas gibberellin GA 4 was distinctively altered, showing a sharp decrease in RNA interference knockdown lines. Expression analysis by reverse transcriptase-quantitative polymerase chain reaction expression analysis further confirmed transcriptional regulation of key factors involved in brassinosteroid and gibberellin interaction. These findings suggest that a differential modulation of phytohormones may be involved in the contrasting effects on growth following phloridzin reduction. The present study also illustrates how CRISPR/Cas9 genome editing can be applied to dissect the contribution of genes involved in phloridzin biosynthesis in apple., (© 2022 Society for Experimental Biology and John Wiley & Sons Ltd.)

Inflammation and oxidative stress are two mechanisms involved in the pathogenesis of celiac disease (CD). Since the direct effect of gliadin on the intestinal epithelia is less studied, the aims of this study were the development of a specific cellular model based on the use of gliadin as a pro-inflammatory stimulus and the evaluation of the potential antioxidant and anti-inflammatory properties of extracts from different black rice in the framework of CD. The rice extracts were in vitro digested, characterized in terms of phenolic compounds and antioxidant capacity, and tested on Caco-2 cells to investigate their inhibitory effect on Reactive Oxygen Species, the NF-κB transcription and the CXC chemokines (sICAM-1, IL-8, and CXCL-10). In addition, the role of the extracts in modulating the activation of epithelial cells in CD was confirmed by applying the K562(S) agglutination test. The black rice extracts showed inhibitory effects on the production of the oxidative and the inflammatory mediators considered, with particular reference to lymphocyte-attracting CXCL-10 both before and after digestion. The presence of anthocyanins and their digestion metabolites may account for the observed anti-inflammatory activity after in vitro digestion. This work provided preliminary data supporting the use of black rice as a healthy food or ingredient of food supplements for celiacs., Competing Interests: The authors declare no conflict of interest.

Plants rich in hydrolyzable tannins were traditionally used all over the world for a variety of chronic inflammatory disorders, including arthritis, colitis, and dermatitis. However, the knowledge of their immunological targets is still limited though fundamental for their rational use in phytotherapy. The recent advances regarding the pathogenesis of inflammatory-based diseases represent an opportunity to elucidate the pharmacological mechanism of plant-derived metabolites with immunomodulatory activity. This review collects recent articles regarding the role of hydrolyzable tannins and their gut metabolites in Th1, Th2, and Th17 inflammatory responses. In line with the traditional use, rheumatoid arthritis (RA), inflammatory bowel diseases (IBDs), psoriasis, atopic dermatitis (AD), and asthma were the most investigated diseases. A substantial body of in vivo studies suggests that, beside innate response, hydrolyzable tannins may reduce the levels of Th-derived cytokines, including IFN-γ, IL-17, and IL-4, following oral administration. The mode of action is multitarget and may involve the impairment of inflammatory transcription factors (NF-κB, NFAT, STAT), enzymes (MAPKs, COX-2, iNOS), and ion channels. However, their potential impact on pathways with renewed interest for inflammation, such as JAK/STAT, or the modulation of the gut microbiota demands dedicate studies.

L-Dopa (LD), a substance used medically in the treatment of Parkinson's disease, is found in several natural products, such as Vicia faba L., also known as broad beans. Due to its low chemical stability, LD analysis in plant matrices requires an appropriate optimization of the chosen analytical method to obtain reliable results. This work proposes an HPLC-UV method, validated according to EURACHEM guidelines as regards linearity, limits of detection and quantification, precision, accuracy, and matrix effect. The LD extraction was studied by evaluating its aqueous stability over 3 months. The best chromatographic conditions were found by systematically testing several C 18 stationary phases and acidic mobile phases. In addition, the assessment of the best storage treatment of Vicia faba L. broad beans able to preserve a high LD content was performed. The best LD determination conditions include sun-drying storage, extraction in HCl 0.1 M, chromatographic separation with a Discovery C 18 column, 250 × 4.6 mm, 5 µm particle size, and 99% formic acid 0.2% v / v and 1% methanol as the mobile phase. The optimized method proposed here overcomes the problems linked to LD stability and separation, thus contributing to the improvement of its analytical determination.

Hamamelis virginiana L. bark extract is a traditional remedy for skin affections, including atopic dermatitis/eczema (AD). Hamamelis preparations contain tannins, including hamamelitannin (HT), although their pharmacological role in AD is still unknown. This study aimed to study the rational for its topical use by considering the impact of crucial biomarkers on AD pathogenesis. A standardized extract (HVE) (0.5−125 μg/mL) was compared to hamamelitannin (HT), its main compound (0.5−5 μg/mL), in a model of human keratinocytes (HaCaTs), challenged with an AD-like cytokine milieu (TNF-α, IFN-γ, and IL-4). HVE inhibited the release of mediators involved in skin autoimmunity (IL-6 and IL-17C) and allergy (TSLP, IL-6, CCL26, and MMP-9) with a concentration-dependent fashion (IC50s < 25 μg/mL). The biological mechanism was ascribed, at least in part, to the impairment of the NF-κB-driven transcription. Moreover, HVE counteracted the proliferative effects of IL-4 and recovered K10, a marker of skin differentiation. Notably, HT showed activity on well-known targets of IL-4 pathway (CCL26, K10, cell proliferation). To the best of our knowledge, this work represents the first demonstration of the potential role of Hamamelis virginiana in the control of AD symptoms, such as itch and skin barrier impairment, supporting the relevance of the whole phytocomplex.

Cannabidiol (CBD) is a phytocannabinoid contained in the Cannabis sativa plant, devoid of psychotomimetic effects but with a broad-spectrum pharmacological activity. Because of its pharmacological profile and its ability to counteract the psychoactive Δ 9 -tetrahydrocannabinol (Δ9THC), CBD may be a potential treatment for several psychiatric and neurodegenerative disorders. In this study, we performed a dose-response evaluation of CBD modulatory effects on BDNF, a neurotrophin subserving pleiotropic effects on the brain, focusing on the cortico-striatal pathway for its unique role in the brain trafficking of BDNF. Male adult rats were exposed to single and repeated CBD treatments at different dosing regimen (5, 15, and 30 mg/kg), to investigate the rapid modulation of the neurotrophin (1 h after the single treatment) as well as a potential drug-free time point (24 h after the repeated treatment). We show here, for the first time, that CBD can be found in the rat brain and, specifically, in the medial prefrontal cortex (mPFC) following single or repeated exposure. In fact, we found that CBD is present in the mPFC of rats treated either acutely or repeatedly with the phytocannabinoid, with a clear dose-response profile. From a molecular standpoint, we found that single, but not repeated, CBD exposure upregulates BDNF in the mPFC, while the repeated exposure increased BDNF only in the striatum, with a slight decrease in the mPFC. Together, these data reveal a CBD dose-dependent and anatomically specific modulation of BDNF, which may be functionally relevant and may represent an added value for CBD as a supplement., Competing Interests: Mikael Sodergren is a Clinical Senior Lecturer at Imperial College and Chief Medical Officer at Curaleaf International, from whom he receives payments as a consultant. The other authors declare no conflicts of interests.

Cutibacterium acnes ( C. acnes ) is recognized as one of the main triggers of the cutaneous inflammatory response in acne vulgaris, a chronic skin disorder with a multifactorial origin. Witch hazel ( Hamamelis virginiana L.) is a plant widely used for skin inflammatory conditions, with some preliminary anti-inflammatory evidence on the skin, but lacking data on acne conditions. This study aimed to evaluate the effect of a glycolic extract from Hamamelis virginiana bark (HVE) versus C. acnes -induced inflammation in human keratinocytes (HaCaT). Phytochemical investigations of HVE identified hamamelitannin (HT) and proanthocyanidins as the most abundant compounds (respectively, 0.29% and 0.30% w / w extract ). HVE inhibited C. acnes -induced IL-6 release (IC 50 : 136.90 μg/mL), by partially impairing NF-κB activation; however, no antibacterial or antibiofilm activities were found. In addition, HVE showed greater anti-inflammatory activity when TNF-α was used as a proinflammatory stimulus (IC 50 of 38.93 μg/mL for IL-8 release), partially acting by antioxidant mechanisms, as shown for VEGF inhibition. The effects of HVE are primarily based on the proanthocyanidin content, as HT was found inactive on all the parameters tested. These results suggest further investigations of HVE in other inflammatory-based skin diseases.

The use of Cannabis sativa is currently recognized to ease certain types of chronic pain, reduce chemotherapy-induced nausea, and improve anxiety. Nevertheless, few studies highlighted the therapeutic potential of C. sativa extracts and related phytocannabinoids for a variety of widespread skin disorders including acne, atopic dermatitis, psoriasis, pruritus, and pain. This review summarized the current evidence on the effects of phytocannabinoids at the cutaneous level through the collection of in vitro, in vivo, and clinical studies published on PubMed, Scopus, Embase, and Web of Science until October 2020. Phytocannabinoids have demonstrated potential anti-inflammatory, antioxidant, anti-aging, and anti-acne properties by various mechanisms involving either CB 1 / 2 -dependent and independent pathways. Not only classical immune cells, but also several skin-specific actors, such as keratinocytes, fibroblasts, melanocytes, and sebocytes, may represent a target for phytocannabinoids. Cannabidiol, the most investigated compound, revealed photoprotective, antioxidant, and anti-inflammatory mechanisms at the cutaneous level, while the possible impact on cell differentiation, especially in the case of psoriasis, would require further investigation. Animal models and pilot clinical studies supported the application of cannabidiol in inflammatory-based skin diseases. Also, one of the most promising applications of non-psychotropic phytocannabinoids is the treatment of seborrheic disorders, especially acne. In conclusion, the incomplete knowledge of the role of the endocannabinoid system in skin disorders emerged as an important limit for pharmacological investigations. Moreover, the limited studies conducted on C. sativa extracts suggested a higher potency than single phytocannabinoids, thus stimulating new research on phytocannabinoid interaction., Competing Interests: The authors declare that they have no conflicts of interest., (Thieme. All rights reserved.)

Epidermal Patterning Factor Like 9 (EPFL9), also known as STOMAGEN, is a cysteine-rich peptide that induces stomata formation in vascular plants, acting antagonistically to other epidermal patterning factors (EPF1, EPF2). In grapevine there are two EPFL9 genes, EPFL9-1 and EPFL9-2 sharing 82% identity at protein level in the mature functional C-terminal domain. In this study, CRISPR/Cas9 system was applied to functionally characterize VvEPFL9-1 in 'Sugraone', a highly transformable genotype. A set of plants, regenerated after gene transfer in embryogenic calli via Agrobacterium tumefaciens , were selected for evaluation. For many lines, the editing profile in the target site displayed a range of mutations mainly causing frameshift in the coding sequence or affecting the second cysteine residue. The analysis of stomata density revealed that in edited plants the number of stomata was significantly reduced compared to control, demonstrating for the first time the role of EPFL9 in a perennial fruit crop. Three edited lines were then assessed for growth, photosynthesis, stomatal conductance, and water use efficiency in experiments carried out at different environmental conditions. Intrinsic water-use efficiency was improved in edited lines compared to control, indicating possible advantages in reducing stomatal density under future environmental drier scenarios. Our results show the potential of manipulating stomatal density for optimizing grapevine adaptation under changing climate conditions., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Clemens, Faralli, Lagreze, Bontempo, Piazza, Varotto, Malnoy, Oechel, Rizzoli and Dalla Costa.)

Sumac (Rhus coriaria L.) is a spice and medicinal herb traditionally used in the Mediterranean region and the Middle East. Since we previously demonstrated Sumac biological activity in a model of tumor necrosis factor alpha (TNF-α)-induced skin inflammation, the present work is aimed at further demonstrating a potential role in inflammatory disorders, focusing on gastritis. For this purpose, different polar extracts (water-W, ethanol-water-EW, ethanol-E, ethanol macerated-Em, acetone-Ac, ethylacetate-EtA) were investigated in gastric epithelial cells (GES-1) challenged by TNF-α or H. pylori infection. The ethanolic extracts (E, EW, Em) showed the major phenolic contents, correlating with lower half maximal inhibitory concentrations (IC50s) on the release of interleukin-8 (IL-8, <15 μg/mL) and interleukin-6 (IL-6, <20 μg/mL) induced by TNF-α. Similarly, they inhibited IL-8 release (IC50s < 70 μg/mL) during Helicobacter pylori (H. pylori) infection and exhibited a direct antibacterial activity at comparable concentrations (minimum inhibitory concentration (MIC) = 100 μg/mL). The phenolic content and the bioactivity of EW were maintained after simulated gastric digestion and were associated with nuclear factor kappa B (NF-κB) impairment, considered the main putative anti-inflammatory mechanism. On the contrary, an anti-urease activity was excluded. To the best of our knowledge, this is the first demonstration of the potential role of Sumac as a nutraceutical useful in H. pylori-related gastritis.

Metabolic syndrome is a complex condition associated with a series of pathologies featuring glucose intolerance, diabetes, high blood pressure, dyslipidemia, microalbuminuria, overweight, and obesity. It is also related to nonalcoholic fatty liver disease (NAFLD), recognized as the most familiar cause of chronic liver disease worldwide. The overall prevalence of metabolic syndrome and, consequently, the one of NAFLD is constantly increasing worldwide. The initial management of these diseases involves lifestyle modifications, including changes in diet and physical exercise. In addition to conventional drugs like orlistat, botanicals are traditionally used to counteract these disorders, and some of them are currently under evaluation. The present work evaluated the in vivo beneficial effects of hydroalcoholic extracts of two Cameroonian spices, focusing on obesity-related hepatic lipid injury in high-fat-fed C57BL/6 mice. Hydroethanolic extracts were prepared and characterized by reverse phase-high-performance liquid chromatography (HPLC)-photodiode array detection and ultra performance liquid chromatography-triple time-of-flight electrospray ionization tandem mass spectroscopy (TOF-ESI-MS/MS) analysis. Plant extracts were orally administered for 30 days at different dose levels (100 and 200 mg kg -1 body weight (BW)) to obese C57BL/6 mice. Food intake (FI) and BW were recorded daily. Plasma biochemical parameters and lipid content were estimated at the beginning and at the end of the experiment. Liver tissues were subjected to histological examinations, lipid content, as well as oxidative stress markers, and FAME (fatty acid methyl esters) were estimated. Oral administration of extracts at 200 mg kg -1 BW significantly reduced FI and prevented BW gain. A decrease in the weight of the liver and a decrease in the hepatic and plasma lipid content were observed. Plasma enzyme (serum glutamic-oxaloacetic transaminase, SGOT; serum glutamic pyruvic transaminase, SGPT; alkaline phosphatase, ALP) activities were not indicative of any organ damage. Chemical analysis suggested that phenolic acids (4-caffeoylquinic acid, p-coumaric acid 4-O-glucoside, 5-caffeoylshikimic acid, caffeic acid hexose, and 4-O-methyl gallic acid) and flavonoids (morusin derivatives, naringenin-7-O-glucoside, and homoisoflavanone) identified in the extracts could potentially justify the biological properties observed. The main findings of this study showed that Xylopia parviflora (A. Rich.) Benth and Aframomum citratum (Pereira ex Oliv. et Hanb.) K. Shum decreased hepatic lipid accumulation in high-fat-diet (HFD)-induced obese C57BL/6 mice and confirmed, at least in part, our previous in vitro and ex vivo studies. The molecular mechanisms underlying these effects are still unclear and will be explored in the future., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

Human epidermis responds to ultraviolet (UV)B-induced damage by tolerating it, restoring it, or undergoing programmed cell death when the damage is massive. Recently, compounds rich in polyphenols, such as Vitis vinifera L. leaf extract (VVLe), have attracted a lot of interest for skin protection. We investigated the effect of VVLe pre-treatment (1 h) in a 2D model of HaCaT cells and in 3D organotypic cultures of normal human skin exposed to a single UVB dose to study the immediate specific events 1 h and the response orchestrated in the epidermal layer 24 h after irradiation, respectively. In both models, transmission electron microscopy analysis was carried out. The expression of the inducible keratin K17, the activation of both pSTAT3 and Nuclear Factor (NF)-κB signalling pathways, and the epidermal distribution of Toll-Like Receptor (TLR) 4 were assessed by immunofluorescence in the 2D and 3D model. In 3D organotypic cultures, thanks to the preservation of a multi-layered structure, the epidermal distribution of the differentiation biomarkers K10 and K14 as well as of K16 was analysed by immunofluorescence, while the release of interleukin (IL)-8 was evaluated by ELISA. In skin bioptic fragments, cytotoxicity and genotoxicity were investigated by LDH assay and Alkaline Comet assay, respectively, and then compared to cell proliferation. The epidermal distribution of the histone γ-H2AX, indicating the fragmented DNA, was analysed by immunofluorescence. In both experimental models, VVLe tuned UVB-induced K17 expression to a different extent in HaCaT cells and in the skin. In HaCaT cells, pSTAT3 activation was induced by UVB and reverted by VVLe pre-treatment. TLR4 expression was triggered by UVB in both models, but VVLe pre-treatment abolished this event only in HaCaT cells. NF-κB immunostaining increased both in the nucleus and in the cytoplasm only in HaCaT cells after UVB irradiation. In all irradiated skin samples, VVLe pre-treatment was not able to revert the inhibition of epidermal proliferation, K16 expression, and IL-8 secretion. The effectiveness of VVLe in contrasting the irradiation-induced genotoxicity still remains unclear. In conclusion, our study clearly shows that K17 is a robust marker induced in keratinocytes upon UVB stimulation and that this event can be reverted by a pre-treatment with VVLe. On the whole, these observations represent a novelty in the scenario of the complex relationships between the effects exerted by UVB rays on human skin and significantly improve the knowledge regarding the modulation of the early epidermal response induced by a single exposure to UVB in the presence of VVLe., (© 2021 S. Karger AG, Basel.)

The molecular pathophysiology of cardiometabolic diseases is known to be influenced by dysfunctional ectopic adipose tissue. In addition to lifestyle improvements, these conditions may be managed by novel nutraceutical products. This study evaluatedthe effects of 11 Cameroonian medicinal spice extracts on triglyceride accumulation, glucose uptake, reactive oxygen species (ROS) production and interleukin secretion in SW 872 human adipocytes after differentiation with 100 µM oleic acid. Triglyceride content was significantly reduced by all spice extracts. Glucose uptake was significantly increased by Tetrapleura tetraptera , Aframomum melegueta and Zanthoxylum leprieurii . Moreover, Xylopia parviflora , Echinops giganteus and Dichrostachys glomerata significantly reduced the production of ROS. Concerning pro-inflammatory cytokine secretion, we observed that Tetrapleura tetraptera , Echinops giganteus , Dichrostachys glomerata and Aframomum melegueta reduced IL-6 secretion. In addition, Xylopia parviflora , Monodora myristica , Zanthoxylum leprieurii , and Xylopia aethiopica reduced IL-8 secretion, while Dichrostachys glomerata and Aframomum citratum increased it. These findings highlight some interesting properties of these Cameroonian spice extracts in the modulation of cellular parameters relevant to cardiometabolic diseases, which may be further exploited, aiming to develop novel treatment options for these conditions based on nutraceutical products.

Nowadays, medicinal herbs and their phytochemicals have emerged as a great therapeutic option for many disorders. However, poor bioavailability and selectivity might limit their clinical application. Therefore, bioavailability is considered a notable challenge to improve bio-efficacy in transporting dietary phytochemicals. Different methods have been proposed for generating effective carrier systems to enhance the bioavailability of phytochemicals. Among them, nano-vesicles have been introduced as promising candidates for the delivery of insoluble phytochemicals. Due to the easy preparation of the bilayer vesicles and their adaptability, they have been widely used and approved by the scientific literature. The first part of the review is focused on introducing phytosome technology as well as its applications, with emphasis on principles of formulations and characterization. The second part provides a wide overview of biological activities of commercial and non-commercial phytosomes, divided by systems and related pathologies. These results confirm the greater effectiveness of phytosomes, both in terms of biological activity or reduced dosage, highlighting curcumin and silymarin as the most formulated compounds. Finally, we describe the promising clinical and experimental findings regarding the applications of phytosomes. The conclusion of this study encourages the researchers to transfer their knowledge from laboratories to market, for a further development of these products., Competing Interests: The authors report no conflicts of interest for this work., (© 2021 Barani et al.)

Background: A number of cellular processes have evolved in metazoans that increase the proteome repertoire in relation to the genome, such as alternative splicing and translation recoding. Another such process, translational stop codon readthrough (SCR), generates C-terminally extended protein isoforms in many eukaryotes, including yeast, plants, insects, and humans. While comparative genome analyses have predicted the existence of programmed SCR in many species including humans, experimental proof of its functional consequences are scarce., Results: We show that SCR of the Drosophila POU/Oct transcription factor Ventral veins lacking/Drifter (Vvl/Dfr) mRNA is prevalent in certain tissues in vivo, reaching a rate of 50% in the larval prothoracic gland. Phylogenetically, the C-terminal extension is conserved and harbors intrinsically disordered regions and amino acid stretches implied in transcriptional activation. Elimination of Vvl/Dfr translational readthrough by CRISPR/Cas9 mutagenesis changed the expression of a large number of downstream genes involved in processes such as chromatin regulation, neurogenesis, development, and immune response. As a proof-of-principle, we demonstrate that the C-terminal extension of Vvl/Dfr is necessary for correct timing of pupariation, by increasing the capacity to regulate its target genes. The extended Vvl/Dfr isoform acts in synergy with the transcription factor Molting defective (Mld) to increase the expression and biosynthesis of the steroid hormone ecdysone, thereby advancing pupariation. Consequently, late-stage larval development was prolonged and metamorphosis delayed in vvl/dfr readthrough mutants., Conclusions: We demonstrate that translational recoding of a POU/Oct transcription factor takes place in a highly tissue-specific and temporally controlled manner. This dynamic and regulated recoding is necessary for normal expression of a large number of genes involved in many cellular and developmental processes. Loss of Vvl/Dfr translational readthrough negatively affects steroid hormone biosynthesis and delays larval development and progression into metamorphosis. Thus, this study demonstrates how SCR of a transcription factor can act as a developmental switch in a spatiotemporal manner, feeding into the timing of developmental transitions between different life-cycle stages., (© 2021. The Author(s).)

Fire blight disease, caused by the bacterium Erwinia amylovora (E. amylovora), is responsible for substantial losses in cultivated apples worldwide. An important mechanism of plant immunity is based on the recognition of conserved microbial molecules, named pathogen-associated or microbe-associated molecular patterns (PAMPs or MAMPs), through pattern recognition receptors (PRRs), leading to pattern-triggered immunity (PTI). The interspecies transfer of PRRs represents a promising strategy to engineer broad-spectrum and durable disease resistance in crops. EFR, the Arabidopsis thaliana PRR for the PAMP elf18 derived from the elongation factor thermal unstable (EF-Tu) proved to be effective in improving bacterial resistance when expressed into Solanaceae and other plant species. In this study, we tested whether EFR can affect the interaction of apple with E. amylovora by its ectopic expression in the susceptible apple rootstock M.26. Stable EFR expression led to the activation of PAMP-triggered immune response in apple leaves upon treatment with supernatant of E. amylovora, as measured by the production of reactive oxygen species and the induction of known defense genes. The amount of tissue necrosis associated with E. amylovora infection was significantly reduced in the EFR transgenic rootstock compared to the wild-type. Our results show that the expression of EFR in apple rootstock may be a valuable biotechnology strategy to improve the resistance of apple to fire blight., (© 2021. The Author(s).)

Ribes nigrum L. (blackcurrant) leaf extracts, due to high levels of flavonols and anthocyanins, have been shown to exhibit beneficial effects in inflammatory diseases. However, whereas their traditional use has been investigated and validated in several models of inflammation and oxidative stress, the possible impact on skin disorders is still largely unknown. The purpose of this work was to elucidate the effects of R. nigrum leaf extract (RNLE) on keratinocyte-derived inflammatory mediators, elicited by a Th1 or Th2 cytokine milieu. HaCaT cells were challenged with TNF-α, either alone or in combination with the costimulatory cytokines IFN-γ or IL-4, and the release of proinflammatory cytokines and mediators (IL-8, IL-6, s-ICAM-1, and TSLP) was evaluated. The results showed that RNLE preferentially interferes with IFN-γ signaling, demonstrating only negligible activity on TNF-α or IL-4. This effect was attributed to flavonols, which might also account for the ability of RNLE to impair TNF-α/IL-4-induced TSLP release in a cAMP-independent manner. These results suggest that RNLE could have an antiallergic effect mediated in keratinocytes via mechanisms beyond histamine involvement. In conclusion, the discovery of RNLE preferential activity against IFN-γ-mediated inflammation suggests potential selectivity against Th1 type response and the possible use in Th1 inflammatory diseases.

Cardiac toxicity still represents a common adverse outcome causing drug attrition and post-marketing withdrawal. The development of relevant in vitro models resembling the human heart recently opened the path towards a more accurate detection of drug-induced human cardiac toxicity early in the drug development process. Organs-on-chip have been proposed as promising tools to recapitulate in vitro the key aspects of the in vivo cardiac physiology and to provide a means to directly analyze functional readouts. In this scenario, a new device capable of continuous monitoring of electrophysiological signals from functional in vitro human hearts-on-chip is here presented. The development of cardiac microtissues was achieved through a recently published method to control the mechanical environment, while the introduction of a technology consisting in micro-electrode coaxial guides allowed to conduct direct and non-destructive electrophysiology studies. The generated human cardiac microtissues exhibited synchronous spontaneous beating, as demonstrated by multi-point and continuous acquisition of cardiac field potential, and expression of relevant genes encoding for cardiac ion-channels. A proof-of-concept pharmacological validation on three drugs proved the proposed model to potentially be a powerful tool to evaluate functional cardiac toxicity., (Creative Commons Attribution license.)

Objective: To investigate the main chemical components and the anti-inflammatory activity of extracts of Adelia ricinella L. aerial parts., Methods: Three extracts obtained by soxhlet extraction and ethanol/water mixtures were evaluated in their chemical composition by UPLC-DAD-MS/MS. The in vitro anti-inflammatory activity of the prepared extracts was assessed through three different assays: COX-1 and COX-2 enzymatic inhibition, cell-based COX assays on RAW264.7 macrophages (ATCC) measuring the COX-2 protein expression by Western blot and the measurement of the PGE2 concentration in the supernatants of the culture medium. Also was determinate the effect of the three extracts on the RAW 264.7 cell viability., Key Findings: Few differences in the phytochemical profile were found between the three prepared extracts, identifying a blend of thirteen flavonoids derived from luteolin and apigenin, with orientin as main constituent. Plant extracts (alcoholic and aqueous) did not affect the macrophage cell viability (IC50 > 256 μg/ml) and significantly reduced COX-1 and COX-2 enzyme activities. Additionally, COX-2 expression and PGE2 release were suppressed after 24 h of LPS stimulation and treatment with plant extracts (8-64 µg/ml)., Conclusions: A. ricinella extracts showed the ability to reduce the inflammatory effect exerted by LPS in murine macrophages. However, further studies should confirm their anti-inflammatory activity., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Royal Pharmaceutical Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Coccoloba cowellii Britton (Polygonaceae) is an endemic and critically endangered plant that only grows in Camagüey, a province of Cuba. In this study, a total of 13 compounds were identified in a methanolic leaf extract, employing a dereplication of the UHPLC-HRMS data by means of feature-based molecular networking (FBMN) analysis in the Global Natural Products Social Molecular Network (GNPS), together with the interpretation of the MS/MS data and comparison with the literature. The major constituents were glucuronides and glycosides of myricetin and quercetin, as well as epichatechin-3- O -gallate, catechin, epicatechin and gallic acid, all of them being reported for the first time in C. cowellii leaves. The leaf extract was also tested against various microorganisms, and it showed a strong antifungal effect against Candida albicans ATCC B59630 (azole-resistant) (IC 50 2.1 µg/mL) and Cryptococcus neoformans ATCC B66663 (IC 50 4.1 µg/mL) with no cytotoxicity (CC 50 > 64.0 µg/mL) on MRC-5 SV2 cells, determined by the resazurin assay. Additionally, the extract strongly inhibited COX-1 and COX-2 enzyme activity using a cell-free experiment in a dose-dependent manner, being significantly more active on COX-1 (IC 50 4.9 µg/mL) than on COX-2 (IC 50 10.4 µg/mL). The constituents identified as well as the pharmacological activities measured highlight the potential of C. cowellii leaves, increasing the interest in the implementation of conservation strategies for this species.

In Cameroon, local plants are traditionally used as remedies for a variety of ailments. In this regard, several papers report health benefits of Cameroonian spices, which include antioxidant and anti-microbial properties, whereas gastric anti-inflammatory activities have never been previously considered. The present study investigates the antioxidant and anti-inflammatory activities of hydro-alcoholic extracts of eleven Cameroonian spices in gastric epithelial cells (AGS and GES-1 cells). The extracts showed antioxidant properties in a cell-free system and reduced H 2 O 2 -induced ROS generation in gastric epithelial cells. After preliminary screening on TNFα-induced NF-κB driven transcription, six extracts from Xylopia parviflora , Xylopia aethiopica , Tetrapleura tetraptera , Dichrostachys glomerata , Aframomum melegueta , and Aframomum citratum were selected for further studies focusing on the anti-inflammatory activity. The extracts reduced the expression of some NF-κB-dependent pro-inflammatory mediators strictly involved in the gastric inflammatory process, such as IL-8, IL-6, and enzymes such as PTGS2 (COX-2), without affecting PTGS1 (COX-1). In conclusion, the selected extracts decreased pro-inflammatory markers by inhibiting the NF-κB signaling in gastric cells, justifying, in part, the traditional use of these spices. Other molecular mechanisms cannot be excluded, and further studies are needed to better clarify their biological activities at the gastric level.

Genome editing via CRISPR/Cas9 is a powerful technology, which has been widely applied to improve traits in cereals, vegetables and even fruit trees. For the delivery of CRISPR/Cas9 components into dicotyledonous plants, Agrobacterium tumefaciens mediated gene transfer is still the prevalent method, although editing is often accompanied by the integration of the bacterial T-DNA into the host genome. We assessed two approaches in order to achieve T-DNA excision from the plant genome, minimizing the extent of foreign DNA left behind. The first is based on the Flp/FRT system and the second on Cas9 and synthetic cleavage target sites (CTS) close to T-DNA borders, which are recognized by the sgRNA. Several grapevine and apple lines, transformed with a panel of CRISPR/SpCas9 binary vectors, were regenerated and characterized for T-DNA copy number and for the rate of targeted editing. As detected by an optimized NGS-based sequencing method, trimming at T-DNA borders occurred in 100% of the lines, impairing in most cases the excision. Another observation was the leakage activity of Cas9 which produced pierced and therefore non-functional CTS. Deletions of genomic DNA and presence of filler DNA were also noticed at the junctions between T-DNA and genomic DNA. This study proved that many factors must be considered for designing efficient binary vectors capable of minimizing the presence of exogenous DNA in CRISPRed fruit trees.

Malus x domestica microRNA MdmiR285N is a potential key regulator of plant immunity, as it has been predicted to target 35 RNA transcripts coding for different disease resistance proteins involved in plant defense to pathogens. In this study, the promoter region of MdmiR285N was isolated from the apple genome and analyzed in silico to detect potential regulatory regions controlling its transcription. A complex network of putative regulatory elements involved in plant growth and development, and in response to different hormones and stress conditions, was identified. Activity of the β-Glucoronidase ( GUS ) reporter gene driven by the promoter of MdmiR285N was examined in transgenic apple, demonstrating that MdmiR285N was expressed during the vegetative growth phase. Similarly, in transgenic Arabidopsis thaliana , spatial and temporal patterns of GUS expression revealed that MdmiR285N was differentially regulated during seed germination, vegetative phase change, and reproductive development. To elucidate the role of MdmiR285N in plant immunity, MdmiR285N expression in wild-type apple plants and GUS activity in transgenic apple and Arabidopsis thaliana plants were monitored in response to Erwinia amylovora and Pseudomonas syringae pv. Tomato DC3000. A significant decrease of MdmiR285N levels and GUS expression was observed during host-pathogen infections. Overall, these data suggest that MdmiR285N is involved in the biotic stress response, plant growth, and reproductive development., Competing Interests: Conflict of interestThe authors declare that they have no conflict of interest., (© The Author(s) 2020.)

During the last 30 years, berries have gained great attention as functional food against several risk factors in chronic diseases. The number of related publications on Pubmed rose from 1000 items in 1990 to more than 11,000 in 2019. Despite the fact that a common and clear definition of "berries" is not shared among different scientific areas, the phytochemical pattern of these fruits is mainly characterized by anthocyanins, flavanols, flavonols, and tannins, which showed antioxidant and anti-inflammatory properties in humans. Skin insults, like wounds, UV rays, and excessive inflammatory responses, may lead to chronic dermatological disorders, conditions often characterized by long-term treatments. The application of berries for skin protection is sustained by long traditional use, but many observations still require a clear pharmacological validation. This review summarizes the scientific evidence, published on EMBASE, MEDLINE, and Scholar, to identify extraction methods, way of administration, dose, and mechanism of action of berries for potential dermatological treatments. Promising in vitro and in vivo evidence of Punica granatum L. and Vitis vinifera L. supports wound healing and photoprotection, while Schisandra chinensis (Turcz.) Baill. and Vaccinium spp. showed clear immunomodulatory effects. Oral or topical administrations of these berries justify the evaluation of new translational studies to validate their efficacy in humans.

Oxidative stress plays a relevant role in the progression of chronic conditions, including cardiometabolic diseases. Several Cameroonian plants, including spices, are traditionally used as herbal medicines for the treatment of diseases where oxidative stress contributes to insulin resistance, like type 2 diabetes mellitus. This study evaluated the antioxidant capacity and the effects on oxidative-stress-induced impairment of glucose uptake of 11 Cameroonian spice extracts. H 2 O 2 -induced reactive oxygen species (ROS) production by human HepG2 cells was significantly reduced by 8/11 extracts. The most effective extracts, Xylopia parviflora , Echinops giganteus , and Dichrostachys glomerata , showed a concentration-dependent ROS-scavenging activity, which involved Nrf2 translocation into the nucleus. Xylopia parviflora , Tetrapleura tetraptera , Dichrostachys glomerata , Aframomum melegueta , and Aframomum citratum extracts showed the highest antioxidant capacity, according to oxygen radical absorbance capacity (ORAC) (2.52-88 μM Trolox Eq/g of extract), ferric-reducing antioxidant power (FRAP) (40.23-233.84 mg gallic acid Eq/g of extract), and total phenol (8.96-32.96% mg gallic acid Eq/g of extract) assays. In HepG2 cells, glucose uptake was stimulated by 4/11 extracts, similarly to insulin and metformin. H 2 O 2 -induced oxidative stress reduced glucose uptake, which was rescued by pretreatment with Xylopia aethiopica , Xylopia parviflora , Scorodophloeus zenkeri , Monodora myristica , and Dichrostachys glomerata extracts. The ROS-scavenging ability of the spice extracts may reside in some secondary metabolites observed by phytochemical profiling (reverse-phase high-performance liquid chromatography coupled to a diode array detector (HPLC-UV-DAD)). Further studies are needed to better clarify their biological activities and potential use to control oxidative stress and promote insulin sensitivity.

Rhus coriaria L. (sumac) is a small plant widely diffused in the Mediterranean region. Its fruit are often consumed as a spice but are also present in traditional medicine of several countries. Recently, interest in this plant has increased and many scientific works reported its beneficial effects including antioxidant and anti-inflammatory properties. Plant extracts can be successfully used against ultraviolet rays, which are able to reach and damage the human skin; however, sumac extracts were never applied to this usage. Thus, in this study, we used a macerated ethanol extract of Rhus coriaria L. dried fruit (mERC) to demonstrate its preventive role against the damage induced by ultraviolet-A rays (UV-A) on microvascular endothelial cells (HMEC-1). In vitro effects of the extract pre-treatment and UV-A exposure were evaluated in detail. The antioxidant capacity was assessed by reactive oxygen species (ROS) formation and cellular antioxidant activity measurement. Genoprotective effects of mERC were investigated as well. Our findings indicate that the extract acts as a cell cycle inhibitor or apoptosis inducer, according to the level of damage. The present work provides new insights into the usage of Rhus coriaria extracts against skin injuries.

Vitis vinifera L. water extract from red grapevine leaves contains high levels of polyphenols in quantities similar to those found in red grape and grape seeds. Phenolic compounds are the largest group of natural antioxidants with also an anti-inflammatory activity, widely demonstrated both in vitro and in vivo. Interestingly, their antioxidant effect relies not only on the direct radical scavenging activity but also on their ability in modulating cellular signalling transduction pathways. UV radiation exerts multiple effects on skin cells inducing apoptosis, senescence and carcinogenesis. The aim of this study was to investigate the antioxidant and the DNA protective potentials of Vitis vinifera L. water extract against UV-A and UV-B radiation in HaCaT cells, a human keratinocytes cell line. Comet and ɣH2AX assays were used to assess DNA damage in UV irradiated cells pre-treated or not with the extract (100 μg/mL). For UV-B, DNA damage resulted significantly increased at 40 mJ/cm 2 dose determining cell cycle arrest and apoptosis. For UV-A, DNA damage was significant at 10 J/cm 2 while cell cycle arrest and apoptosis were evident only at 25 J/cm 2 . The extract (1h of pre-treatment) highlights the antioxidant and scavenger activity on the UV-A, while the maintenance of the apoptosis with both UV-A and UV-B must be interpreted as an anti-mutagenic effect., Competing Interests: Declaration of Competing Interest None, (Copyright © 2020 Elsevier B.V. All rights reserved.)

The bacterium Erwinia amylovora, the causal agent of fire blight disease in apple, triggers its infection through the DspA/E effector which interacts with the apple susceptibility protein MdDIPM4. In this work, MdDIPM4 knockout has been produced in two Malus × domestica susceptible cultivars using the CRISPR/Cas9 system delivered via Agrobacterium tumefaciens. Fifty-seven transgenic lines were screened to identify CRISPR/Cas9-induced mutations. An editing efficiency of 75% was obtained. Seven edited lines with a loss-of-function mutation were inoculated with the pathogen. Highly significant reduction in susceptibility was observed compared to control plants. Sequencing of five potential off-target sites revealed no mutation event. Moreover, our construct contained a heat-shock inducible FLP/FRT recombination system designed specifically to remove the T-DNA harbouring the expression cassettes for CRISPR/Cas9, the marker gene and the FLP itself. Six plant lines with reduced susceptibility to the pathogen were heat-treated and screened by real-time PCR to quantify the exogenous DNA elimination. The T-DNA removal was further validated by sequencing in one plant line. To our knowledge, this work demonstrates for the first time the development and application of a CRISPR/Cas9-FLP/FRT gene editing system for the production of edited apple plants carrying a minimal trace of exogenous DNA., (© 2019 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

Skin inflammatory diseases result from complex events that include dysregulation and abnormal expression of inflammatory mediators or their receptors in skin cells. The present study investigates the potential effect of a Cannabis sativa L. ethanolic extract standardized in cannabidiol as antiinflammatory agent in the skin, unraveling the molecular mechanisms in human keratinocytes and fibroblasts. The extract inhibited the release of mediators of inflammation involved in wound healing and inflammatory processes occurring in the skin. The mode of action involved the impairment of the nuclear factor-kappa B (NF-κB) pathway since the extract counteracted the tumor necrosis factor-alpha-induced NF-κB-driven transcription in both skin cell lines. Cannabis extract and cannabidiol showed different effects on the release of interleukin-8 and vascular endothelial growth factor, which are both mediators whose genes are dependent on NF-κB. The effect of cannabidiol on the NF-κB pathway and metalloproteinase-9 (MMP-9) release paralleled the effect of the extract thus making cannabidiol the major contributor to the effect observed. Down-regulation of genes involved in wound healing and skin inflammation was at least in part due to the presence of cannabidiol. Our findings provide new insights into the potential effect of Cannabis extracts against inflammation-based skin diseases., (© 2019 John Wiley & Sons, Ltd.)

Ethnopharmacological Relevance: Rhus coriaria L. (R. coriaria) is a medicinal herb native to the middle east and Mediterranean region and well-known as "sumac" or "sicilian sumac". This herb has a wide range of traditional applications, covering its topical use to treat skin burns or eczemas and to promote wound healing., Aim of the Study: The present research aims to investigate the potential anti-inflammatory activity of Rhus coriaria L. fruit extracts in human keratinocytes (HaCaT cells), evaluating extracts prepared using different techniques., Materials and Methods: Water (WRC), ethanol-water (EWRC) and two types of ethanol extracts (mERC and ERC) were prepared. The HaCaT cells were challenged by TNF-α (10 ng/mL) and IL-8, ICAM-1, VEGF, and MMP-9 release, as well as NF-κB translocation, were measured by ELISA assays. The most active extracts were chemically profiled through HPLC-UV-DAD analysis., Results: Althought all the extracts inhibited the TNF-α-induced IL-8 release, just mERC and EWRC suppressed NF-κB activation, ICAM-1, and MMP-9 secretion. EWRC showed higher inhibition on ICAM-1 and MMP-9 with IC 50 s of 1.76 ± 0.24 and 1.24 ± 0.33 μg/mL, respectively (mean ± s.d.). On the contrary, mERC significantly decreased VEGF levels whereas EWRC did not show any effect. The HPLC-UV profile of the extracts revealed higher amount of anthocyanins in EWRC in comparison with mERC., Conclusions: Our results suggest the potential positive effect of R. coriaria fruit extracts, mostly mERC, as preventive agent in the treatment of keratinocyte inflammation through their inhibitory effect on the production of skin pro-inflammatory mediators., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Atherosclerosis is characterized by interaction between immune and vascular endothelial cells which is mediated by adhesion molecules occurring on the surface of the vascular endothelium leading to massive release of proinflammatory mediators. Ginkgo biloba L. (Ginkgoaceae) standardized extracts showing beneficial effects are commonly prepared by solvent extraction, and acetone is used according to the European Pharmacopoeia recommendations; the well-known Ginkgo biloba acetone extract EGb761® is the most clinically investigated. However, in some countries, the allowed amount of solvent is limited to ethanol, thus implying that the usage of a standardized Ginkgo biloba ethanol extract may be preferred in all those cases, such as for food supplements. The present paper investigates if ethanol and acetone extracts, with comparable standardization, may be considered comparable in terms of biological activity, focusing on the radical scavenging and anti-inflammatory activities. Both the extracts showed high inhibition of TNF α -induced VCAM-1 release (41.1-43.9  μ g/mL), which was partly due to the NF- κ B pathway impairment. Besides ROS decrease, cAMP increase following treatment with ginkgo extracts was addressed and proposed as further molecular mechanism responsible for the inhibition of endothelial E-selectin. No statistical difference was observed between the extracts. The present study demonstrates for the first time that ethanol and acetone extracts show comparable biological activities in human endothelial cell, thus providing new insights into the usage of ethanol extracts in those countries where restrictions in amount of acetone are present.

Psoriasis is a chronic cutaneous condition characterized by the release of pro-inflammatory mediators and oxidative stress. The reduction of these factors is currently the most effective strategy to inhibit the symptoms of pathology. Antioxidants from natural sources are increasingly used to improve skin conditions. Dried red leaves from grapevine ( Vitis vinifera L., cv Teinturiers) showed anti-inflammatory and anti-bacterial activities, but their possible effects on keratinocytes have not been previously investigated. In this study we tested the ability of a water extract from grapevine leaves (VVWE) to inhibit inflammatory conditions in human keratinocytes (HaCaT cells), challenged with proinflammatory (tumor necrosis factor-α (TNF-α) or lipopolysaccharide (LPS)) or prooxidant (ultraviolet B radiation (UVB) or H 2 O 2 ) mediators. VVWE inhibited interleukin-8 (IL-8) secretion induced by proinflammatory stimuli, acting on the IL-8 promoter activity, but the effect was lower when prooxidant mediators were used. The effect was partly explained by the reduction of nuclear factor-κB (NF-κB)-driven transcription and nuclear translocation. Furthermore, vascular endothelial growth factor (VEGF) secretion, a regulator of angiogenesis, was inhibited by VVWE, but not matrix metalloproteinase-9 (MMP-9), a protease involved in matrix remodeling. VVWE, assayed on Franz diffusion cell system, showed a marked reduction of High Performance Liquid Chromatography (HPLC)-identified compounds. Pure molecules individually failed to reduce TNF-α-induced IL-8 release, suggesting synergistic effects or the presence of other bioactive compounds still unknown.