Aqueous extract of fermented Eucommia ulmoides leaves alleviates hyperlipidemia by maintaining gut homeostasis and modulating metabolism in high-fat diet fed rats.

BACKGROUND: As a traditional Chinese medicinal herb, the lipid-lowing biological potential of Eucommia ulmoides leaves (EL) has been demonstrated. After fermentation, the EL have been made into various products with lipid-lowering effects and antioxidant activity. However, the anti-hyperlipidemic mechanism of fermented Eucommia ulmoides leaves (FEL) is unclear now. PURPOSE: To evaluate the effects of FEL on hyperlipidemia and investigate the mechanism based on regulating gut homeostasis and host metabolism. METHODS: Hyperlipidemia animal model in Wistar rats was established after 8 weeks high-fat diet (HFD) fed. The administered doses of aqueous extract of FEL (FELE) were 128, 256 and 512 mg/kg/d, respectively. Serum biochemical parameters detection, histopathological sections analysis, 16S rDNA sequencing of gut microbiota and untargeted fecal metabolomics analysis, were performed to determine the therapeutic effects and predict related pathways of FELE on hyperlipidemia. The changes of proteins and genes elated to lipid were detected by Immunofluorescence (IF) and quantitative real-time polymerase chain reaction (qRT-PCR). RESULTS: 56 Components in FELE were identified by UPLC-MS, with organic acids, flavonoids and phenolic acids accounting for the majority. The intervention of FELE significantly reduced the body weight, lipid accumulation and the levels of total cholesterol (TC), triglycerides (TG), and low-density lipoprotein-cholesterol (LDL-C) in hyperlipidemia rats, while increased the level of High-density lipoprotein-cholesterol (HDL-C). Meanwhile, FELE improved the inflammatory makers and oxidative stress factors, which is tumor necrosis factor-α (TNF-α), monocyte chemotactic protein-1 (MCP-1), interleukin-6 (IL-6), malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT). These results demonstrated that FETE can effectively reduce blood lipids and alleviate inflammation and oxidative damage caused by hyperlipidemia. Mechanistically, FELE restore the homeostasis of gut microbiota by reducing the Firmicutes/Bacteroidetes ratio and increasing the abundance of probiotics, especially Lactobacillus, Rombousia, Bacteroides, Roseburia, Clostridia_UCG-014_Unclassified, while modulated metabolism through amino acid, bile acid and lipid-related metabolism pathways. In addition, the Pearson correlation analysis found that the upregulated bilirubin, threonine, dopamine and downregulated lipocholic acid, d-sphingosine were key metabolites after FELE intervention. IF and qRT-PCR analysis showed that FELE upregulated the expression of fatty acid oxidation proteins and genes (PPARα, CPT1A), bile acid synthesis and excretion proteins and genes (LXRα, CYP7A1, FXR), and downregulated the expression of adipogenic gene (SREBP-1c) by regulating gut microbiota to improve metabolism and exert a lipid-lowering effect. CONCLUSION: This work filled the lipid-lowering mechanism gap of FEL. FELE can improve HFD-induced hyperlipidemia by regulating the gut microbiota homeostasis and metabolism. Thus, FEL has the potential to develop into the novel raw material of lipid-lowering drugs.

CsCuAO1 Associated with CsAMADH1 Confers Drought Tolerance by Modulating GABA Levels in Tea Plants.

Our previous study showed that COPPER-CONTAINING AMINE OXIDASE (CuAO) and AMINOALDEHYDE DEHYDROGENASE (AMADH) could regulate the accumulation of γ-aminobutyric acid (GABA) in tea through the polyamine degradation pathway. However, their biological function in drought tolerance has not been determined. In this study, Camellia sinensis (Cs) CsCuAO1 associated with CsAMADH1 conferred drought tolerance, which modulated GABA levels in tea plants. The results showed that exogenous GABA spraying effectively alleviated the drought-induced physical damage. Arabidopsis lines overexpressing CsCuAO1 and CsAMADH1 exhibited enhanced resistance to drought, which promoted the synthesis of GABA and putrescine by stimulating reactive oxygen species' scavenging capacity and stomatal movement. However, the suppression of CsCuAO1 or CsAMADH1 in tea plants resulted in increased sensitivity to drought treatment. Moreover, co-overexpressing plants increased GABA accumulation both in an Agrobacterium-mediated Nicotiana benthamiana transient assay and transgenic Arabidopsis plants. In addition, a GABA transporter gene, CsGAT1, was identified, whose expression was strongly correlated with GABA accumulation levels in different tissues under drought stress. Taken together, CsCuAO1 and CsAMADH1 were involved in the response to drought stress through a dynamic GABA-putrescine balance. Our data will contribute to the characterization of GABA's biological functions in response to environmental stresses in plants.

The effect of intercropping leguminous green manure on theanine accumulation in the tea plant: A metagenomic analysis.

Intercropping is a widely recognised technique that contributes to agricultural sustainability. While intercropping leguminous green manure offers advantages for soil health and tea plants growth, the impact on the accumulation of theanine and soil nitrogen cycle are largely unknown. The levels of theanine, epigallocatechin gallate and soluble sugar in tea leaves increased by 52.87% and 40.98%, 22.80% and 6.17%, 22.22% and 29.04% in intercropping with soybean-Chinese milk vetch rotation and soybean alone, respectively. Additionally, intercropping significantly increased soil amino acidnitrogen content, enhanced extracellular enzyme activities, particularly ß-glucosidase and N-acetyl-glucosaminidase, as well as soil multifunctionality. Metagenomics analysis revealed that intercropping positively influenced the relative abundances of several potentially beneficial microorganisms, including Burkholderia, Mycolicibacterium and Paraburkholderia. Intercropping resulted in lower expression levels of nitrification genes, reducing soil mineral nitrogen loss and N2 O emissions. The expression of nrfA/H significantly increased in intercropping with soybean-Chinese milk vetch rotation. Structural equation model analysis demonstrated that the accumulation of theanine in tea leaves was directly influenced by the number of intercropping leguminous green manure species, soil ammonium nitrogen and amino acid nitrogen. In summary, the intercropping strategy, particularly intercropping with soybean-Chinese milk vetch rotation, could be a novel way for theanine accumulation.

Quality Marker Discovery and Quality Evaluation of Eucommia ulmoides Pollen Using UPLC-QTOF-MS Combined with a DPPH-HPLC Antioxidant Activity Screening Method.

Pollen, as an important component of Eucommia ulmoides (EUP), is rich in nutrients and is receiving increasing attention. At present, there are no reports on research related to the chemical composition and quality standards of EUP, and there are significant quality differences and counterfeit phenomena in the market. This study used a UPLC-QTOF-MS system to identify 49 chemical components in EUP for the first time. In the second step, 2,2-diphenyl-1-picrylhydrazyl (DPPH)-HPLC antioxidant activity screening technology was used to identify the main active components of EUP, quercetin-3-O-sophoroside (QSH), quercetin-3-O-sambubioside (QSB), and quercetin 3-O-neohesperidoside (QNH), and their purification, preparation, and structure identification were carried out. Third, molecular docking was used to predict the activity of these components. Fourth, the intracellular ROS generation model of RAW264.7 induced by H2O2 was used to verify and evaluate the activity of candidate active ingredients to determine their feasibility as Q-markers. Finally, a quality control method for EUP was constructed using the three selected components as Q-markers. The identification of chemical components and the discovery, prediction, and confirmation of characteristic Q-markers in EUP provide important references for better research on EUP and the effective evaluation and control of its quality. This approach provides a new model for the quality control of novel foods or dietary supplements.

Iron metabolism: An emerging therapeutic target underlying the anti-Alzheimer's disease effect of ginseng.

Finding neuroprotective drugs with fewer side effects and more efficacy has become a major problem as the global prevalence of Alzheimer's disease (AD) rises. Natural drugs have risen to prominence as potential medication candidates. Ginseng has a long history of use in China, and it has a wide range of pharmacological actions that can help with neurological issues. Iron loaded in the brain has been linked to AD pathogenesis. We reviewed the regulation of iron metabolism and its studies in AD and explored how ginseng might regulate iron metabolism and prevent or treat AD. Researchers utilized network pharmacology analysis to identify key factive components of ginseng that protect against AD by regulating ferroptosis. Ginseng and its active ingredients may benefit AD by regulating iron metabolism and targeting ferroptosis genes to inhibit the ferroptosis process. The results present new ideas for ginseng pharmacological studies and initiatives for further research into AD-related drugs. To provide comprehensive information on the neuroprotective use of ginseng to modulate iron metabolism, reveal its potential to treat AD, and provide insights for future research opportunities.

Pharmacokinetic evaluation of Sinisan containing vinegar-processed products in depressive rats, a comprehensive perspective of 'individual herb, herb-pair, and herbal formula'.

ETHNOPHARMACOLOGICAL RELEVANCE: As a classical formula for the treatment of depression, the clinical application of vinegar-processed products of Bupleuri Radix (Bupleurum chinense DC., BR) and Paeoniae Radix Alba (Paeonia lactiflora Pall., PRA) contained in Sinisan (SNS) is still controversial. AIM OF THE STUDY: Three levels of 'individual herb, herb-pair, and herbal formula' were employed to investigate whether and how the processing of main drugs affected the active constituents of pharmacokinetics in SNS, as well as their impacts on the hepatic CYP450 enzyme. MATERIALS AND METHODS: Rats were subjected to construct a chronic unpredictable mild stimulation (CUMS) model. A rapid and sensitive ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS/MS) analytical method was developed and validated for simultaneously quantitative evaluation of thirteen potential active compounds of SNS in depressive rat plasma, and successfully applied to a holistic comparison of pharmacokinetics. The differences in pharmacokinetic parameters based on three different forms of drug composition from BR and PRA before and after vinegar-processing were compared. Meanwhile, qRT-PCR and Western Blot were utilized to explore the metabolic activity of three isoforms of CYP450 enzyme scattered in the livers of depressive rats. RESULTS: The characteristic pharmacokinetics profiles of thirteen representative constituents in CUMS rats were influenced by vinegar-processing of BR and PRA and/or the compatibility. In detail, there were significant differences in the Cmax, AUC0-24, AUC0-∞, t1/2, and MRT0-24 of most constituents among the three different forms of drug composition from BR and PRA before and after vinegar-processing, with the most obvious changes in six constituents from the adjuvant and mediating guide drugs. And also, the pharmacokinetic parameters of seven constituents from BR and PRA in SNS containing vinegar-processed products obviously changed after compatibility. Additionally, the mRNA and protein levels of CYP1A2, CYP2E1, and CYP3A1 were observed to increase significantly with the processing of BR and PRA and the combination/formulation. CONCLUSIONS: In conclusion, SNS containing vinegar-processed products was more conducive to the absorption of most activated constituents compared to the original formula in vivo. The vinegar-processing of BR and PRA and the compatibility co-contribute to the pharmacokinetic variability of active compounds of SNS in CUMS rats, and the extent of contribution varies among drugs, which might be related to the regulation of the hepatic drug metabolizing enzymes. The finding of the investigation could help to better understand how active compounds metabolized in vivo, which might be helpful for guiding the clinical application of SNS containing vinegar-processed products.

Does acupuncture therapy affect peripheral inflammatory cytokines of major depressive disorder? A protocol for the systematic review and meta-analysis.

Background: Acupuncture is widely used as adjuvant therapy for major depressive disorder (MDD). There is robust evidence that inflammation is closely associated with MDD. To date, only a few numbers of studies have investigated the potential relationship between acupuncture and the change of inflammatory biomarkers in patients with MDD. Additionally, the results are inconsistent among studies. The current study aims to provide a comprehensive, systematic review of the association between acupuncture and changes in peripheral inflammation of patients with MDD, and clarify the alterations of inflammatory cytokines before and after acupuncture treatment by meta-analysis. Methods and analysis: This study will be conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. Eligible randomized controlled trials (RCTs) reporting acupuncture, with inflammatory cytokines as the outcome measured before and after intervention in patients with MDD, were searched in electronic databases, such as PubMed, Embase, Cochrane, SINOMED, Wanfang, China national knowledge infrastructure (CNKI), and Chongqing VIP (CQVIP). Primary outcomes of interest will be validated to measure the levels of inflammatory cytokines before and after acupuncture treatment in patients with MDD. Discussion: Acupuncture can drive anti-inflammatory effects, as well as symptom changes in MDD, which may represent a viable, multi-faceted treatment approach in MDD. Systematic review registration: [PROSPERO], identifier [CRD42021289207 on 04 December 2021].

A UPLC-Q-TOF-MS-Based Metabolomics Approach to Screen out Active Components in Prepared Rhubarb for Its Activity on Noxious Heat Blood Stasis Syndrome.

Background: Prepared rhubarb was obtained by steaming raw rhubarb with wine. Different from raw rhubarb with a purgative effect, prepared rhubarb shows effects of promoting blood circulation and removing blood stasis. However, the mechanisms of its action through regulating endogenous metabolites remain unclear. Purpose: The purpose of this study was to explore active chemical components in prepared rhubarb for its activity on noxious heat blood stasis syndrome (NHBS) by comprehensive metabolomics profiling. Study design: Plant extracts usually show their activities in a synergistic way; therefore, integrated omics was developed as a rational way for a better understanding of their biological effects and potential active compounds. Methods: The activities of prepared rhubarb were evaluated by biochemical and metabolomic analysis; meanwhile, serum chemical profiles were sought using UHPLC-Q-TOF-MS. Gray correlation analysis (GCA) was used for calculating the underlying correlations between them. Results: The metabolomics profiles of rat plasma from model and control groups were significantly different, with 31 endogenous metabolites changed by NHBS. Then, after the administration of prepared rhubarb, 18 of them were regulated. Multiple metabolic pathways were disturbed after NHBS modeling and restored by prepared rhubarb, among which had a greater impact on sphingolipid metabolism. A total of 28 compounds from prepared rhubarb absorbed into the plasma were identified, including nine prototypes and 19 metabolites. Statistical results suggested that rhein and its metabolites accounted for half of the top 10 active compounds in prepared rhubarb for its biomedical activities. Conclusion: This study presented evidence for the therapeutic effects and active chemicals of prepared rhubarb on NHBS in the way of metabolomics.

CsAAP7.2 is involved in the uptake of amino acids from soil and the long-distance transport of theanine in tea plants (Camellia sinensis L.).

Tea plant roots can uptake both inorganic nitrogen (NH4+ and NO3-) and organic nitrogen (amino acids) from the soil. These amino acids are subsequently assimilated into theanine and transported to young shoots through the xylem. Our previous study showed that CsLHT1 and CsLHT6 transporters take up amino acids from the soil, and CsAAPs participate in the transport of theanine. However, whether other amino acid transporters are involved in this process remains unknown. In this study, we identified two new CsAAPs homologous to CsAAP7, named CsAAP7.1 and CsAAP7.2. Heterologous expression of CsAAP7.1 and CsAAP7.2 in the yeast mutant 22Δ10α showed that CsAAP7.2 had the capacity to transport theanine and other amino acids, whereas CsAAP7.1 had no transport activity. Transient expression of the CsAAP7.2-GFP fusion protein in tobacco leaf epidermal cells confirmed its localization to the endoplasmic reticulum. Tissue-specific analysis showed that CsAAP7.2 was highly expressed in roots and stems. In addition, CsAAP7.2 overexpression lines were more sensitive to high concentrations of theanine due to the high accumulation of theanine in seedlings. Taken together, these findings suggested that CsAAP7.2 plays an important role in the uptake of amino acids from soil and the long-distance transport of theanine. These results provide valuable tools for nitrogen nutrition studies and enrich our understanding of theanine transport in tea plants.

Effects of heat-processed Gynostemma pentaphyllum on high-fat diet-fed mice of obesity and functional analysis on network pharmacology and molecular docking strategy.

ETHNOPHARMACOLOGICAL RELEVANCE: Gynostemma pentaphyllum has been used as traditional medicine for many diseases, including metabolic syndrome (Mets), aging, diabetes, neurodegenerative diseases in China, some East Asian and Southeast Asian countries. It was shown that G. pentaphyllum and gypenosides had anti-obesity and cholesterol-lowering effects too. However, its main active ingredients are still unclear. AIMS: The objective of this study was to compare the effects of gypenosides before and after heat-processing on high fat obese mice, and to analyze the function of G. pentaphyllum saponin via network pharmacology and molecular docking. METHODS: The leaves of G. pentaphyllum were heat processed at 120 °C for 3 h to obtain heat-processed G. pentaphyllum. Gypenosides (Gyp) and heat-processed gypenosides (HGyp) were prepared by resin HP-20 chromatography and analyzed using LC-MS from the extracts of G. pentaphyllum before and after heat-processing, respectively. Obesity model was made with high fat diet (HFD). Gyp and HGyp were administrated at 100 mg/kg for 12 weeks in HFD obese mice and the body weight, energy intake, and levels of total cholesterol (TC), triglyceride (TG), low-density lipoprotein (LDL), high-density lipoprotein (HDL) were compared. HGyp was administrated at a dose of 50,100,200 mg/kg for 12 weeks in HFD obese mice and the perirenal adipose, epididymal adipose, abdominal adipose, shoulder brown adipose, inguinal adipose were measured. Moreover, the potential targets, hub genes and pathways of damulin A, damulin B, gypenoside L, gypenoside LI for treating Mets were screened out via network pharmacology. According to the results of network pharmacology, core targets of treating Mets were docking with damulin A, gypenoside L, damulin B, gypenoside LI via molecular docking. RESULTS: HGyp showed stronger effects on body weight loss and lipid-lowering in obese mice than Gyp. The contents of gypenoside L, gypenoside LI, damulin A and damulin B of G. pentaphyllum were increased by heat-processing. HGyp significantly decreased the body weight, calorie intake, and levels of TC, TG, LDL, HDL on the obese mice. It up-regulated PPARα and PPARγ in the liver tissues. HGyp reduced significantly the size of adipocytes in inguinal, abdominal, epididymal adipose and increased the proportion of interscapular brown fat. Network pharmacology results showed that 21 potential targets and 12 related-pathways were screened out. HMGCR, ACE, LIPC, LIPG, PPARα PPARδ, PPARγ were the core targets of HGyp against lipid metabolism by molecular docking. The putative functional targets of HGyp may be modulated by AGE-RAGE, TNF, glycerolipid metabolism, lipid and atherosclerosis, cholesterol metabolism, PPAR, fat digestion and absorption, cell adhesion molecules signaling pathway. CONCLUSIONS: Gyp and HGyp are valuable for inhibition obesity, lipid-lowering, metabolic regulation. Especially, the effect of HGyp is better than that of Gyp.

Tea plantation intercropping green manure enhances soil functional microbial abundance and multifunctionality resistance to drying-rewetting cycles.

Climate change leads to more serious drying-rewetting alternation disturbance, which furtherly affects soil ecosystem function and agriculture production. Intercropping green manure, as an ancient agricultural practice, can improve the physical, chemical, and biological fertility of soil in tea plantation. However, the effects of intercropping green manure on soil multifunctional resistance to drying-rewetting disturbance in tea plantation has not been reported. In this study, the effects of different green manure practices over four years (tea plant monoculture, tea plant and soybean intercropping, tea plant and soybean + milk vetch intercropping) on soil multifunctionality resistance to drying-rewetting cycles, and the pivotal influencing factors were investigated. We used quantitative PCR array and analysis of multiple enzyme activities to characterize the abundance of functional genes and ecosystem multifunctionality, respectively. Compared with tea plantation monoculture, tea plant intercropping soybean and soybean + milk vetch significantly increased multifunctionality resistance by 12.07% and 25.86%, respectively. Random forest analysis indicated that rather than the diversity, the abundance of functional genes was the major drive of multifunctionality resistance. The structure equation model further proved that tea plantation intercropping green manure could improve the abundance of C cycling related functional genes mediated by soil properties, and ultimately increased multifunctionality resistance to drying-rewetting disturbance. Therefore, tea plantation intercropping green manure is an effective approach to maintain the multifunctionality resistance, which is conducive to maintain the soil nutrient supply capacity and tea production under the disturbance of drying-rewetting alternation.

Application and Development of Intelligent Medicine in Traditional Chinese Medicine.

As modern science and technology constantly progresses, the fields of artificial intelligence, mixed reality technology, remote technology, etc. have rapidly developed. Meanwhile, these technologies have been gradually applied to the medical field, leading to the development of intelligent medicine. What's more, intelligent medicine has greatly promoted the development of traditional Chinese medicine (TCM), causing huge changes in the diagnosis of TCM ailments, remote treatment, teaching, etc. Therefore, there are both opportunities and challenges for inheriting and developing TCM. Herein, the related research progress of intelligent medicine in the TCM in China and abroad over the years is analyzed, with the purpose of introducing the present application status of intelligent medicine in TCM and providing reference for the inheritance and development of TCM in a new era.

[Roles of processing and compatibility in antidepression of Sini Powder:a metabonomics study].

The antidepressant mechanism of Sini Powder was investigated by metabonomics based on UHPLC-Q-TOF-MS, and the roles of processing and compatibility in the antidepression of Sini Powder were discussed in the present study. The chronic unpredictable mild stress(CUMS) model of depression was induced in the model group, the Bupleuri Radix group, the Paeoniae Radix Alba group, the herb-pair group(Bupleuri Radix-Paeoniae Radix Alba), the Sini Powder group, and the vinegar-processed Sini Powder group(Bupleuri Radix and Paeoniae Radix Alba were vinegar-processed). After the establishment of the model, the rats in each group were continuously administered with corresponding drugs(ig) at a dose of 9.6 g·kg~(-1) for eight days [the rats in the model group and the normal group(without model induction) received the same volume of normal saline at the same time]. Following the last administration, the differential metabolites were identified to analyze metabolic pathways based on the rat plasma samples collected from each group. A total of sixteen potential biomarkers were identified. The metabolites with significant changes were involved in many biological metabolic pathways, such as amino acid metabolism, pentose phosphate pathway, glycerol phospholipid metabolism, sphingolipid metabolism, and purine metabolism. After drug intervention, some biomarkers returned to normal levels. Further comparisons of processing and compatibility revealed that the vinegar-processed Sini Powder group had the most total metabolic pathways where differential metabolites were returned to normal. Compared with the individual herbs, the herb-pair significantly improved the recovery of differential metabolites in the pentose phosphate and purine metabolic pathways. Compared with the Sini Powder, the vinegar-processed Sini Powder facilitated the recovery of differential metabolites in the arginine biosynthesis, and pyrimidine and pentose phosphate metabolic pathways. As indicated by the results, Sini Powder may interfere with depression by regulating lipid and nucleotide metabolisms. The processing and compatibility of Chinese herbal medicines can potentiate the intervention on depression by regulating nucleotide, energy, and amino acid metabolisms to a certain extent.

[Simultaneous quantitative analysis of nine saponins in Gynostemma pentaphyllum before and after heat processing based on UPLC-Q-Trap-MS].

Heat-processed Gynostemma pentaphyllum has strong biological activity, and saponins are the main components. To investigate the changes of saponins in G. pentaphyllum before and after heat processing, the present study determined and analyzed the content of nine saponins in G. pentaphyllum from Zhangzhou of Fujian and Jinxiu of Guangxi by ultra-high performance liquid chromatography with quadrupole ion-trap mass spectrometry(UPLC-Q-Trap-MS). The separation of the analytes was performed on an ACQUITY UPLC BEH C_(18) column(2.1 mm×50 mm, 1.7 µm) at 30 ℃, with acetonitrile and 0.1% formic acid in water as the mobile phase by gradient elution, and the flow rate was 0.3 mL·min~(-1). Quantitative analysis was performed using electrospray ionization source(ESI) in the multiple reaction-monitoring(MRM) mode. The results showed that the content of saponins with biological activities increased after heat processing. Specifically, gypenoside L, gypenoside LI, damulin A, damulin B, ginsenoside Rg_3(S), and ginsenoside Rg_3(R) in G. pentaphyllum produced in Zhangzhou of Fujian increased by 7.369, 8.289, 12.155, 7.587, 0.929, and 1.068 µg·g~(-1), respectively, while the content of ginsenoside Rd, gypenoside LVI, and gypenoside XLVI, which were abundant in the raw materials, decreased by 0.779, 19.37, and 9.19 µg·g~(-1), respectively. The content of gypenoside L, gypenoside LI, damulin A, damulin B, ginsenoside Rg_3(S), and ginsenoside Rg_3(R) in G. pentaphyllum produced in Jinxiu of Guangxi increased by 0.100, 0.161, 0.317, 0.228, 3.280, and 3.395 µg·g~(-1), respectively, while the content of ginsenoside Rd, gypenoside LVI, and gypenoside XLVI in the raw materials was reduced by 1.661, 0.014, and 0.010 µg·g~(-1), respectively. The results suggest that heat processing is an effective way to transform rare gypenosides. Furthermore, it is found that there are great differences in the content of gypenosides in different regions.

The effects of tea plants-soybean intercropping on the secondary metabolites of tea plants by metabolomics analysis.

BACKGROUND: Intercropping, especially with legumes, as a productive and sustainable system, can promote plants growth and improves the soil quality than the sole crop, is an essential cultivation pattern in modern agricultural systems. However, the metabolic changes of secondary metabolites and the growth in tea plants during the processing of intercropping with soybean have not been fully analyzed. RESULTS: The secondary metabolomic of the tea plants were significant influence with intercropping soybean during the different growth stages. Especially in the profuse flowering stage of intercropping soybean, the biosynthesis of amino acids was significantly impacted, and the flavonoid biosynthesis, the flavone and flavonol biosynthesis also were changed. And the expression of metabolites associated with amino acids metabolism, particularly glutamate, glutamine, lysine and arginine were up-regulated, while the expression of the sucrose and D-Glucose-6P were down-regulated. Furthermore, the chlorophyll photosynthetic parameters and the photosynthetic activity of tea plants were higher in the tea plants-soybean intercropping system. CONCLUSIONS: These results strengthen our understanding of the metabolic mechanisms in tea plant's secondary metabolites under the tea plants-soybean intercropping system and demonstrate that the intercropping system of leguminous crops is greatly potential to improve tea quality. These may provide the basis for reducing the application of nitrogen fertilizer and improve the ecosystem in tea plantations.

Uncovering the anti-NSCLC effects and mechanisms of gypenosides by metabolomics and network pharmacology analysis.

ETHNOPHARMACOLOGICAL RELEVANCE: Lung cancer is the chief reason of cancer death worldwide, and non-small cell lung cancer (NSCLC) make up the majority of lung cancers. Gypenosides are the main active constituents from Gynostemma pentaphyllum. Previous studies showed that they were used to remedy many cancers. The effect of gypenosides on NSCLC has never been studied from the perspective of network pharmacology and metabolomics. The mechanism is still not clear and remains to be explored. AIM OF THE STUDY: To explore the anti-NSCLC activity and mechanism of gypenosides in A549 cells. MATERIAL/METHODS: Gypenosides of G. pentaphyllum were detected by HPLC-MS. The cytotoxicity was detected by MTT assay. The migration, cell cycle and apoptosis of gypenosides were studied by wound healing assay, JC-1 assay and flow cytometry. The mechanism of gypenosides on NSCLC was studied by metabolomics and network pharmacology. Some key proteins and pathways were further confirmed by Western blot. RESULTS: Eleven gypenosides were detected by HPLC-MS. Gypenosides could suppress the proliferation of A549 cells, inhibit the migration of A549 cells, induce apoptosis and arrest cell cycle in G0/G1 phase. Metabolomics and network pharmacology approach revealed that gypenosides might affect 17 metabolite related proteins by acting on 9 candidate targets (STAT3, VEGFA, EGFR, MMP9, IL2, TYMS, FGF2, HPSE, LGALS3), thus resulting in the changes of two metabolites (uridine 5'-monophosphate, D-4'-Phosphopantothenate) and two metabolic pathways (pyrimidine metabolism; pantothenate and CoA biosynthesis). Western blotting indicated that gypenosides might inhibit A549 cells through MMP9, STAT3 and TYMS to indirectly affect the pathways of pyrimidine metabolism, pantothenate and CoA biosynthesis. CONCLUSIONS: This study revealed that metabolomics combined with network pharmacology was conducive to understand the anti-NSCLC mechanism of gypenosides.

Gypenoside LI arrests the cell cycle of breast cancer in G0/G1 phase by down-regulating E2F1.

ETHNOPHARMACOLOGICAL RELEVANCE: Gynostemma pentaphyllum (Thunb.) Makino, a traditional medicine in China, has been widely used for the treatment of various diseases. Gypenoside LI (Gyp LI) is a major constituent from steamed G. pentaphyllum. Previous studies have shown that gypnenoside LI possess inhibitory effect on the growth of many cancer cells. However, its pharmacological effect in breast cancer and the mechanism have not been reported yet. AIM OF THE STUDY: To investigate the anti-breast cancer activity of gypenoside LI and underlying mechanisms of gypenoside LI in MDA-MB-231 and MCF-7 cells. MATERIAL/METHODS: The cytotoxicity of gypenoside LI was determined by MTT, colony-formation and three-dimensional spheroid assay. The migration, cell apoptosis and the cell cycle were investigated through cell morphology observation, flow cytometry analysis and key proteins detection. The anticancer mechanisms of gypenoside LI were detected by RNA sequencing (RNA-seq) and Gene Set Enrichment Analysis (GSEA) transcriptome analysis. RESULTS: Gypenoside LI inhibited cell proliferation, migration, induced cell apoptosis and cell cycle arrest. Gypenoside LI arrested cell cycle at G0/G1 phase by regulating E2F1. It also inhibited tumor proliferation by regulating the expression of ERCC6L. Interestingly, we found that E2F1 siRNA also down-regulated the expression of ERCC6L. Gypenoside LI showed potential anti-breast cancer cells activity, especially on triple-negative breast cancer cells. CONCLUSIONS: These data indicate that gypenoside LI could inhibit human breast cancer cells through inhibiting proliferation and migration, inducing apoptosis, arresting cell cycle at G0/G1 phase by regulating E2F1. It could be used as potential multi-target chemopreventive agents for cancer.

Mechanism of gypenosides of Gynostemma pentaphyllum inducing apoptosis of renal cell carcinoma by PI3K/AKT/mTOR pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Gynostemma pentaphyllum (Thunb.) Makino is a traditional medicine commonly used in China, East Asia and Southeast Asia. In clinic, it is mainly used for hyperlipidemia and antitumor. Its antitumor activity was first recorded in "Illustrated Catalogue of Plants". Gypenosides were the main active ingredients of G. pentaphyllum. The anticancer activity of gypenosides in vivo and in vitro had been widely reported. However, the mechanism of gypenosides in renal cell carcinoma (RCC) still unclear. AIM OF THE STUDY: In this study, we tried to investigate the active constituents from G. pentaphyllum and potential mechanisms in RCC treatment through network pharmacology and in vitro experiments. MATERIAL/METHODS: Active compounds and their targets were evaluated and screened through TCMSP and Swiss Target Prediction database. Notably, nine preliminary screened components obtained from database were identified by LC-MS and LC-MS/MS. The targets associated with RCC were obtained from OMIM, TTD and GeneCards database. The PPI network and active component/target/pathway networks were constructed to identify the potential drug targets using String database and Cytoscape software. The functions and pathways of targets were analyzed through DAVID database. Finally, AutoDockTools 1.5.6 was used for molecular docking to assess the binding ability between compounds and targets. To support our prediction, we then explore the antitumor effect and mechanism of gypenosides by vitro experiments. CCK8 and flow cytometry were performed to evaluate cell death treated with gypenosides. Quantitative real-time PCR and Western blot were conducted to detect the changes of PI3K/AKT/mTOR signaling pathway. RESULTS: Nine saponins and 68 targets have been screened. The hub targets covered PIK3CA, VEGFA, STAT3, JAK2, CCND1 and MAPK3. Enrichment analysis showed that the pathways mainly contained PI3K/Akt/mTOR, HIF-1, TNF, JAK-STAT and MAPK signaling pathways. Gypenosides extracted from G. pentaphyllum showed strong activity against 786-O and Caki-1 cells, and cell apoptosis were detected through Annexin V/PI dual staining assay. RT-qPCR showed that gypenosides downregulated the levels of PIK3CA, Akt and mTOR in Caki-1 and 786-O cells. Mechanistically, gypenosides induced apoptosis of RCC cells through regulating PI3K/Akt/mTOR signaling pathway which was implemented though decreasing the phosphorylation level of Akt and mTOR. CONCLUSIONS: Gypenosides induced apoptosis of RCC cells by modulating PI3K/Akt/mTOR signaling pathway.

Effectiveness of acupuncture combined mecobalamin in the treatment of elderly diabetic peripheral neuropathy: A protocol of systematic review and meta-analysis.

BACKGROUND: Although previous studies have reported the effectiveness of acupuncture combined mecobalamin (AM) in the treatment of elderly diabetic peripheral neuropathy (EDPN), no systematic study has assessed its effectiveness and safety. Thus, this study will evaluate the effectiveness and safety of AM for the treatment of patients with EDPN. METHODS: Bibliographic electronic databases will be searched as follows: Cochrane Library, PUBMED, EMBASE, CINAHL, PsycINFO, WANGFANG, and China National Knowledge Infrastructure. All of them will be searched from each database initial to March 1, 2020 without language restrictions. All study selection, information extracted, and study quality evaluation will be performed by 2 independent authors. Any disagreements between 2 authors will be resolved by a third author via discussion. RevMan 5.3 software will be used for data pooling and meta-analysis performance if it is possible. RESULTS: This study will provide synthesis of current evidence of AM for patients with EDPN through primary outcome of glycemic profile, and secondary of neuropathic pain intensity, plantar tactile sensitivity, sensory nerve conduction velocity and motor nerve conduction velocity, health-related quality of life, and adverse events. CONCLUSION: This study will provide helpful reference for the efficacy and safety of AM for the treatment of patients with EDPN to the clinicians and further studies.Study registration number: INPLASY202040094.

[Responses of shallow soil water content in Artemisia ordosica community to different rainfall patterns]. / æ²¹è’¿çŒä¸›ç¾¤è½æµ å±‚åœŸå£¤æ°´åˆ†å¯¹ä¸åŒé™é›¨æ ¼å±€çš„å“åº”.

The shrub species, Artemisia ordosica, commonly occurs in the eastern Hobq desert. Here, we used a micrometeorological observation system to continuously monitor the rainfall and soil water content in 0-10, 10-30, and 30-50 cm soil layers during the growing season from 2016 to 2018. The dynamic spatial and temporal changes in soil water content under different rainfall patterns were examined, and the replenishing effects of rainfall events on soil water content and water infiltration characteristics were analyzed. The results showed that soil water content of the surface layer in the A. ordosica community had significant seasonal and vertical variation under rainfall fluctuation. Rainfall amount and soil water content before rain were the main factors controlling soil water replenishment and infiltration. The soil surface layer (0-10 cm) was sensitive to rainfall, and the rainfall of 3.8 mm began to replenish this layer. The responses of 10-30 cm soil layer to rainfall was slower, more than 8.6 mm rainfall being needed for effective replenishment. The response of the 30-50 cm soil layer to rainfall was even more delayed, and replenishment at this depth could not be achieved until the rainfall exceeded 11.8 mm. The water infiltration rate increased with rainfall amount and decreased with soil depth, while water infiltration depth was positively correlated with the rainfall amount and soil water content before rainfall. During the study period, rainfall of <10 mm occurred predominantly, accounting for 78.4% of the total rainfall events. The rainfall mainly replenished soil layer above 30 cm, and the replenishment of deep soil was very limited, which was not conducive to the growth of deep-rooted species. Therefore, rainfall patterns directly affected the composition, distribution, and succession of plant communities in this area.