Integrating plasma metabolomics and gut microbiome to reveal the mechanisms of Huangqi Guizhi Wuwu Decoction intervene diabetic peripheral neuropathy.

ETHNOPHARMACOLOGICAL RELEVANCE: Huangqi Guizhi Wuwu Decoction (HGWD) is a classic traditional Chinese herbal formula from "Synopsis of Golden Chamber," which is used to treat blood stagnation and has been used for alleviating diabetic peripheral neuropathy (DPN) in the clinic. However, the mechanisms of HGWD intervention DPN are still to be discovered. AIM OF THE STUDY: This study aims to explore the mechanism of HGWD intervention DPN by integrating plasma metabolomics and gut microbiome. MATERIALS AND METHODS: BKS Cg-m+/+Leprdb/J (db/db) mice with DPN were at 16 weeks of age. The indices of DPN phenotypes in db/db mice, pathomorphology of the sciatic nerve, intraepithelial nerve fibers (IENF) of the foot pad, levels of blood lipids and oxidative stress, and inflammatory reaction were used to appraise the HGWD efficacy. Finally, the pharmacological mechanisms of HGWD intervening DPN were explored by metabolomics and 16S rRNA gene sequencing. RESULTS: HGWD reversed DPN phenotypes in db/db mice, improved peripheral nerve structure, ameliorated the level of blood lipids and nerve growth factor in plasma, enhanced antioxidant capacity, and alleviated inflammatory responses. Plasma metabolomics disclosed that HGWD remarkably regulated the unusual levels of thirty-seven metabolites involved in sphingolipid metabolism, biosynthesis of unsaturated fatty acids, arachidonic acid metabolism, and amino acid biosynthesis pathways. The gut microbiome showed that nine bacteria were highly correlated with the efficacy of HGWD in DPN. Integrating analysis of microbiome and metabolomics demonstrated that the interaction of four bacteria with four metabolic pathways might be the significant mechanism of HGWD intervention in DPN. CONCLUSIONS: The mediation of gut microbiota and plasma metabolism may be the potential mechanism of HGWD ameliorating DPN in db/db mice. The interaction of Lactobacillus, Alloprevotella, Bacteroides, and Desulfovibio with four metabolic pathways might be the critical mechanism for HGWD treating DPN.

[Diosgenin alleviates NAFLD induced by a high-fat diet in rats via mTOR/SREBP-1c/HSP60/MCAD/SCAD signaling pathway].

This study aims to observe the effects of diosgenin on the expression of mammalian target of rapamycin(mTOR), sterol regulatory element-binding protein-1c(SREBP-1c), heat shock protein 60(HSP60), medium-chain acyl-CoA dehydrogenase(MCAD), and short-chain acyl-CoA dehydrogenase(SCAD) in the liver tissue of the rat model of non-alcoholic fatty liver disease(NAFLD) and explore the mechanism of diosgenin in alleviating NAFLD. Forty male SD rats were randomized into five groups: a control group, a model group, low-(150 mg·kg~(-1)·d~(-1)) and high-dose(300 mg·kg~(-1)·d~(-1)) diosgenin groups, and a simvastatin(4 mg·kg~(-1)·d~(-1)) group. The rats in the control group were fed with a normal diet, while those in the other four groups were fed with a high-fat diet. After feeding for 8 weeks, the body weight of rats in the high-fat diet groups increased significantly. After that, the rats were administrated with the corresponding dose of diosgenin or simvastatin by gavage every day for 8 weeks. The levels of triglyceride(TG), total cholesterol(TC), alanine transaminase(ALT), and aspartate transaminase(AST) in the serum were determined by the biochemical method. The levels of TG and TC in the liver were measured by the enzyme method. Oil-red O staining was employed to detect the lipid accumulation, and hematoxylin-eosin(HE) staining to detect the pathological changes in the liver tissue. The mRNA and protein levels of mTOR, SREBP-1c, HSP60, MCAD, and SCAD in the liver tissue of rats were determined by real-time fluorescence quantitative polymerase chain reaction(RT-qPCR) and Western blot, respectively. Compared with the control group, the model group showed increased body weight, food uptake, liver index, TG, TC, ALT, and AST levels in the serum, TG and TC levels in the liver, lipid deposition in the liver, obvious hepatic steatosis, up-regulated mRNA and protein expression levels of mTOR and SREBP-1c, and down-regulated mRNA and protein expression levels of HSP60, MCAD, and SCAD. Compared with the model group, the rats in each treatment group showed obviously decreased body weight, food uptake, liver index, TG, TC, ALT, and AST levels in the serum, TG and TC levels in the liver, lessened lipid deposition in the liver, ameliorated hepatic steatosis, down-regulated mRNA and protein le-vels of mTOR and SREBP-1c, and up-regulated mRNA and protein levels of HSP60, MCAD, and SCAD. The high-dose diosgenin outperformed the low-dose diosgenin and simvastatin. Diosgenin may prevent and treat NAFLD by inhibiting the expression of mTOR and SREBP-1c and promoting the expression of HSP60, MCAD, and SCAD to reduce lipid synthesis, improving mitochondrial function, and promoting fatty acid ß oxidation in the liver.

ITRAQ-based proteomics analysis of human ectopic endometrial stromal cells treated by Maqian essential oil.

BACKGROUND: Endometriosis is a common and complex syndrome characterized by the presence of endometrial-like tissue outside the uterus. Chinese medicine has been recently found to show good efficacy in treating endometriosis. Our previous results revealed that Maqian fruit essential oil (MQEO) could inhibit the proliferation and induce apoptosis of ectopic endometrial stromal cells (EESCs), but the mechanisms remain unclear. In this study, we aim to explore the molecular mechanism of MQEO's specific effects in EESCs. METHODS: We conducted a quantitative proteomics analysis by iTRAQ on EESCs treated with MQEO or DMSO. Then deep analysis was performed based on differentially expressed proteins, including Gene Ontology enrichment analysis, pathway enrichment analysis and protein interaction analysis. Candidate protein targets were subsequently verified by western blotting. RESULTS: Among 6575 identified proteins, 435 proteins exhibited altered expression levels in MQEO-treated EESCs. Of these proteins, most were distributed in signal transduction as well as immune system and the most significantly altered pathway was complement and coagulation cascades. Moreover, two differentially expressed proteins (Heme oxygenase 1 and Acyl-CoA 6-desaturase) were verified and they can be potential biomarkers for endometriosis treatment. CONCLUSIONS: Our proteomic analysis revealed distinct protein expression patterns induced by MQEO treatment in EESCs, highlighting the potential of MQEO for endometriosis treatment and biomarker discovery.

The hidden costs of emotional labor on withdrawal behavior: the mediating role of emotional exhaustion, and the moderating effect of mindfulness.

BACKGROUND: Employees' withdrawal behavior concerns organization leaders and policymakers in many countries. However, the specific mechanism which emotional labor affects withdrawal behavior has yet to be thoroughly discussed. There needs to be systematic research on how different emotional labor strategies affect work withdrawal, whether directly or through individual perception, and how to respond. METHODS: A total of 286 hotel and catering service employees participated in our study. A series of hierarchical moderated regression analyses were performed to test the hypothesis. RESULTS: The results indicated that surface acting positively affected withdrawal behavior, while deep acting had a negative effect. Emotional exhaustion mediated in this relationship of surface acting with withdrawal behavior and deep acting with withdrawal behavior. Mindfulness showed moderation effects between emotional exhaustion and withdrawal behavior. CONCLUSIONS: Emotional labor and emotional exhaustion are significant in predicting employees' intentions to withdraw, given that emotional exhaustion partially mediates the effects of emotional labor on withdrawal behavior. Significantly, the relationship between emotional exhaustion and withdrawal behavior is weakened by mindfulness.

[Mechanism of heat-clearing prescriptions in alleviating type 2 diabetes mellitus:a review].

Type 2 diabetes mellitus(T2DM), a common chronic metabolic disease, is often accompanied by internal heat syndrome. Heat-clearing prescriptions are widely used to treat different heat syndromes of T2DM from the aspects of clearing stagnant heat, excess heat, damp heat, phlegm heat, and heat toxin, demonstrating remarkable effects. The mechanism of blood sugar-lowering agents has always been a hotspot of research. Recently, the basic studies of heat-clearing prescriptions from different perspectives have been increasing year by year. To clarify the mechanisms of heat-clearing prescriptions and find specific mechanisms, we systematically reviewed the basic studies of heat-clearing prescriptions commonly used for the treatment of T2DM in the past decade, intending to provide a reference for related research.

[Effect of diosgenin on mTOR/FASN/HIF-1α/VEGFA expression in rats with non-alcoholic fatty liver disease].

The present study aimed to investigate the effect of diosgenin on mammalian target of rapamycin(mTOR), fatty acid synthase(FASN), hypoxia inducible factor-1α(HIF-1α), and vascular endothelial growth factor A(VEGFA) expression in liver tissues of rats with non-alcoholic fatty liver disease(NAFLD) and explore the mechanism of diosgenin on lipogenesis and inflammation in NAFLD. Forty male SD rats were divided into a normal group(n=8) fed on the normal diet and an experimental group(n=32) fed on the high-fat diet(HFD) for the induction of the NAFLD model. After modeling, the rats in the experimental group were randomly divided into an HFD group, a low-dose diosgenin group(150 mg·kg~(-1)·d~(-1)), a high-dose diosgenin group(300 mg·kg~(-1)·d~(-1)), and a simvastatin group(4 mg·kg~(-1)·d~(-1)), with eight rats in each group. The drugs were continuously given by gavage for eight weeks. The levels of triglyceride(TG), total cholesterol(TC), low-density lipoprotein cholesterol(LDL-C), alanine transaminase(ALT), and aspartate transaminase(AST) in the serum were detected by the biochemical method. The content of TG and TC in the liver was detected by the enzyme method. Enzyme-linked immunosorbent assay(ELISA) was used to measure interleukin 1ß(IL-1ß) and tumor necrosis factor α(TNF-α) in the serum. Lipid accumulation in the liver was detected by oil red O staining. Pathological changes of liver tissues were detected by hematoxylin-eosin(HE) staining. The mRNA and protein expression levels of mTOR, FASN, HIF-1α, and VEGFA in the liver of rats were detected by real-time fluorescence-based quantitative polymerase chain reaction(PCR) and Western blot, respectively. Compared with the normal group, the HFD group showed elevated body weight and levels of TG, TC, LDL-C, ALT, AST, IL-1ß, and TNF-α(P<0.01), increased lipid accumulation in the liver(P<0.01), obvious liver steatosis, up-regulated mRNA expression levels of mTOR, FASN, HIF-1α, and VEGFA(P<0.01), and increased protein expression levels of p-mTOR, FASN, HIF-1α, and VEGFA(P<0.01). Compared with the HFD group, the groups with drug treatment showed lowered body weight and levels of TG, TC, LDL-C, ALT, AST, IL-1ß, and TNF-α(P<0.05, P<0.01), reduced lipid accumulation in the liver(P<0.01), improved liver steatosis, decreased mRNA expression levels of mTOR, FASN, HIF-1α, and VEGFA(P<0.05, P<0.01), and declining protein expression levels of p-mTOR, FASN, HIF-1α, and VEGFA(P<0.01). The therapeutic effect of the high-dose diosgenin group was superior to that of the low-dose diosgenin group and the simvastatin group. Diosgenin may reduce liver lipid synthesis and inflammation and potentiate by down-regulating the mTOR, FASN, HIF-1α, and VEGFA expression, playing an active role in preventing and treating NAFLD.

Sculponeatin A promotes the ETS1-SYVN1 interaction to induce SLC7A11/xCT-dependent ferroptosis in breast cancer.

BACKGROUND: E26 transformation specificity-1 (ETS1) is a transcription factor that is overexpressed in breast cancer (BC) and promotes tumor progression. Sculponeatin A (stA), a new diterpenoid extracted from Isodon sculponeatus, has no reported antitumor mechanism. PURPOSE: Here, we explored the antitumor activity of stA in BC and further clarified its mechanism. METHODS: Ferroptosis was detected by flow cytometric, glutathione, malondialdehyde, and iron determination assays. The effect of stA on the upstream signaling pathway of ferroptosis was detected by Western blot, gene expression, gene alterations and other approaches. The binding of stA and ETS1 was examined through a microscale thermophoresis assay and a drug affinity responsive target stability assay. An in vivo mouse model experiment was performed to evaluate the therapeutic and potential mechanism of stA. RESULTS: stA exhibits therapeutic potential in BC by inducing SLC7A11/xCT-dependent ferroptosis. stA decreases the expression of ETS1, which is responsible for xCT-dependent ferroptosis in BC. stA inhibits the transcriptional expression of xCT by directly binding to the ETS domain of the ETS1 protein. In addition, stA promotes proteasomal degradation of ETS1 by triggering ubiquitin ligase synoviolin 1 (SYVN1)-mediated ubiquitination. The K318 site of ETS1 mediates ubiquitination of ETS1 by SYVN1. In a mouse model, stA inhibits tumor growth without causing obvious toxicity. CONCLUSION: Taken together, the results confirm that stA promotes the ETS1-SYVN1 interaction to induce ferroptosis in BC mediated by ETS1 degradation. stA is expected to be used in research of candidate drugs for BC and drug design based on ETS1 degradation.

Clinical efficacy and complications of transurethral resection of the prostate versus plasmakinetic enucleation of the prostate.

BACKGROUND: Benign prostatic hyperplasia (BPH) is a common disease in elderly males, and many kinds of minimally invasive procedures can be used for the treatment of BPH. However, various procedures have caused some controversies regarding clinical outcomes, so more studies are needed to validate these controversial topics. AIMS: This study aimed to explore differences of clinical efficacy, surgical features, and complications between transurethral resection of the prostate (TURP) and plasmakinetic enucleation of the prostate (PKEP) for BPH. METHODS: A total of eligible 850 cases of BPH underwent TURP (the TURP group, 320 cases) or PKEP (the PKEP group, 530 cases) in the urology department of our hospital from March 2015 to 2018 were involved in this study. Then, the baseline data, surgical characteristics, IPSS, QoL, PVR, Qmax, IIEF-5, and documented complications were compared between the two groups. RESULTS: The operative time, intraoperative irrigation volume, postoperative hemoglobin, decrease in hemoglobin, postoperative irrigation time and volume, catheterization time, and hospital stay of the PKEP group were significantly less than those of the TURP group (all P < 0.05). At 3 months, 1, 2, and 3 years after operation, no significant differences were observed in IPSS, QoL, PVR, but the results of Qmax and IIEF-5 in the PKEP group were significantly higher than those parameters in the TURP group (all P < 0.05). The incidences of massive blood loss, postoperative secondary bleeding, blood transfusion, capsular perforation, urinary tract irritation, bladder spasm, clot retention, urinary tract infection, transient incontinence, erectile dysfunction, and the incidences of II, III grade of Clavien-Dindo classification in the PKEP group were significantly lower than those of the TURP group (all P < 0.05). CONCLUSION: The clinical efficacy of PKEP is compared favorably with TURP during midterm follow-up. Given the merits such as less blood loss and hospital stay, lower complications, PKEP should be given a priority for BPH.

Predation effect on copepods by the giant jellyfish Nemopilema nomurai during the early occurrence stage in May in the northern East China Sea and southern Yellow Sea, China.

Massive blooms of Nemopilema nomurai have occurred recently across East Asian waters. They are potentially important as zooplankton predators, as well as being competitors for prey with zooplanktivorous fish. Few studies have estimated the predation effects on zooplankton by N. nomurai in situ. To quantify the natural diets and feeding rates and estimate the predation effects, N. nomurai medusae were collected in the northern East China Sea and southern Yellow Sea, one of the principal nursery grounds of this jellyfish, during May 2019. The gut contents indicated that copepods were an important food source for N. nomurai; copepods <1000 µm represented the bulk of total prey intake in number (> 99 %). Linear regression analyses showed that the copepods number in the gut contents was significantly influenced by medusa diameter and prey abundance. Calculations using the above data indicated that one medusa (mean diameter: 26.06 ± 9.73 cm) consumed approximately 5248 ± 2768 of copepods daily. However, even the maximum predation pressure was <0.1 % of the total copepods standing stock daily due to the small diameter and low density/biomass of N. nomurai medusae in May 2019. The data presented here suggested that the predation effects of N. nomurai on copepods were low and might not reduce prey availability to fish with diets consisting mainly of copepods during the early occurrence stage of the N. nomurai population.

Mechanism of heat-clearing prescriptions in alleviating type 2 diabetes mellitus:a review / 中国中药杂志

Type 2 diabetes mellitus(T2DM), a common chronic metabolic disease, is often accompanied by internal heat syndrome. Heat-clearing prescriptions are widely used to treat different heat syndromes of T2DM from the aspects of clearing stagnant heat, excess heat, damp heat, phlegm heat, and heat toxin, demonstrating remarkable effects. The mechanism of blood sugar-lowering agents has always been a hotspot of research. Recently, the basic studies of heat-clearing prescriptions from different perspectives have been increasing year by year. To clarify the mechanisms of heat-clearing prescriptions and find specific mechanisms, we systematically reviewed the basic studies of heat-clearing prescriptions commonly used for the treatment of T2DM in the past decade, intending to provide a reference for related research.

[Ethoxysanguinarine directly targets CIP2A to inhibit proliferation and induce autophagy of SGC7901/DDP cells].

This study aims to investigate the effect of ethoxysanguinarine(Eth) on cisplatin(DDP)-resistant human gastric cancer cells and decipher the underlying mechanism. The human gastric cancer cell line SGC7901 and the DDP-resistant cell line SGC7901/DDP were used as the cell models. Western blot was employed to determine the expression levels of multidrug resistance-related proteins, and methyl thiazolyl tetrazolium(MTT) assay to detect the proliferation of SGC7901 and SGC7901/DDP cells exposed to DDP. After treatment with different concentrations of Eth, the proliferation of SGC7901 and SGC7901/DDP cells was detected by MTT assay, trypan blue exclusion assay, colony formation assay, and high-content imaging and analysis system. The apoptosis of SGC7901/DDP cells was detected by flow cytometry with Annexin V-FITC/PI staining. GFP-LC3 transfection was carried out to detect the effect of Eth on the autophagy of SGC7901/DDP cells. The expression levels of the multidrug resistance-related protein P-glycoprotein(P-gp), the apoptosis-related proteins [caspase-9, caspase-3, and poly(ADP-ribose) polymerase(PARP)], the autophagy-related protein light chain 3-Ⅱ(LC3-Ⅱ), the key effectors [mammalian target of rapamycin(mTOR), 70 kDa ribosomal protein S6 kinase(P70 S6 K), and 4 E binding protein 1(4 E-BP1)] of the mammalian target of rapamycin complex 1(mTORC1) signaling pathway, cancerous inhibitor of protein phosphatase 2A(CIP2A), and protein kinase B(Akt) were measured by Western blot. The mRNA level of CIP2A in the SGC7901/DDP cells exposed to Eth for 24 h was analyzed by RT-qPCR. After SGC7901/DDP cells were transfected with CIP2A expression vector pcDNA3.1-HA-CIP2A and treated with different concentrations of Eth, MTT assay was used to determine the prolife-ration of SGC7901/DDP cells and Western blot to detect the expression levels of related proteins. The interaction sites of Eth and CIP2A were predicted by molecular docking. The affinity between Eth and CIP2A was determined by drug affinity responsive target stability(DARTS) assay. The pharmacokinetic properties and drug-like activity of Eth were predicted by SwissADME. The results indicated that SGC7901/DDP cells were more sensitive to Eth than SGC7901 cells. Eth significantly inhibited proliferation and colony formation and changed the morphology, roundness, and area of SGC7901/DDP cells. Eth treatment caused the nucleus shrinking and significantly increased the apoptosis rate of the cells. Furthermore, Eth down-regulated the expression of caspase-9 and caspase-3 precursors and promoted the cleavage of PARP, which suggested that Eth induced the apoptosis of SGC7901/DDP cells. The GFP-LC3 in Eth-treated cells showed speckled aggregation. The up-regulated expression of LC3-Ⅱ by Eth indicated that Eth activated the autophagy of SGC7901/DDP cells. Eth down-regulated the expression of P-gp, the phosphorylation of mTOR, P70 S6K, and 4E-BP1, the expression of CIP2A, and the phosphorylation of Akt. Additionally, it increased the activity of PP2A, and had no significant effect on the expression of CIP2A in SGC7901/DDP cells. CIP2A overexpression antagonized the inhibition of cell proliferation and the activation of autophagy by Eth. Molecular docking suggested that Eth bound to CIP2A. The results of DARTS assay further proved the above binding effect. Eth has potential drug-like activity. The above results demonstrated that Eth inhibited the proliferation, induced the apoptosis, and activated the autophagy of SGC7901/DDP cells by targeting CIP2A and then down-regulating PP2A/mTORC1 signaling pathway. This study provided a new target for the treatment of cisplatin-resistant gastric cancer.

Higher Circulating Vitamin D Levels Are Associated With Decreased Migraine Risk: A Mendelian Randomization Study.

Background: Evidence showed the supplementation of vitamin D might have beneficial effects for migraine patients. We aimed to investigate the causal effects of serum vitamin D levels on migraine risk using two-sample Mendelian randomization (MR) method. Methods: A total of 184 independent genetic instruments for serum vitamin D levels were selected from a study in 417,580 Europeans from UK biobank. Six variants from an independent study were obtained to perform replication analysis. Summary-level data for migraine were obtained from three studies with 48,975 migraine cases, 28,852 migraine cases and 10,536 migraine cases, respectively. Results: The estimated odds ratios (ORs) per standard deviation increase in circulating vitamin D levels based on the three migraine datasets were 0.948 (95% CI = 0.883-1.016, p = 0.133), 0.902 (95% confidence intervals [CI] = 0.825-0.986, p = 0.023), and 0.880 (95% CI = 0.786-0.984, p = 0.025), respectively. Using pooled migraine summary data with no sample overlap, MR analysis showed per standard deviation increase in circulating vitamin D levels was significantly associated with a decreased migraine risk (OR = 0.916, 95% CI = 0.859-0.977, p = 0.008). Multivariable MR analyses, sensitivity analyses and replication analysis confirmed the association. MR analyses showed similar estimates for migraine with aura and migraine without aura but with wider 95% CIs. Mediation analysis showed the effect of vitamin D on migraine risk via pathway of serum calcium was corresponding to an OR of 1.003 (95% CI = 1.001-1.005) and a proportion mediated of 3.42%. The reverse MR analysis showed migraine might not affect vitamin D levels. Conclusion: This two-sample MR study showed genetically determined increased circulating vitamin D levels are associated with decreased migraine risk. The effect seems consistent across different migraine subtypes. In addition, the role of serum calcium in mediating the association between vitamin D and migraine is negligible. Future large well-designed randomized trials are warranted to assess the effects of vitamin D supplementation for migraine patients, especially in those with vitamin D deficiency.

[The effects of different herbal compound and extracts from different extraction methods on hypoxia tolerance in mice].

Objective: To investigate the effects of different prescription compositions of traditional Chinese medicine and its different extraction methods of compound formula extracts on hypoxia tolerance in mice, in order to preferably select their prescription compositions and preparation extraction methods. Methods: Male BALB/c mice were randomly divided into 6 groups: blank control group, compound danshen group, compound Rhodiola Rosea alcohol-water extract group (Rhodiola rosea, Astragali Radix, Polygonati Rhizoma, Lycii Fructus), compound Rhodiola Rosea water extract group, compound Astragalus alcohol-water extract group (Astragali Radix, Polygonati Rhizoma, Lycii Fructus) and compound Astragalus water extract group, 30 mice in each group. Each group was administered continuously by gavage for 10 d. The blank group was gavaged with sterilized injection water. The mice in the other groups were treated with 0.15 g/kg of compound danshen, 3 g/kg of compound Rhodiola Rosea alcohol-water extract or water extract, and 1.7 g/kg of compound Astragalus alcohol-water extract or water extract, respectively. Each group was subjected to normobaric hypoxia tolerance test, sodium nitrite toxicity survival test and acute cerebral ischemia-hypoxia test 1 h after the last gavage, and the mice brain tissues were used to determine the activity of antioxidant enzymes and metabolites related to oxidative stress. Results: Compared with the blank control group, in normobaric hypoxia tolerance test, the survival time of mice in the compound danshen group and the compound Astragalus alcohol-water extract group and water extraction group was prolonged significantly (P＜0.01), and the number of open-mouth gasping after cerebral ischemia and hypoxia was increased significantly (P＜0.05). There was no statistical difference in survival time after sodium nitrite injection in each group. Compared with the blank control group, the activities of T-AOC, SOD, GSH and CAT were increased significantly (P＜0.05, P＜0.01) and the content of MDA was decreased significantly (P＜0.01) in the compound Astragalus water extract group. Compared with the compound danshen group, the activities of SOD, CAT and GSH were increased significantly (P＜0.01, P＜0.05) and the content of MDA was decreased significantly (P＜0.05). Conclusion: Compound Astragalus water extraction has the best effect of hypoxia tolerance, compound Rhodiola Rosea can eliminate Rhodiola rosea and consists of Astragali Radix, Polygonati Rhizoma, Lycii Fructus and its extraction method is water extraction.

Preparation of Tea Aroma Precursor Glycosides: An Efficient and Sustainable Approach via Chemical Glycosidation.

Tea aroma precursor glycosides are plant-derived natural products with great economic value. However, the preparation of these glycosides remains largely overlooked in the past decades. Herein, we report a mild, efficient, and sustainable chemocatalytic procedure for the production of tea aroma precursor glycosides. During the study of the glycosidation, the catalysts were found to be decisive in the product formation favoring different reaction pathways; in addition, the influence of molecular sieves was elucidated. With regard to these findings, the serious problem of the competing orthoester formation side reaction was successfully overcome with low catalyst loading (1 mol %) and the use of 5 Å molecular sieves, leading to the preparation of a variety of tea aroma precursor ß-d-glucopyranosides and ß-primeverosides on a gram scale in high yields in an economical way. Taken together, the current approach features catalytic glycosidation with non-toxic and low-cost catalysts, demonstrates highly favorable greenness and sustainability, and promises industrial production of tea aroma precursor glycosides.

The genome of medicinal leech (Whitmania pigra) and comparative genomic study for exploration of bioactive ingredients.

BACKGROUND: Leeches are classic annelids that have a huge diversity and are closely related to people, especially medicinal leeches. Medicinal leeches have been widely utilized in medicine based on the pharmacological activities of their bioactive ingredients. Comparative genomic study of these leeches enables us to understand the difference among medicinal leeches and other leeches and facilitates the discovery of bioactive ingredients. RESULTS: In this study, we reported the genome of Whitmania pigra and compared it with Hirudo medicinalis and Helobdella robusta. The assembled genome size of W. pigra is 177 Mbp, close to the estimated genome size. Approximately about 23% of the genome was repetitive. A total of 26,743 protein-coding genes were subsequently predicted. W. pigra have 12346 (46%) and 10295 (38%) orthologous genes with H. medicinalis and H. robusta, respectively. About 20 and 24% genes in W. pigra showed syntenic arrangement with H. medicinalis and H. robusta, respectively, revealed by gene synteny analysis. Furthermore, W. pigra, H. medicinalis and H. robusta expanded different gene families enriched in different biological processes. By inspecting genome distribution and gene structure of hirudin, we identified a new hirudin gene g17108 (hirudin_2) with different cysteine patterns. Finally, we systematically explored and compared the active substances in the genomes of three leech species. The results showed that W. pigra and H. medicinalis exceed H. robusta in both kinds and gene number of active molecules. CONCLUSIONS: This study reported the genome of W. pigra and compared it with other two leeches, which provides an important genome resource and new insight into the exploration and development of bioactive molecules of medicinal leeches.

Coronary artery calcification and risk of mortality and adverse outcomes in patients with COVID-19: a Chinese multicenter retrospective cohort study.

Background: Coronary artery calcification (CAC) is an independent risk factor of major adverse cardiovascular events; however, the impact of CAC on in-hospital death and adverse clinical outcomes in patients with coronavirus disease 2019 (COVID-19) remains unclear. Objective: To explore the association between CAC and in-hospital mortality and adverse events in patients with COVID-19. Methods: This multicenter retrospective cohort study enrolled 2067 laboratory-confirmed COVID-19 patients with definitive clinical outcomes (death or discharge) admitted from 22 tertiary hospitals in China between January 3, 2020 and April 2, 2020. Demographic, clinical, laboratory results, chest CT findings, and CAC on admission were collected. The primary outcome was in-hospital death and the secondary outcome was composed of in-hospital death, admission to intensive care unit (ICU), and requiring mechanical ventilation. Multivariable Cox regression analysis and Kaplan-Meier plots were used to explore the association between CAC and in-hospital death and adverse clinical outcomes. Results: The mean age was 50 years (SD,16) and 1097 (53.1%) were male. A total of 177 patients showed high CAC level, and compared with patients with low CAC, these patients were older (mean age: 49 vs. 69 years, P < 0.001) and more likely to be male (52.0% vs. 65.0%, P = 0.001). Comorbidities, including cardiovascular disease (CVD) ([33.3%, 59/177] vs. [4.7%, 89/1890], P < 0.001), presented more often among patients with high CAC, compared with patients with low CAC. As for laboratory results, patients with high CAC had higher rates of increased D-dimer, LDH, as well as CK-MB (all P < 0.05). The mean CT severity score in high CAC group was also higher than low CAC group (12.6 vs. 11.1, P = 0.005). In multivariable Cox regression model, patients with high CAC were at a higher risk of in-hospital death (hazard ratio [HR], 1.731; 95% CI 1.010-2.971, P = 0.046) and adverse clinical outcomes (HR, 1.611; 95% CL 1.087-2.387, P = 0.018). Conclusion: High CAC is a risk factor associated with in-hospital death and adverse clinical outcomes in patients with confirmed COVID-19, which highlights the importance of calcium load testing for hospitalized COVID-19 patients and calls for attention to patients with high CAC. Supplementary Information: The online version contains supplementary material available at 10.1007/s42058-021-00072-4.

Paris saponin VII, a Hippo pathway activator, induces autophagy and exhibits therapeutic potential against human breast cancer cells.

Dysregulation of the Hippo signaling pathway seen in many types of cancer is usually associated with a poor prognosis. Paris saponin VII (PSVII) is a steroid saponin isolated from traditional Chinese herbs with therapeutic action against various human cancers. In this study we investigated the effects of PSVII on human breast cancer (BC) cells and its anticancer mechanisms. We showed that PSVII concentration-dependently inhibited the proliferation of MDA-MB-231, MDA-MB-436 and MCF-7 BC cell lines with IC50 values of 3.16, 3.45, and 2.86 µM, respectively, and suppressed their colony formation. PSVII (1.2-1.8 µM) induced caspase-dependent apoptosis in the BC cell lines. PSVII treatment also induced autophagy and promoted autophagic flux in the BC cell lines. PSVII treatment decreased the expression and nuclear translocation of Yes-associated protein (YAP), a downstream transcriptional effector in the Hippo signaling pathway; overexpression of YAP markedly attenuated PSVII-induced autophagy. PSVII-induced, YAP-mediated autophagy was associated with increased active form of LATS1, an upstream effector of YAP. The activation of LATS1 was involved the participation of multiple proteins (including MST2, MOB1, and LATS1 itself) in an MST2-dependent sequential activation cascade. We further revealed that PSVII promoted the binding of LATS1 with MST2 and MOB1, and activated LATS1 in the BC cell lines. Molecular docking showed that PSVII directly bound to the MST2-MOB1-LATS1 ternary complex. Microscale thermophoresis analysis and drug affinity responsive targeting stability assay confirmed the high affinity between PSVII and the MST2-MOB1-LATS1 ternary complex. In mice bearing MDA-MB-231 cell xenograft, administration of PSVII (1.5 mg/kg, ip, 4 times/week, for 4 weeks) significantly suppressed the tumor growth with increased pLATS1, LC3-II and Beclin 1 levels and decreased YAP, p62 and Ki67 levels in the tumor tissue. Overall, this study demonstrates that PSVII is a novel and direct Hippo activator that has great potential in the treatment of BC.

The use of miR122 and its target sequence in adeno-associated virus-mediated trichosanthin gene therapy.

OBJECTIVE: Plant-derived cytotoxic transgene expression, such as trichosanthin (tcs), regulated by recombinant adeno-associated virus (rAAV) vector is a promising cancer gene therapy. However, the cytotoxic transgene can hamper the vector production in the rAAV producer cell line, human embryonic kidney (HEK293) cells. Here, we explored microRNA-122 (miR122) and its target sequence to limit the expression of the cytotoxic gene in the rAAV producer cells. METHODS: A miR122 target (122T) sequence was incorporated into the 3' untranslated region of the tcs cDNA sequence. The firefly luciferase (fluc) transgene was used as an appropriate control. Cell line HEK293-mir122 was generated by the lentiviral vector-mediated genome integration of the mir122 gene in parental HEK293 cells. The effects of miR122 overexpression on cell growth, transgene expression, and rAAV production were determined. RESULTS: The presence of 122T sequence significantly reduced transgene expression in the miR122-enriched Huh7 cell line (in vitro), fresh human hepatocytes (ex vivo), and mouse liver (in vivo). Also, the normal liver physiology was unaffected by delivery of 122T sequence by rAAV vectors. Compared with the parental cells, the miR122-overexpressing HEK293-mir122 cell line showed similar cell growth rate and expression of transgene without 122T, as well as the ability to produce liver-targeting rAAV vectors. Fascinatingly, the yield of rAAV vectors carrying the tcs-122T gene was increased by 77.7-fold in HEK293-mir122 cells. Moreover, the tcs-122T-containing rAAV vectors significantly reduced the proliferation of hepatocellular carcinoma cells without affecting the normal liver cells. CONCLUSION: HEK293-mir122 cells along with the 122T sequence provide a potential tool to attenuate the cytotoxic transgene expression, such as tcs, during rAAV vector production.

Exploration of the mechanism of Zisheng Shenqi decoction against gout arthritis using network pharmacology.

BACKGROUND: In this study, the network pharmacological methods were used to predict the target of effective components of compounds in Zisheng Shenqi Decoction (ZSD, or Nourishing Kidney Qi Decoction) in the treatment of gouty arthritis (GA). METHOD: The main effective components and corresponding key targets of herbs in the ZSD were discerned through the Traditional Chinese Medicine Systems Pharmacology Database and Analysis (TCMSP), Bioinformatics Analysis Tool for Molecular mechanism of Traditional Chinese Medicine (BATMAN-TCM) database. UniProt database and Swiss Target Prediction (STP) database was used to rectify and unify the target names and supply the target information. The targets related to GA were obtained by using GeneCards database. After we discovered the potential common targets between ZSD and GA, the interaction network diagram of "ZSD-component-GA-target" was constructed by Cytoscape software (Version 3.7.1). Subsequently, the Protein-protein interaction (PPI) network of ZSD effective components-targets and GA-related targets was constructed by Search Tool for the Retrieval of Interacting Genes Database (STRING). Bioconductor package "org.Hs.eg.db" and "cluster profiler" package were installed in R software (Version 3.6.0) which used for Gene Ontology analysis and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway enrichment analysis. RESULTS: 146 components and 613 targets of 11 herbal medicines in the ZSD were got from TCMSP database and BATMAN-TCM database. 987 targets of GA were obtained from GeneCards database. After intersected and removed duplications, 132 common targets between ZSD and GA were screened out by Cytoscape software (Version 3.7.1). These common targets derived from 81 effective components of 146 components, such as quercetin, stigmasterol and kaempferol. They were closely related to anti-inflammatory, analgesic and anti oxidative stress and the principal targets comprised of Purinergic receptor P2X, ligand-gated ion channel 7 (P2x7R), Nod-like receptor protein 3 (NLRP3) and IL-1ß. GO enrichment analysis and KEGG pathway enrichment analysis by R software (Version 3.6.0) showed that the key target genes had close relationship with oxidative stress, reactive oxygen species (ROS) metabolic process and leukocyte migration in aspects of biological process, cell components and molecular function. It also indicated that ZSD could decrease inflammatory reaction, alleviate ROS accumulation and attenuate pain by regulating P2 × 7R and NOD like receptor signaling pathway of inflammatory reaction. CONCLUSION: A total of 81 effective components and 132 common target genes between ZSD and GA were screened by network pharmacology. The PPI network, GO enrichment analysis and KEGG pathway enrichment analysis suggested that ZSD can exerte anti-inflammatory and analgesic effects on the treatment of GA by reducing decreasing inflammatory reaction, alleviating ROS accumulation, and attenuating pain. The possible molecular mechanism of it mainly involved multiple components, multiple targets and multiple signaling pathways, which provided a comprehensive understanding for further study. In general, the network pharmacological method applied in this study provides an alternative strategy for the mechanism of ZSD in the treatment of GA.

Multi-Frequency Ultrasonic Extraction of Anthocyanins from Blueberry Pomace and Evaluation of Its Antioxidant Activity.

BACKGROUND: A large amount of blueberry pomace is discarded and wasted. OBJECTIVE: To make full use of blueberry pomace and extract anthocyanins. METHOD: Multi-frequency ultrasound was used to extract anthocyanins from blueberry pomace. The effects of different extraction conditions on the anthocyanin yields were studied. The antioxidant activity of anthocyanins was evaluated, and the main anthocyanin components were identified by HPLC-electrospray ionization (ESI)-MS/MS. RESULTS: The optimal anthocyanin extraction conditions were as follows: dual-frequency ultrasound of 40 + 80 kHz, ultrasonic power of 350 W, extraction temperature of 50°C, and extraction time of 40 min. Compared with single frequency (25 kHz) and tri-frequency ultrasound (25 + 40 + 80 kHz), the anthocyanin yield with dual-frequency ultrasound increased by 15.26% and 5.45% respectively. Furthermore, the antioxidant activities (DPPH, hydroxyl radical scavenging ability, and reducing power) of anthocyanins extracted with dual-frequency ultrasound were all higher than those without ultrasound. Seven kinds of anthocyanins were identified by HPLC-ESI-MS/MS. CONCLUSIONS: Dual-frequency ultrasound had a higher extraction yield than single frequency ultrasound, tri-frequency ultrasound, and no ultrasound. HIGHLIGHTS: Dual-frequency ultrasonic technology provides an effective approach for improving the extraction yield of anthocyanins from blueberry pomace.