Moscatilin inhibits vascular calcification by activating IL13RA2-dependent inhibition of STAT3 and attenuating the WNT3/ß-catenin signalling pathway.

INTRODUCTION: Vascular calcification, a devastating vascular complication accompanying atherosclerotic cardiovascular disease and chronic kidney disease, increases the incidence of adverse cardiovascular events and compromises the efficacy of vascular interventions. However, effective therapeutic drugs and treatments to delay or prevent vascular calcification are lacking. OBJECTIVES: This study was designed to test the therapeutic effects and mechanism of Moscatilin (also known as dendrophenol) from Dendrobium huoshanense (an eminent traditional Chinese medicine) in suppressing vascular calcification in vitro, ex vivo and in vivo. METHODS: Male C57BL/6J mice (25-week-old) were subjected to nicotine and vitamin D3 (VD3) treatment to induce vascular calcification. In vitro, we established the cellular model of osteogenesis of human aortic smooth muscle cells (HASMCs) under phosphate conditions. RESULTS: By utilizing an in-house drug screening strategy, we identified Moscatilin as a new naturally-occurring chemical entity to reduce HASMC calcium accumulation. The protective effects of Moscatilin against vascular calcification were verified in cultured HASMCs. Unbiased transcriptional profiling analysis and cellular thermal shift assay suggested that Moscatilin suppresses vascular calcification via binding to interleukin 13 receptor subunit A2 (IL13RA2) and augmenting its expression. Furthermore, IL13RA2 was reduced during HASMC osteogenesis, thus promoting the secretion of inflammatory factors via STAT3. We further validated the participation of Moscatilin-inhibited vascular calcification by the classical WNT/ß-catenin pathway, among which WNT3 played a key role in this process. Moscatilin mitigated the crosstalk between WNT3/ß-catenin and IL13RA2/STAT3 to reduce osteogenic differentiation of HASMCs. CONCLUSION: This study supports the potential of Moscatilin as a new naturally-occurring candidate drug for treating vascular calcification via regulating the IL13RA2/STAT3 and WNT3/ß-catenin signalling pathways.

Is there a role for traditional and complementary medicines in managing chronic fatigue? a systematic review of randomized controlled trials.

Introduction: Chronic fatigue syndrome (CFS) is an increasingly common condition that is challenging to treat due to unclear etiology and a lack of consensus on clinical diagnosis and treatment guidance. Many affected people resorted to using traditional and complementary medicines (T&CMs). However, the evidence for T&CMs for CFS has been inconclusive and continues to evolve. The study aims to identify, summarize and assess the most recent evidence on the efficacy and safety of T&CMs for CFS. Methods: Randomized controlled trials (RCTs) investigating T&CMs for CFS published in English of Chinese between 1 January 2013 and 31 December 2022 were searched from 7 databases. RCTs comparing T&CMs with no treatment, placebo, or pharmacological medicine were included, irrespective of language or blinding. The Consolidated Standards of Reporting Trials Statement extensions for Chinese herbal medicine Formulas (CONSORT-CHM) and the Cochrane Collaboration's Risk of Bias tool were used to evaluate the quality and risk of bias of included studies. Results: A total of 62 RCTs investigating 43 types of T&CMs and involving 5,231 participants with CFS were included in this review. The primary outcome measures mainly included the scoring of fatigue symptoms using the validated tool Fatigue Scale-14 (FS-14) or the TCM syndrome score. The main interventions showing overall efficacy were Chaihu Guizhi Decoction and Buzhong Yiqi combined with Xiao Chaihu Decoction, and 148 ingredients were identified, including Astragali Radix, Glycyrrhizae Radix et Rhizoma, Atractylodis Macrocephalae Rhizoma, and Bupleuri Radix. The most significant effect was the improvement of fatigue, followed by TCM-diagnosed symptoms and other psychological conditions. No serious adverse effect had been reported. However, the quality of the RCTs included RCTs were found to be suboptimal, and the risk of bias remained uncertain. Conclusion: Some evidence from RCTs supported the efficacy and safety of T&CM in CFS. However, given the methodological and quality heterogenicity of the included studies, the recommendations of T&CMs in treating CFS remain inconclusive. To develop better quality evidence about T&CMs for CFS, future studies should employ more objective diagnosis standards and outcome measurements, larger sample size, and better bias control, and ensure the compliance with the corresponding reporting guidelines. Systematic Review Registration: https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42022362268, identifier CRD42022362268.

Efficacy and safety of traditional Chinese medicine for cancer-related fatigue: a systematic literature review of randomized controlled trials.

BACKGROUND: Cancer-related fatigue (CRF) is an extremely common and long-term condition that affects the physical and mental health of oncology patients. While the treatment for CRF with western medicine and non-pharmacological therapy remains uncertain and challenging, traditional Chinese medicine (TCM) has become a trending option for the patients. Based on the findings from randomized controlled trials (RCTs), this study aims to identify and evaluate the evidence about the efficacy and safety of TCM for CRF. METHODS: A systematic literature search was conducted according to the PRISMA literature research guidelines. Seven electronic databases including PubMed, the Cochrane Library, Embase, Web of Science, Scopus, China National Knowledge Infrastructure (CNKI) and Wanfang database were searched to identify RCTs which investigated TCM in the treatment of CRF published since inception to December 2022. RCTs comparing TCM with no treatment, placebo, or pharmacological interventions were considered eligible for this review. The Consolidated Standards of Reporting Trials Statement extensions for Chinese herbal medicine Formulas (CONSORT-CHM) and the Cochrane Collaboration's Risk of Bias tool were used in this review to evaluate the quality and the risk of bias of all included trials. RESULTS: A total of 82 RCTs were included in this review, regardless of whether they were published in English or Chinese. After data extraction and results evaluation, 78 trials demonstrated overall efficacy in using TCM for CRF patients compared with the control group, in which 33 trials showed that the efficacy rate was statistically significant (p < 0.05 or p < 0.01). TCM was also shown to be beneficial in improving the scores of relevant scales (e.g., PFS, QoL, TCM syndrome score, other fatigue scales etc.) or physical tests indicators (e.g., cytokines, blood test etc.). The most common herbs found in Chinese medicine were Astragali Radix, Ginseng Radix and Codonopsis Radix. Some TCM products, such as Kangai Injection, Buzhong Yiqi Decoction and Shenqi Fuzheng Injection could provide a reference for medication in this review. A range of non-serious, reversible adverse effects associated with the use of TCM was also reported. However, the result of evaluation showed that none of the trials fully met all the CONSORT-CHM criteria, the quality of included trials was generally poor and the risk of bias was mostly uncertain. CONCLUSION: There is some evidence supporting the efficacy and safety of TCM in managing CRF in this systematic review. However, no clear conclusion can be made due to the inadequate reporting of efficacy and adverse reactions. In view of some concerns about the existing evidence after the evaluation, it is essential to standardize the comprehensive identification and efficacy measurement standards, improve the quality of RCTs and conduct more multicomponent therapies to provide an updated reference for CRF patients medication in the future. The protocol of this systematic review has been registered on PROSPERO (CRD42023413625). [ https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42023413625 ].

A systemic review on Liubao tea: A time-honored dark tea with distinctive raw materials, process techniques, chemical profiles, and biological activities.

Liubao tea (LBT) is a unique microbial-fermented tea that boasts a long consumption history spanning 1500 years. Through a specific post-fermentation process, LBT crafted from local tea cultivars in Liubao town Guangxi acquires four distinct traits, namely, vibrant redness, thickness, aging aroma, and purity. The intricate transformations that occur during post-fermentation involve oxidation, degradation, methylation, glycosylation, and so forth, laying the substance foundation for the distinctive sensory traits. Additionally, LBT contains multitudinous bioactive compounds, such as ellagic acid, catechins, polysaccharides, and theabrownins, which contributes to the diverse modulation abilities on oxidative stress, metabolic syndromes, organic damage, and microbiota flora. However, research on LBT is currently scattered, and there is an urgent need for a systematical recapitulation of the manufacturing process, the dominant microorganisms during fermentation, the dynamic chemical alterations, the sensory traits, and the underlying health benefits. In this review, current research progresses on the peculiar tea varieties, the traditional and modern process technologies, the substance basis of sensory traits, and the latent bioactivities of LBT were comprehensively summarized. Furthermore, the present challenges and deficiencies that hinder the development of LBT, and the possible orientations and future perspectives were thoroughly discussed. By far, the productivity and quality of LBT remain restricted due to the reliance on labor and experience, as well as the incomplete understanding of the intricate interactions and underlying mechanisms involved in processing, organoleptic quality, and bioactivities. Consequently, further research is urgently warranted to address these gaps.

Epigallocatechin gallate (EGCG) alleviates the inflammatory response and recovers oral microbiota in acetic acid-induced oral inflammation mice.

The oral microbiota, the second largest microbiome in the human body, plays an integral role in maintaining both the local oral and systemic health of the host. Oral microecological imbalances have been identified as a potential risk factor for numerous oral and systemic diseases. As a representative component of tea, epigallocatechin gallate (EGCG) has demonstrated inhibitory effects on most pathogens in single-microbial models. In this study, the regulatory effect of EGCG on more complex oral microbial systems was further explored through a mouse model of acetic acid-induced oral inflammation. Acetic acid induces histological damage in the cheek pouch, tongue, and throat, such as broken mucosa, submucosal edema, and muscular disorders. These detrimental effects were ameliorated significantly following EGCG treatment. Additionally, EGCG reduced the levels of the inflammatory cytokines interleukin-6 and tumor necrosis factor-α to alleviate the inflammation of the tongue, cheek pouch, and throat. According to the 16S rDNA gene sequencing data, EGCG treatment contributed to increased diversity of the oral microbiota and the reversal of oral microecological disorder. This study demonstrates the regulatory effect of EGCG on dysregulated oral microbiota, providing a potential option for the prevention and treatment of oral-microbiota-associated diseases.

Green tea extracts alleviate acetic acid-induced oral inflammation and reconstruct oral microbial balance in mice.

Oral cavity contains the second largest microbial community in the human body. Due to the highly vascularized feature of mouth, oral microbes could directly access the bloodstream and affect the host healthy systemically. The imbalance of oral microbiota is closely related to various oral and systemic diseases. Green tea extracts (GTE) mainly contain tea polyphenols, alkaloids, amino acid, flavones, and so on, which equipped with excellent anti-inflammatory activities. Previous studies have demonstrated the beneficial effects of GTE on oral health. However, most researches used in vitro models or focused on limited microorganisms. In this study, the regulatory effect of GTE on oral microbiome and the alleviative effect on oral inflammation in vivo were evaluated. The results showed that GTE could efficiently alleviate the inflammations of the tongue, cheek pouch, as well as throat. GTE effectively inhibited the activation of NF-κB through the upregulation of the anti-inflammatory cytokine interleukin (IL)-10, consequently leading to reduced expression of pro-inflammatory cytokines IL-6 and tumor necrosis factor-α. The indexes of spleen and thymus were also elevated by GTE in stomatitis mice. Moreover, GTE promoted the growth of probiotics Lactobacillus and Bacillus, inhibited the reproduction of pathogens Achromobacter, reversing the microbiota disorders in oral cavity. This study not only presents a novel approach for enhancing oral microecology but also facilitates the wider adoption of tea consumption.

Mollugin activates GLP-1R to improve cognitive dysfunction in type 2 diabetic mice.

AIMS: The incidence of diabetic cognitive dysfunction is increasing year by year, and it has gradually become a research hot spot. Studies have shown that glucagon-like peptide-1 receptor (GLP-1R) agonists can improve cognitive dysfunction in diabetic patients. This study focuses on whether small molecule GLP-1R agonists from traditional Chinese medicine (TCM) can improve the diabetic cognitive dysfunction. MATERIALS AND METHODS: The small molecules from TCM were screened by cell membrane chromatography (CMC) with GLP-1R-HEK293 cell membrane column. MTT assay, flow cytometry, immunofluorescence cytochemistry and other methods were used to determine the effects of mollugin on the apoptosis rate and reactive oxygen species (ROS) level of high glucose (HG)/hydrogen peroxide (H2O2) induced PC12 cells. Real-Time PCR was used to detect mRNA expression in mouse cerebral cortex. Water maze test was further used to confirm the effect of mollugin on cognitive dysfunction in T2DM mice. KEY FINDINGS: Mollugin bound to GLP-1R, promoted Ca2+ influx, increased insulin secretion and cAMP content in ß-TC-6 cells. Mollugin enhanced the cell viability, ameliorated apoptosis, reduced intracellular ROS levels in HG/H2O2-injured PC12 cells. Mollugin reduced the T2DM mice's escape latency, improved neuronal cell damage, decreased the expression of Pik3ca, Akt1 and Mapk1 mRNA in the cerebral cortex tissue. SIGNIFICANCE: The results suggest that mollugin could improve cognitive dysfunction in T2DM mice through activating GLP-1R/cAMP/PKA signal pathway.

Schisandrin B, a potential GLP-1R agonist, exerts anti-diabetic effects by stimulating insulin secretion.

Diabetes mellitus is a metabolic disease that is characterized by elevated blood sugar. Although glucagon-like peptide-1 receptor agonists (GLP-1RA) lower blood glucose in a glucose-dependent manner, most of them are macromolecule polypeptides. Macromolecular peptides are relatively expensive and inconvenient compared with small molecules. Therefore, this study sought to identify the small molecules binding to GLP-1R via cell membrane chromatography (CMC), confirm their agonistic activity, and further study its beneficial effects in a mouse model of type 2 diabetes mellitus (T2DM) induced by a combination of high-fat diet and streptozotocin. We used CMC, calcium imaging and molecular docking techniques to screen and identify the potential small molecule Schisandrin B (Sch B), which exhibits a strong binding effect to GLP-1R, from the small molecule library of traditional Chinese medicine. Through in-vitro experiments, we found that Sch B stimulated insulin secretion in ß-TC-6 cells, while GLP-1R antagonist Exendin9-39, adenylate cyclase inhibitor SQ22536, and protein kinase A (PKA) inhibitor H89 could significantly inhibit the insulin secretion induced by Sch B. In vivo, Sch B significantly improved fasting blood glucose levels, intraperitoneal glucose tolerance test damage, and the status of pancreatic tissue damage, and reduced serum insulin levels, total cholesterol, triglyceride and low density lipoprotein in T2DM mice. These results indicate that Sch B alleviates T2DM by promoting insulin release through the GLP-1R/cAMP/PKA signaling pathway, suggesting that Sch B may be a potential GLP-1RA, which is expected to provide a new therapeutic strategy for the prevention and treatment of T2DM.

Current insight into enhanced strategies and interaction mechanisms of hydrogel materials for phosphate removal and recovery from wastewater.

Phosphorus plays a crucial role in modern society but often pollutes the environment through raising eutrophication and has particularly devastating effects on the water environment. As a promising material platform, the three-dimensional network structure and the tailorable nature of hydrogels provide infinite application possibilities. Thereinto, phosphate removal and recovery from wastewater using hydrogel materials have gained momentum since their rapid reactivity, ease of operation, low cost and simplicity of recovery compared to traditional techniques. In this review, current strategies for functional enhancement of hydrogel materials are systematically summarized from different perspectives. Following, based on the discussion of different interaction mechanisms between phosphates and hydrogels, the phosphate mass transfer and performance of hydrogels and their current application are critically reviewed. This review aims to present mechanistic insight into the recent development in phosphate removal and recovery using hydrogel materials and provides new ideas for constructing high-efficient hydrogels and laying the foundations for the practical application of this technology.

Photothermal-responsive Prussian blue nanocages loaded with thrombin for tumor starvation therapy and photothermal therapy.

The goal of inhibiting tumor growth can be achieved by cutting off the supply of nutrients in the blood vessels of a tumor site, but finding ways to effectively, accurately and safely deliver drugs that can induce vascular embolism remains a challenge. Phase change materials (PCM) can undergo solid-liquid transformation at the phase change temperature. This study reports on a near-infrared ray (NIR)-responsive nano-drug delivery platform based on Prussian blue (PB) nanoparticles. The PCM (lauric acid) can encapsulate thrombin (Thr) in the Prussian blue nanocage (PB Cage), and effectively avoid the pre-leakage of Thr during blood circulation. When the (Thr/PCM)@PB Cage is accumulated at the tumor site and irradiated with NIR, the thermal effect induced by the PB Cage causes the PCM to undergo a solid-liquid state transition, rapidly releasing the encapsulated Thr and inducing coagulation in the tumor blood vessels. Based on the safe delivery and precisely controlled release of Thr, the proliferation of tumor cells can be inhibited without damaging other tissues and organs. In addition, PB Cage-induced photothermal therapy can also ablate tumor cells. Thr-induced "starvation therapy" based on PB Cage loading provides a good reference for precise controlled-release drug delivery systems.

[Molecular mechanism of ligustilide attenuating OGD/R injury in PC12 cells by inhibiting ferroptosis].

The aim of this study is to explore the mechanism of ligustilide, the main active constituent of essential oils of traditional Chinese medicine Angelicae Sinensis Radix, on alleviating oxygen-glucose deprivation/reperfusion(OGD/R) injury in PC12 cells from the perspective of ferroptosis. OGD/R was induced in vitro, and 12 h after ligustilide addition during reperfusion, cell viability was detected by cell counting kit-8(CCK-8) assay. DCFH-DA staining was used to detect the level of intracellular reactive oxygen species(ROS). Western blot was employed to detect the expression of ferroptosis-related proteins, glutathione peroxidase 4(GPX4), transferrin receptor 1(TFR1), and solute carrier family 7 member 11(SLC7A11), and ferritinophagy-related proteins, nuclear receptor coactivator 4(NCOA4), ferritin heavy chain 1(FTH1), and microtubule-associated protein 1 light chain 3(LC3). The fluorescence intensity of LC3 protein was analyzed by immunofluorescence staining. The content of glutathione(GSH), malondialdehyde(MDA), and Fe was detected by chemiluminescent immunoassay. The effect of ligustilide on ferroptosis was observed by overexpression of NCOA4 gene. The results showed that ligustilide increased the viability of PC12 cells damaged by OGD/R, inhibited the release of ROS, reduced the content of Fe and MDA and the expression of TFR1, NCOA4, and LC3, and improved the content of GSH and the expression of GPX4, SLC7A11, and FTH1 compared with OGD/R group. After overexpression of the key protein NCOA4 in ferritinophagy, the inhibitory effect of ligustilide on ferroptosis was partially reversed, indicating that ligustilide may alleviate OGD/R injury of PC12 cells by blocking ferritinophagy and then inhibiting ferroptosis. The mechanism by which ligustilide reduced OGD/R injury in PC12 cells is that it suppressed the ferroptosis involved in ferritinophagy.

Identification of the active compounds in the Yi-Fei-San-Jie formula using a comprehensive strategy based on cell extraction/UPLC-MS/MS, network pharmacology, and molecular biology techniques.

BACKGROUND: Chinese herbal formulae has multiple active constituents and targets, and the good clinical response is encouraging more scientists to explore the bio-active ingredients in such complex systems. Yi-Fei-San-Jie formula (YFSJF) is commonly used to treat patients with lung cancer in South China; however, its bio-active ingredients remain unknown. PURPOSE: We investigated the bio-active ingredients of the YFSJF using a novel comprehensive strategy. METHODS: A549 cell extraction coupled with ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS/MS) was used for the screening of potential bio-active ingredients. Network pharmacology approach and molecular dynamics simulation were performed for the screening of targets. Surface plasmon resonance (SPR) assay and molecular biology techniques were used to verify the targets. RESULTS: Nine A549 cell membrane-binding compounds were identified through cell extraction/UPLC-MS/MS. Five compounds, namely ginsenoside Ro, ginsenoside Rb1, ginsenoside Rc, peimisine, and peimine were cytotoxic to A549 cells, and they were considered the bio-active ingredients of the YFSJF in vitro. Network pharmacology analysis revealed that TGFBR2 is the key target and the TGFß pathway is the key pathway targeted by YFSJF in non-small cell lung cancer. Peimisine showed an affinity to TGFBR2 using molecular docking and dynamic stimulation, which was confirmed using surface plasmon resonance spectroscopy. The molecular biology-based analysis further confirmed that peimisine targets TGFBR2 and can reverse A549 epithelial-mesenchymal transition by inhibiting the TGFß pathway. CONCLUSION: Taken together, cell extraction/UPLC-MS/MS, network pharmacology, and molecular biology-based analysis comprise a feasible strategy to explore active ingredients in YFSJF.

Systematic analysis of randomised controlled trials of Chinese herb medicine for non-alcoholic steatohepatitis (NASH): implications for future drug development and trial design.

BACKGROUND: Non-alcoholic steatohepatitis (NASH) is a liver disease currently lacking an approved therapy, resulting in significant clinical demand. Traditional Chinese medicines (TCMs) have been commonly used to manage NASH. This study aimed to systematically analyse the randomised controlled trials (RCTs) using TCMs for NASH management. METHODS: A systematic literature review was performed by following PRISMA guidelines 2020 in six electronic databases: PubMed, Web of Science, Scopus, Embase, the Cochrane Library, and China National Knowledge Infrastructure, from inception until August 2022. RCTs using TCMs for NASH were included in the analysis, irrespective of language or blinding. RESULTS: 112 RCTs were included in this review, with 10,573 NASH participants. 108 RCTs were conducted in China, and 4 RCTs were in other countries. Herbal medicine decoction was the major dosage form used for treating NASH (82/112). 11 TCMs products have been approved for NASH treatment (8 in China, 2 in Iran, and 1 in Japan). Classic prescriptions, such as "Huang Lian Jie Du decoction", "Yin Chen Hao decoction", and "Yi Guan Jian" were used in some studies. The TCMs treatment of NASH involved the use of 199 different plants, with the top 5 herbs being Salviae Miltiorrhizae Radix Et Rhizoma, Alismatis Rhizoma, Bupleuri Radix, Poria, and Curcumae Radix. "Salviae Miltiorrhizae Radix Et Rhizoma + Bupleuri Radix/Alismatis Rhizoma" were the mostly common drug-pair in the herbs network analysis. Nowadays, "Bupleuri Radix/Alismatis Rhizoma + Atractylodis Macrocephalae Rhizoma" are increasingly applied in herbal formulas for NASH. Based on the PICOS principles, the included studies varied in terms of the population, intervention, comparator, outcomes, and study design. However, some studies reported unstandardised results and failed to report diagnostic standards, inclusion or exclusion criteria, or sufficient patient information. CONCLUSION: Adopting Chinese classic prescriptions or drug-pair may provide a basis for developing new drugs of NASH management. Further research is needed to refine the clinical trial design and obtain more convincing evidence for using TCMs to treat NASH.

Dextran-Sulfate-Sodium-Induced Colitis-Ameliorating Effect of Aqueous Phyllanthus emblica L. Extract through Regulating Colonic Cell Gene Expression and Gut Microbiomes.

The anti-inflammation effect of aqueous Phyllanthus emblica L. extract (APE) and its possible underlying mechanism in dextran sulfate sodium (DSS)-induced mice chronic colonic inflammation were studied. APE treatment significantly improved the colitic symptoms, including ameliorating the shortening of the colon, increasing the DSS-induced body weight loss, reducing the disease activity index, and reversing the condition of colon tissue damage of mucus lost and goblet cell reduction. Overproduction of serum pro-inflammatory cytokines were suppressed by the treatment of APE. Gut microbiome analysis showed that APE remodeled the structure of gut bacteria in phylum and genus levels, upregulating the abundance of phylum Bacteroidetes, family Muribaculaceae, and genus Bacteroides and downregulating the abundance of phylum Firmicutes. The reshaped gut microbiome caused metabolic functions and pathway change with enhanced queuosine biosynthesis and reduced polyamine synthesis pathway. Colon tissue transcriptome analysis further elucidated APE-inhibited mitogen-activated protein kinase (MAPK), cytokine-cytokine receptor interaction, and tumor necrosis factor (TNF) signaling pathways and the expressions of the genes that promote the progress of colorectal cancer. It turned out that APE reshaped the gut microbiome and inhibited MAPK, cytokine-cytokine receptor interaction, and TNF signaling pathways as well as the colorectal-cancer-related genes to exert its colitis protective effect.

Magnesium promotes tea plant growth via enhanced glutamine synthetase-mediated nitrogen assimilation.

Acidic tea (Camellia sinensis) plantation soil usually suffers from magnesium (Mg) deficiency, and as such, application of fertilizer containing Mg can substantially increase tea quality by enhancing the accumulation of nitrogen (N)-containing chemicals such as amino acids in young tea shoots. However, the molecular mechanisms underlying the promoting effects of Mg on N assimilation in tea plants remain unclear. Here, both hydroponic and field experiments were conducted to analyze N, Mg, metabolite contents, and gene expression patterns in tea plants. We found that N and amino acids accumulated in tea plant roots under Mg deficiency, while metabolism of N was enhanced by Mg supplementation, especially under a low N fertilizer regime. 15N tracing experiments demonstrated that assimilation of N was induced in tea roots following Mg application. Furthermore, weighted gene correlation network analysis (WGCNA) analysis of RNA-seq data suggested that genes encoding glutamine synthetase isozymes (CsGSs), key enzymes regulating N assimilation, were markedly regulated by Mg treatment. Overexpression of CsGS1.1 in Arabidopsis (Arabidopsis thaliana) resulted in a more tolerant phenotype under Mg deficiency and increased N assimilation. These results validate our suggestion that Mg transcriptionally regulates CsGS1.1 during the enhanced assimilation of N in tea plant. Moreover, results of a field experiment demonstrated that high Mg and low N had positive effects on tea quality. This study deepens our understanding of the molecular mechanisms underlying the interactive effects of Mg and N in tea plants while also providing both genetic and agronomic tools for future improvement of tea production.

[Mechanism of total flavonoids of Rhododendra simsii in alleviating ischemic brain injury].

This study explores the effect of total flavonoids of Rhododendra simsii(TFR) on middle cerebral artery occlusion(MCAO)-induced cerebral injury in rats and oxygen-glucose deprivation/reoxygenation(OGD/R) injury in PC12 cells and the underlying mechanism. The MCAO method was used to induce focal ischemic cerebral injury in rats. Male SD rats were randomized into sham group, model group, and TFR group. After MCAO, TFR(60 mg·kg~(-1)) was administered for 3 days. The content of tumor necrosis factor-α(TNF-α), interleukin-1(IL-1), and interleukin-6(IL-6) in serum was detected by enzyme-linked immunosorbent assay(ELISA). The pathological changes of brain tissue and cerebral infarction were observed based on hematoxylin and eosin(HE) staining and 2,3,5-triphenyltetrazolium chloride(TTC) staining. RT-qPCR and Western blot were used to detect the mRNA and protein levels of calcium release-activated calcium channel modulator 1(ORAI1), stromal interaction molecule 1(STIM1), stromal intera-ction molecule 2(STIM2), protein kinase B(PKB), and cysteinyl aspartate specific proteinase 3(caspase-3) in brain tissues. The OGD/R method was employed to induce injury in PC12 cells. Cells were randomized into the normal group, model group, gene silencing group, TFR(30 µg·mL~(-1)) group, and TFR(30 µg·mL~(-1))+gene overexpression plasmid group. Intracellular Ca~(2+) concentration and apoptosis rate of PC12 cells were measured by laser scanning confocal microscopy and flow cytometry. The effect of STIM-ORAI-regulated store-operated calcium entry(SOCE) pathway on TFR was explored based on gene silencing and gene overexpression techniques. The results showed that TFR significantly alleviated the histopathological damage of brains in MCAO rats after 3 days of admini-stration, reduced the contents of TNF-α, IL-1, and IL-6 in the serum, down-regulated the expression of ORAI1, STIM1, STIM2, and caspase-3 genes, and up-regulated the expression of PKB gene in brain tissues of MCAO rats. TFR significantly decreased OGD/R induced Ca~(2+) overload and apoptosis in PC12 cells. However, it induced TFR-like effect by ORAI1, STIM1 and STIM2 genes silencing. However, overexpression of these genes significantly blocked the effect of TFR in reducing Ca~(2+) overload and apoptosis in PC12 cells. In summary, in the early stage of focal cerebral ischemia-reperfusion injury and OGD/R-induced injury in PC12 cells TFR attenuates ischemic brain injury by inhibiting the STIM-ORAI-regulated SOCE pathway and reducing Ca~(2+) overload and inflammatory factor expression, and apoptosis.

Development on potential skin anti-aging agents of Cosmos caudatus Kunth via inhibition of collagenase, MMP-1 and MMP-3 activities.

BACKGROUND: Skin aging is associated with degradation of collagen by matrix metalloproteinases (MMPs), which leads to loss of skin elasticity and formation of wrinkles. Cosmos caudatus Kunth (CC) has been traditionally claimed as an anti-aging agent in Malaysia. Despite its well-known antioxidant activity, the anti-aging properties of CC was not validated. PURPOSE: This study aimed to investigate the anti-aging potential of CC extracts and fractions, particularly their inhibition of collagenase, MMP-1 and MMP-3 activities in human dermal fibroblasts CCD-966SK, followed by isolation, identification and analysis of their bioactive constituents. STUDY DESIGN AND METHODS: DPPH assay was firstly used to evaluate the antioxidant activity throughout the bioactivity-guided fractionation. Cell viability was determined using MTS assay. Collagenase activity was examined, while MMP-1 and MMP-3 expression were measured using qRT-PCR and western blotting. Then, chemical identification of pure compounds isolated from CC fractions was done by using ESIMS, 1H and 13C NMR spectroscopies. HPLC analyses were carried out for bioactive fractions to quantify the major components. RESULTS: Throughout the antioxidant activity-guided fractionation, fractions CC-E2 and CC-E3 with antioxidant activity and no toxicity towards CCD-966SK cells were obtained from CC 75% ethanol partitioned layer (CC-E). Both fractions inhibited collagenase activity, MMP-1 and MMP-3 mRNA and protein expression, as well as NF-κB activation induced by TNF-α in CCD-966SK cells. 14 compounds, which mainly consists of flavonoids and their glycosides, were isolated. Quercitrin (14.79% w/w) and quercetin (11.20% w/w) were major compounds in CC-E2 and CC-E3, respectively, as quantified by HPLC. Interestingly, both fractions also inhibited the MMP-3 protein expression synergistically, compared with treatment alone. CONCLUSION: The quantified CC fractions rich in flavonoid glycosides exhibited skin anti-aging effects via the inhibition of collagenase, MMP-1 and MMP-3 activities, probably through NF-κB pathway. This is the first study reported on MMP-1 and MMP-3 inhibitory activity of CC with its chemical profile, which revealed its potential to be developed as anti-aging products in the future.

Huaier granule prolongs overall survival after curative resection of hepatocarcinoma carcinoma: A propensity score analysis.

BACKGROUND: The effectiveness and safety of huaier granules in reducing recurrence after curative resection of HCC have been confirmed, but it is unclear whether huaier granules can significantly prolong overall survival. AIM: To demonstrate the effectiveness of huaier granule for HCC after curative resection over a 5-year follow-up. METHOD: A total of 1265 HCC patients who underwent curative resection from January 2008 to January 2020 were enrolled, 1111 patients were finally enrolled according to the exclusion criteria, and the oncology outcome of Huaier granule was analyzed by propensity score matching method (PSM). RESULT: Before propensity score matching, huaier granule resulted in better 5- year overall survival (61.49% vs 54.92%, p = 0.0099) and recurrence-free survival (45.64% vs 38.42%, p = 0.0042) for HCC patients. For solitary HCC ≤30 mm, huaier granule resulted in similar 5- year recurrence-free survival (54.55% vs 50.13%, p = 0.4403), but better 5- year overall survival (82.42% vs70.08%, p = 0.0189). Similar to overall patients, huaier granule resulted in better 5- year overall survival (54.77% vs 51.37%, p = 0.1530) and recurrence-free survival (42.61% vs 35.62%, p = 0.0082) for solitary HCC >30 mm. After propensity score matching, we did confirm that huaier granules can significantly prolong overall survival by more than 5 years, the exception was recurrence-free survival in the HCC <30 mm cohort.

Comparison of ONT and CCS sequencing technologies on the polyploid genome of a medicinal plant showed that high error rate of ONT reads are not suitable for self-correction.

BACKGROUND: Many medicinal plants are known for their complex genomes with high ploidy, heterozygosity, and repetitive content which pose severe challenges for genome sequencing of those species. Long reads from Oxford nanopore sequencing technology (ONT) or Pacific Biosciences Single Molecule, Real-Time (SMRT) sequencing offer great advantages in de novo genome assembly, especially for complex genomes with high heterozygosity and repetitive content. Currently, multiple allotetraploid species have sequenced their genomes by long-read sequencing. However, we found that a considerable proportion of these genomes (7.9% on average, maximum 23.7%) could not be covered by NGS (Next Generation Sequencing) reads (uncovered region by NGS reads, UCR) suggesting the questionable and low-quality of those area or genomic areas that can't be sequenced by NGS due to sequencing bias. The underlying causes of those UCR in the genome assembly and solutions to this problem have never been studied. METHODS: In the study, we sequenced the tetraploid genome of Veratrum dahuricum (Turcz.) O. Loes (VDL), a Chinese medicinal plant, with ONT platform and assembled the genome with three strategies in parallel. We compared the qualities, coverage, and heterozygosity of the three ONT assemblies with another released assembly of the same individual using reads from PacBio circular consensus sequencing (CCS) technology, to explore the cause of the UCR. RESULTS: By mapping the NGS reads against the three ONT assemblies and the CCS assembly, we found that the coverage of those ONT assemblies by NGS reads ranged from 49.15 to 76.31%, much smaller than that of the CCS assembly (99.53%). And alignment between ONT assemblies and CCS assembly showed that most UCR can be aligned with CCS assembly. So, we conclude that the UCRs in ONT assembly are low-quality sequences with a high error rate that can't be aligned with short reads, rather than genomic regions that can't be sequenced by NGS. Further comparison among the intermediate versions of ONT assemblies showed that the most probable origin of those errors is a combination of artificial errors introduced by "self-correction" and initial sequencing error in long reads. We also found that polishing the ONT assembly with CCS reads can correct those errors efficiently. CONCLUSIONS: Through analyzing genome features and reads alignment, we have found the causes for the high proportion of UCR in ONT assembly of VDL are sequencing errors and additional errors introduced by self-correction. The high error rates of ONT-raw reads make them not suitable for self-correction prior to allotetraploid genome assembly, as the self-correction will introduce artificial errors to > 5% of the UCR sequences. We suggest high-precision CCS reads be used to polish the assembly to correct those errors effectively for polyploid genomes.

Effect of Rubus extract on TLR4/NF-kappa B signaling pathway in alcoholic liver fibrosis rats.

This study aims to explore the effect of the Rubus extract on the TLR4/NF-κB signaling pathway in alcoholic liver fibrosis rats. The alcoholic liver rat model was established by continuous ethanol gavage administration. Rats were divided randomly into six groups (i.e., blank control, model, 0.05g/kg Rubus extract, 0.125g/kg Rubus extract, 0.259 g/kg Rubus extract and positive control groups). Liver tissue and blood were collected after treatment for four weeks. The pathological changes in the liver were observed by HE and Masson staining methods. The hyaluronic acid (HA), TNF-α and IL-6 levels were determined by ELISA kits. The TLR4 and p-p65 protein expression levels in liver were detected by Western blot. The liver lesion degree was significantly decreased in the Rubus extract group, and a high concentration of the Rubus extract indicated a significant improvement. The TNF-α, HA and IL-6 levels in the Rubus extract and positive control groups were significantly lower than those of the model group (P<0.05). The TLR4 and p-p65 protein expression levels were also significantly decreased in the Rubus extract and positive control groups (P< 0.05) with a concentration dependence of Rubus extract. The Rubus extract could delay the development of alcoholic liver fibrosis through inhibiting the TLR4/NF-κB pathway activity.