[Mechanism of "Trichosanthis Fructus-Allii Macrostemonis Bulbus" in treating cardiovascular diseases with syndrome of combined phlegm and stasis based on serum metabolomics].

This study aimed at investigating the mechanism of Trichosanthis Fructus-Allii Macrostemonis Bulbus(GX) in treating cardiovascular diseases in rats with the syndrome of combined phlegm and stasis. The rat model was established by a high-fat diet, ice-water bath combined with subcutaneous injection of adrenalin hydrochloride, and the syndrome score was determined. The serum samples of rats in the control, model, and GX groups were collected. Ultra-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry(UPLC-Q-TOF-MS) was employed to analyze the metabolic profiles of the serum samples. The differential metabolites were screened and identified by partial least squares-discriminant analysis(PLS-DA) and orthogonal partial least squares-discriminant analysis(OPLS-DA). The intervention targets of GX-regulated metabolites and their metabolic pathways were searched against MetaboAnalyst. Gene Ontology enrichment was carried out to predict the biological pathways associated with the intervention targets of metabolic pathways. A total of 129 potential biomarkers were detected in the rat model with the syndrome of combined phlegm and stasis via metabolomics, and GX regulated 54 metabolites in several metabolic pathways such as linoleic acid metabolism, sphingolipid metabolism, and tricarboxylic acid cycle. The further screening against MetaboAnalyst showed that GX recovered the levels of nine metabolites associated with cardiovascular diseases with the syndrome of combined phlegm and stasis, which involved 69 targets in the pathways regarding cholesterol metabolism, fatty acid metabolism, inflammatory response, and glucose homeostasis and metabolism. The above-mentioned results suggested that GX can alleviate the symptoms of the rat model of cardiovascular diseases with the syndrome of combined phlegm and stasis by regulating the metabolism of linoleic acid, sphingosine, docosahexaenoic acid, rosemary acid, succinic acid, adenine, L-phenylalanine, L-valine and modulating the biological pathways such as cholesterol metabolism, fatty acid metabolism, inflammatory response, and glucose homeostasis and metabolism.

Transcriptomic analysis of Vibrio alginolyticus challenged by Rhizoma coptidis reveals mechanisms of virulence genes.

Rhizoma coptidis, a Chinese herbal medicine widely used to treat various bacterial infections, has the potential to develop antibiotic substitutes to overcome the drug resistance of Vibrio alginolyticus. To study the inhibitory effect of R. coptidis on V. alginolyticus, we sequenced the transcriptomes of three groups of samples of wild-type V. alginolyticus (CK) and V. alginolyticus, which were stressed by 5 mg/mL R. coptidis for 2 h (RC_2 h) and 4 h (RC_4 h). CK was compared with RC_2 h and RC_4 h, respectively, and a total of 1565 differentially expressed genes (DEGs) (988 up-regulated and 577 down-regulated) and 1737 DEGs (1152 up-regulated and 585 down-regulated) were identified. Comparing RC_2 h with RC_4 h, 156 DEGs (114 up-regulated and 42 down-regulated) were identified. The ability of biofilm formation and motility of V. alginolyticus altered upon with different concentrations of R. coptidis. Interestingly, relative expression patterns of virulence genes appeared statistically significantly varied, upon different concentrations of R. coptidis extract. DEGs were annotated to the Gene Ontology (GO) database for function enrichment analysis and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, the results showed that the main enriched pathways, was those related to the virulence of V. alginolyticus. This study provides a new perspective for understanding the complex pathogenic mechanism of V. alginolyticus. R. coptidis could potnetially be used as alternative or complimnetary to antibiotics to treat infections after further research.

Jujuboside B alleviates acetaminophen-induced hepatotoxicity in mice by regulating Nrf2-STING signaling pathway.

BACKGROUND: Jujuboside B (JuB) is the main bioactive saponin component of Chinese anti-insomnia herbal medicine Ziziphi Spinosae Semen, which has been reported to possess varied pharmacological functions. Even though it has been traditionally used to treat inflammation- and toxicity-related diseases, the effects of JuB on acetaminophen (APAP) overdose-induced hepatotoxicity have not been determined yet. METHODS: C57BL/6 J mice were pre-treated with JuB (20 or 40 mg/kg) for seven days before APAP (400 mg/kg) injection. After 24 h of APAP treatment, serum, and liver tissues were collected to evaluate the therapeutic effects. To investigate whether the Nrf2-STING signaling pathway is involved in the protective effects of JuB against APAP-induced hepatotoxicity, the mice received the DMXAA (the specific STING agonist) or ML385 (the specific Nrf2 inhibitor) during the administration of JuB, and Hematoxylin-eosin staining, Real-time PCR, immunohistochemical, and western blot were performed. RESULTS: JuB pretreatment reversed APAP-induced CYP2E1 accumulations and alleviated APAP-induced acute liver injury. Furthermore, JuB treatment significantly inhibited oxidative stress and the pro-inflammatory cytokines, as well as alleviated hepatocyte apoptosis induced by APAP. Besides, our result also demonstrated that JuB treatment upregulated the levels of total Nrf2, facilitated its nuclear translocation, upregulated the expression of HO-1 and NQO-1, and inhibited the APAP-induced STING pathway activation. Finally, we verified that the beneficial effects of JuB were weakened by DMXAA and ML385. CONCLUSION: Our study suggested that JuB could ameliorate APAP-induced hepatic damage and verified a previously unrecognized mechanism by which JuB prevented APAP-induced hepatotoxicity through adjusting the Nrf2-STING pathway.

["Trichosanthis Fructus-Allii Macrostemonis Bulbus" combination inhibits NLRP3 inflammasome activation and reduces inflammatory response in RAW264.7 macrophage by inducing autophagy].

This study aims to explore the effect of "Trichosanthis Fructus-Allii Macrostemonis" combination(GX) on the activation of NOD-, LRR-, and pyrin domain-containing protein 3(NLRP3) inflammasome, the release of inflammatory cytokines, and the level of autophagy in RAW264.7 macrophage damaged by lipopolysaccharide(LPS), and the mechanism of GX against inflammatory response in macrophages. To be specific, LPS was used to induce the injury of RAW264.7 cells. Cell Counting Kit-8(CCK-8) assay was employed to measure the survival rate of cells, and Western blot to detect the protein expression of NLRP3, apoptosis-associated speck-like protein(ASC), cysteine-aspartic acid protease(caspase)-1, interleukin(IL)-18, IL-1ß, microtubule-associated protein light chain 3(LC3)-Ⅱ, and selective autophagy junction protein p62/sequestosome 1 in RAW264.7 macrophages. ELISA was used to measure the levels of IL-18 and IL-1ß in RAW264.7 cells. Transmission electron microscopy was applied to observe the number of autophagosomes in RAW264.7 cells. Immunofulourescence staining was used to detect the expression of LC3-Ⅱ and p62 in RAW264.7 cells. The result showed that GX significantly reduced the protein expression of NLRP3, ASC, and caspase-1 in RAW264.7 cells, significantly increased the protein expression of LC3Ⅱ, decreased the expression of p62, significantly inhibited the secretion of IL-18 and IL-1ß, significantly increased the number of autophagosomes, significantly enhanced the immunofluorescence of LC3Ⅱ, and reduced the immunofluorescence of p62. Furthermore, 3-methyladenine(3-MA) could reverse the inhibitory effect of GX on NLRP3, ASC, and caspase-1 and reduce the release of IL-18 and IL-1ß. In summary, GX can increase of the autophagy activity of RAW264.7 and inhibit the activation of NLRP3 inflammasome, thereby reducing the release of inflammatory cytokines and suppressing inflammatory response in macrophages.

Reabsorption of intervertebral disc prolapse after conservative treatment with traditional Chinese medicine: A case report.

BACKGROUND: Conservative treatments have been reported to diminish or resolve clinical symptoms of lumbar intervertebral disc herniation (LIDH) within a few weeks. CASE SUMMARY: Computed tomography and magnetic resonance imaging (MRI) of the lumbar region of a 25-year-old male diagnosed with LIDH showed prolapse of the L5/S2 disc. The disc extended 1.0 cm beyond the vertebral edge and hung along the posterior vertebral edge. The patient elected a conservative treatment regimen that included traditional Chinese medicine (TCM), acupuncture, and massage. During a follow-up period of more than 12 mo, good improvement in pain was reported without complications. MRI of the lumbar region after 12 mo showed obvious reabsorption of the herniation. CONCLUSION: A conservative treatment regimen of TCM, acupuncture, and massage promoted reabsorption of a prolapsed disc.

Adolescent Hypertension Induced by Obesity and the Efficacy of Comprehensive Intervention.

Context: With the development of the Chinese economy, people's quality of life has improved, obesity caused by excessive nutrition has increased among teenagers, and the age of patients with obesity-induced hypertension has become younger and younger. Objective: The study intended to analyze the factors affecting hypertension in adolescents to find methods to effectively prevent and treat hypertension in that age group. Design: The research team designed a prospective controlled study. Setting: The study took place at the First People's Hospital of Nantong in Nantong, Jiangsu, China from 2020 to 2022 years. Participants: Participants were 1000 students in Grades 7 to 9 at the Si'an middle school in Nantong, China. Intervention: From the 1000 participants, among of them 500 cases of hypertension. The research team (n = 500) participants who were obese and hypertensive and assigned them to the hypertension group, the control group (n = 500) participants with normal weights and no hypertension assigned them to a control group. Participants with obesity-induced hypertension received a comprehensive intervention. Outcome Measures: The research team used a questionnaire and a physical examination to collect data about participants' ages, heights, weights, demographic characteristics, waist circumferences, hip circumferences, and knowledge of hypertension and blood pressure and analyzed the relationships between those factors. The team measured blood pressure, blood lipids, blood glucose, and body mass index at baseline and postintervention. Results: Significant differences existed between the hypertension and control groups at general data: (1) weight-63.49 ± 13.22 kg and 52.59 ± 10.21 kg, respectively (P = .000); (2) waist circumference-75.44 ± 10.92 cm and 68.73 ± 8.15 cm, respectively (P = .001); (3) hip circumference-92.10 ± 7.98 cm and 85.95 ± 7.91 cm, respectively (P = .000); (4) body mass index (BMI)-22.12 ± 4.02 kg/m2 and 19.58 ± 3.34 kg/m2, respectively (P = .002); (5) waist-hip ratio-0.83 ± 0.08 and 0.81 ± 0.07, respectively (P = .003); and (6) waist-to-height ratio-0.46 ± 0.07 and 0.44 ± 0.06, respectively (P = .000). Only age (p=0.006), hip circumference(p=0.000), and BMI (P = .000) were significantly and positively correlated with hypertension. The regression coefficients for age, hip circumference, and BMI were 0.182, 0.062 and 0.096, respectively. The changes in the hypertension group's mean systolic blood pressure (SBP), diastolic blood pressure (DBP), BMI, body fat, trunk fat, abdominal fat, upper-limb fat, and lower-limb fat between baseline and postintervention were statistically significant. Significant decreases in the hypertension group's triglycerides, total cholesterol, and glycated albumin had occurred between baseline and postintervention (all P < .01), and that group's glucagon (P = .011) had significantly increased. No significant changes had occurred in that group's blood glucose, glycated hemoglobin, insulin, and insulin resistance index between baseline and postintervention (P < .05). Conclusions: Obesity increases the risk of hypertension, and comprehensive interventions can effectively prevent and treat adolescent hypertension.

The Effect of Vitamin D Supplementation in Pregnant Women with Overweight and Obesity: A Randomised Controlled Trial.

The impact of vitamin D supplementation on 25-hydroxyvitamin D (25OHD) levels, metabolic status, and pregnancy outcomes in pregnant women with overweight and obesity (OW/OB) is uncertain. This study aimed to examine whether administrating 800 IU of vitamin D3 orally would improve maternal serum 25OHD levels, lipid profile, and pregnancy outcomes compared to 400 IU. This was a two-arm, parallel, non-blinded randomised controlled trial involving 274 pregnant women recruited from KK Women's and Children's Hospital, with a body mass index of ≥25 kg/m2 within 16 weeks gestation. The participants were randomly assigned to receive 800 IU/day (intervention group) or 400 IU/day (control group) of oral vitamin D3 supplements. The primary outcomes were maternal serum 25OHD and lipid levels at 24-28 weeks gestation. The secondary outcomes included maternal and birth outcomes. Compared with controls (n = 119), the intervention group (n = 112) exhibited higher 25OHD levels at 24-28 weeks gestation (adjusted mean difference 6.52 nmol/L; 95% confidence interval 2.74, 10.31). More women in the intervention group achieved sufficient 25OHD levels (77.7% vs. 55.5%; p < 0.001). No differences were observed in lipid profiles or maternal or birth outcomes between the groups. An additional 400 IU of oral vitamin D3 supplementation increased serum 25OHD levels but did not impact lipid profiles or pregnancy outcomes.

The mechanism of "Trichosanthis Fructus-Allii Macrostemonis Bulbus" on phlegm and blood stasis syndrome-related cardiovascular diseases based on network pharmacology and experimental verification / 药学学报

This study aimed to investigate the mechanism of "Trichosanthis Fructus-Allii Macrostemonis Bulbus" (GX) on phlegm and blood stasis syndrome (PBSS) rats combining the methods of network pharmacology and experimental verification. Animal experiment ethical requirements were approved by the Ethical Committee Experimental Animal Center of Anhui University of Chinese Medicine (grant number: AHUCM-rats-2021070). Based on the HPLC-Q-TOF-MS analysis and database, 69 chemical constituents of GX and 163 targets of GX for the treatment of phlegm and blood stasis-related cardiovascular diseases were obtained. Then, key targets such as serine/threonine kinase 1 (Akt1), tumor necrosis factor (TNF), interleukin 6 (IL6), vascular endothelial growth factor A (VEGFA), cellular tumor antigen p53 (Tp53) were screened. Pathway analysis showed that the targets of GX in the treatment of phlegm and blood stasis-relate cardiovascular diseases were mainly involved in PI3K/Akt signaling pathway, sphingolipid metabolism, platelet activation, hypoxia inducible factor-1 (HIF-1), ras-proximate-1 (rap1) and other signaling pathways. In addition, molecular docking analysis showed that apigenin, cucurbitacin D, linolenic acid and kaempferol and other key components had potential binding ability with Akt1, TNF, IL6, VEGFA and Tp53. In the animal experiments, compared to the phlegm and blood stasis syndrome group, GX could significantly improve the traditional Chinese medicine syndrome score, blood lipid, vascular endothelial structure disorders and reduce serum endothelin-1 (ET-1) level, increase serum nitric oxide (NO) and endothelial nitric oxide synthase (eNOS) levels, which could restore aortic endothelial function. In addition, the expression of intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in aorta could be significantly reduced, which could improve the vascular endothelial injury of aorta. Western blot revealed that GX could significantly decrease the phosphorylation levels of phosphoinositide 3-kinase (PI3K) and Akt in aorta. This study revealed the mechanism of GX in treatment of phlegm and blood stasis-relate cardiovascular diseases is consistent with the characteristics of multiple ingredients, multiple targets and multiple pathways. In addition, this study also clarified that the reversal of pathological of phlegm and blood stasis syndrome rats may be related to GX inhibiting PI3K/Akt signaling pathway, which could improve vascular inflammation and vascular endothelial function injury.

Targets and mechanisms of Alpinia oxyphylla Miquel fruits in treating neurodegenerative dementia.

The dried and ripe fruits of Alpinia oxyphylla and ripe fruits of Alpinia oxyphylla Miquel (AO) have the effects of tonifying kidney-essence and nourishing intelligence and thus have been widely used in treating dementia. Alzheimer's disease (AD) is a typical form of neurodegenerative dementia with kidney-essence deficiency in Traditional Chinese Medicine (TCM). So far, there is a lack of systematic studies on the biological basis of tonifying kidney-essence and nourishing intelligence and the corresponding phytochemicals. In this study, we investigated the targets of AO in tonifying kidney-essence and nourishing intelligence based on the key pathophysiological processes of neurodegenerative dementia. According to ultra-high-performance liquid chromatography with triple quadrupole mass spectrometry data and Lipinski's rule of five, 49 bioactive phytochemicals from AO were identified, and 26 of them were found to target 168 key molecules in the treatment of neurodegenerative dementia. Nine phytochemicals of AO were shown to target acetylcholinesterase (ACHE), and 19 phytochemicals were shown to target butyrylcholinesterase (BCHE). A database of neurodegenerative dementia with kidney-essence deficiency involving 731 genes was constructed. Furthermore, yakuchinone B, 5-hydroxy-1,7-bis (4-hydroxy-3-methoxyphenyl) heptan-3-one (5-HYD), oxyhylladiketone, oxyphyllacinol, butyl-ß-D-fructopyranoside, dibutyl phthalate, chrysin, yakuchinone A, rhamnetin, and rhamnocitrin were identified as the key phytochemicals from AO that regulate the pathogenesis of neurodegenerative dementia in a multitargeted manner. The approach of studying the pharmacological mechanism underlying the effects of medicinal plants and the biological basis of TCM syndrome may be helpful in studying the translation of TCM.

[Compatibility mechanism of Trichosanthis Fructus-Allii Macrostemonis Bulbus combination against atherosclerosis: based on metabolomics and network pharmacology].

This study aims to investigate the compatibility mechanism of Trichosanthis Fructus-Allii Macrostemonis Bulbus combination against atherosclerosis(AS) in apolipoprotein E-deficient(ApoE~(-/-)) mice. To be specific, high-fat diet was used to induce AS in mice. The pathological morphology of mice aorta was evaluated based on hematoxylin-eosin(HE) staining and Masson staining. The metabolic profiling of mouse serum samples was performed with ultra-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry. Multiple statistical analysis methods including partial least squares-discriminant analysis and orthogonal partial least squares-discriminant analysis were employed to screen potential biomarkers in mice. With the techniques in network pharmacology, the metabolites related to AS and the targets in the metabolic pathways were screened out. The results showed that Trichosanthis Fructus alone and the pair all reduced the plaque area of aortic sinus(P<0.05) and collagen area(P<0.05). Compared with the Trichosanthis Fructus alone and Allii Macrostemonis Bulbus alone, the combination significantly decreased the plaque area of aortic sinus(P<0.05) and collagen area(P<0.05). Metabolomics revealed 16 biomarkers in mice. Trichosanthis Fructus re-gulated the abnormal levels of 4 metabolites in glycerophosphatide metabolic pathway. Allii Macrostemonis Bulbus modulated the abnormal levels of 2 metabolites in arachidonic acid metabolic pathway and the combination recovered the levels of 8 metabolites in glycerophosphatide, linoleic acid, arachidonic acid, and pyrimidine metabolic pathways. Network pharmacology suggested that Trichosanthis Fructus regulated 24 targets which related to 2 AS-associated metabolites and involved glycerophosphatide metabolic pathway. Allii Macroste-monis Bulbus modulated 40 targets which related to 2 AS-associated metabolites and involved the arachidonic acid metabolic pathway. The combination regulated 57 targets which related to 6 AS-metabolites and involved linoleic acid metabolic pathway, glycerophosphatide metabolic pathway, and arachidonic acid metabolic pathway. These results indicate that the Trichosanthis Fructus-Allii Macrostemonis Bulbus combination enhances the regulation of linoleic acid metabolism, glycerophosphatide metabolism, and arachido-nic acid metabolism, thereby synergistically alleviating lipid disorder and inflammatory response in AS mice.

[Application regularity and solution strategies of mineral medicine Cinnabaris in pediatric preparations].

Mineral medicine is a characteristic element of advantage of traditional Chinese medicine(TCM), which embodies unique scientific connotation. Cinnabaris is a characteristic drug in Chinese medicinal preparations, especially in Chinese medicinal pediatric preparations. Because of the adverse reactions caused by mercury contained, the safety and application of Cinnabaris have attracted much attention. To explore the application regularity and the value of the pediatric preparations containing Cinnabaris, this study statistically analyzed 32 Cinnabaris-contained pediatric preparations in the 2020 edition of the Chinese Pharmacopoeia and 105 pediatric preparations containing Cinnabaris in the Dictionary of Traditional Chinese Medicine Prescriptions(Vol. Ⅰ and Ⅱ). The statistical results indicated that the pediatric preparations and formulae containing Cinnabaris had great advantages in the treatment of pediatric convulsions, but there were still problems in dosage form, dosage, and quality control. In this study, ICP-MS and LC-AFS were further used to determine the content of total mercury and soluble mercury in 15 commercially available pediatric preparations containing Cinnabaris. It was found that the total mercury content was far higher than soluble mercury content in the sample preparations, and there was no obvious correlation between them. According to the results, the research and application strategies of Cinnabaris were put forward in order to provide references for the rational application of Cinnabaris in pediatric preparations.

Licochalcone B induces DNA damage, cell cycle arrest, apoptosis, and enhances TRAIL sensitivity in hepatocellular carcinoma cells.

Hepatocellular carcinoma (HCC) is a highly fatal disease recognized as a growing global health crisis. Traditional Chinese herbal medicines have been used to treat patients with cancer for many years in China. This study investigated the effects of licochalcone B (LCB), a flavonoid compound isolated from the root of Glycyrrhiza uralensis Fisch., on cell proliferation, DNA damage and TNF-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis in HCC cells. Our results showed that LCB inhibited cell proliferation and induced DNA damage, cell cycle arrest and apoptosis. Treatment with LCB significantly inhibited the Akt/mTOR pathway and activated endoplasmic reticulum (ER) stress and mitogen-activated protein kinase (MAPK) signaling pathway. Moreover, combined treatment with LCB and TRAIL yielded evident enhancements in the viability reduction and apoptosis. LCB upregulated death receptor 4 (DR4) and death receptor 5 (DR5) protein in a concentration- and time-dependent manner. The knockdown of DR5 significantly suppressed TRAIL-induced cleavage of PARP, which was enhanced by LCB. Treatment with an extracellular-regulated kinase (ERK) inhibitor (PD98059) or c-Jun N-terminal kinase (JNK) inhibitor (SP600125) markedly reduced the LCB-induced upregulation of DR5 expression and attenuated LCB-mediated TRAIL sensitization. In summary, LCB exhibits cytotoxic activity through modulation of the Akt/mTOR, ER stress and MAPK pathways in HCC cells and effectively enhances TRAIL sensitivity through the upregulation of DR5 expression in ERK- and JNK-dependent manner. Combination therapy with LCB and TRAIL may be an alternative treatment strategy for HCC.

Identification of circular RNA biomarkers for Pien Tze Huang treatment of CCl4­induced liver fibrosis using RNA­sequencing.

Pien Tze Huang (PZH), a common hepatoprotective Traditional Chinese Medicine that has been found to be an effective treatment for carbon tetrachloride­induced hepatic damage, including liver fibrosis. Circular RNAs (circRNAs) serve a crucial role in regulating gene expression levels via circRNA/micro (mi)RNA/mRNA networks in several human diseases and biological processes. However, whether circRNAs are involved in the underlying mechanism of the therapeutic effects of PZH on liver fibrosis remains unclear. Therefore, the aim of the present study was to investigate these effects using circRNA expression profiles from PZH­treated fibrotic livers in model mice. A case­control study on >59,476 circRNAs from CCl4­induced (control group, n=6) and PZH­treated (case group, n=6) mice was performed using circRNA sequencing in liver tissues. PZH treatment resulted in the differential expression of 91 circRNAs, including 58 upregulated and 33 downregulated circRNAs. Furthermore, the construction of competing endogenous networks also indicated that differentially expressed circRNAs acted as miRNA sponges. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis of miRNA targets demonstrated that PZH­affected circRNAs were mainly involved in biological processes such as 'positive regulation of fibroblast proliferation', 'cellular response to interleukin­1' and 'regulation of DNA­templated transcription in response to stress' and in a number of important pathways, such as 'TNF signaling pathway', 'PI3K­Akt signaling pathway', 'IL­17 signaling pathway' and 'MAPK signaling pathway'. To further validate the bioinformatics data, reverse transcription­-quantitative PCR was performed on seven miRNA targets in a human hepatic stellate LX­2 cell model. The results suggested that seven of the miRNAs exhibited regulatory patterns that were consistent with those of the transcriptome sequencing results. Kaplan­Meier survival analysis demonstrated that the expression levels of dihydrodiol dehydrogenase and solute carrier family 7, member 11 gene were significantly associated with patient survival, 269 patients with liver hepatocellular carcinoma from The Cancer Genome Atlas database. To the best of our knowledge, this was the first study to provide evidence that PZH affects circRNA expression levels, which may serve important roles in PZH­treated fibrotic liver through the regulation of functional gene expression. In conclusion, the present study provided new insights into the mechanism underlying the pathogenesis of liver fibrosis and identified potential novel, efficient, therapeutic targets against liver injury.

[Effects of tHMGR from three different plant sources on mevalonate (MVA) pathway flux in Saccharomyces cerevisiae].

In this study, we used bioinformatic tools to analyze the 3-hydroxy-3-methylglutaryl-CoA reductase(HMGR) genes from Glycyrrhiza uralensis, Artemisia annua, and Arabidopsis thaliana. The results indicated that GuHMGR and AaHMGR contained two transmembrane regions while AtHMGR had three transmembrane regions. GuHMGR, AaHMGR, and AtHMGR all had the active center for catalysis. Three truncated HMGR genes(tHMGRs) of G. uralensi, A. annua, and A. thaliana were respectively ligated to pYES3 vector to construct the recombinant plasmids pYES3-tGuHMGR,pYES3-tAaHMGR,and pYES3-tAtHMGR. Afterwards, the control plasmid pYES3 and the three plasmids and were respectively introduced into Saccharomyces cerevisiae Cen.pk2-1 D, which yielded strains Y0, Y1, Y2, and Y3, respectively. The content of squalene, lanosterol, and ergosterol in these strains was measured by GC-MS. The relative expression of tGuHMGR, tAaHMGR, and tAtHMGR in strains Y1, Y2, and Y3 was determined by quantitative real-time PCR. The results showed that the strain overexpressing tAaHMGR had the highest yield of squalene and the highest total yield of squalene, ergosterol, and lanosterol. The quantitative real-time PCR showed higher relative expression of tAaHMGR than tGuHMGR, consistent with the strain fermentation result. We selected a superior tHMGR by comparing the effects of different tHMGRs on the mevalonate(MVA) pathway flux in S. cerevisiae. The findings can provide a reference for the construction of S. cerevisiae strains with high yields of squalene and terpenoid precursors.

Higher efficacy of resveratrol against advanced breast cancer organoids: A comparison with that of clinically relevant drugs.

The lack of reliable drugs is a therapeutic challenge of advanced breast cancers (ABCs). Resveratrol (Res) exerts inhibitory effects on breast cancer cell lines and animal models, while its efficacy against individual breast cancer cases remains unknown. This study aims to use ABC-derived organoids (ABCOs) as the ex vivo therapeutic platform to clarify the effectiveness of resveratrol against different ABC subtypes. Immunohistochemical staining confirmed that the ABCOs maintained their original tumors' ER, PR, HER2, and Ki67 expression patterns. ABCO proliferation and viability tests showed >50% cell death rates in 79.2% (19/24) of Res-treated, 28.6% (2/7) fulvestrant-treated, 66.7% (4/6) paclitaxel-treated, and 66.7% (6/9) gemcitabine-treated ABCOs. pSTAT3 nuclear translocation was more frequent in Res-sensitive (17/19; 89.47%) than that (1/5; 20%) of Res-insensitive ABCOs, which were suppressed upon Res treatment. Statistical analysis revealed a close correlation of STAT3 activation with the efficacy of Res, but not related to tumor receptor expression patterns (ER, PR, HER2) and pathological classification. We demonstrate for the first time the higher efficacy and broader spectrum of Res against different subtypes of ABCOs in comparison with that of conventional antibreast cancer drugs, providing an alternative approach for better management of ABCs.

[Effect of Pien Tze Huang against EV71].

This study aims to investigate the inhibitory effect of Pien Tze Huang(PZH) on enterovirus 71(EV71). To be speci-fic, chemiluminescence method was adopted to evaluate the toxicity of PZH to African green monkey kidney(Vero) cells and human rhabdomyosarcoma(RD) cells, and cytopathic effect(CPE) method to assess the inhibition on EV71-GFP reporter virus and EV71 C4 wild-type virus. The results showed that PZH had low cytotoxicity to Vero cells and RD cells, with the half-maximal cytotoxic concentration(CC_(50)) of about 0.691 3-0.879 2 mg·mL~(-1) for the two. In addition, PZH can effectively inhibit the replication of EV71 within the non-cytotoxic concentration range, and dose-dependently alleviate the cytopathic changes caused by virus infection, with the half-maximal effective concentration(EC_(50)) of 0.009 2-0.106 3 mg·mL~(-1). On the basis of the above results, the green fluorescent protein(GFP), indirect immunofluorescence assay(IFA), and median tissue culture infective dose(TCID_(50)) were employed to assess and verify the anti-EV71-GFP and anti-EV71 C4 activity of PZH. The results demonstrated that PZH can dose-dependently lower the expression of GFP by EV71-GFP and structural protein VP-1 by EV71 C4 and decrease the production of progeny infectious viruses. The EC_(50) of PZH for EV71-GFP and EV71 C4 was about 0.006 0-0.006 2 mg·mL~(-1) and 0.006 6-0.025 6 mg·mL~(-1), respectively. This study suggested that PZH may exert antiviral activity by acting on EV71 and interfering with the expression of VP-1. At the moment, there is still a lack of specific anti-EV71 drugs. This study proposed a new idea for the symptomatic treatment of EV71 infections such as hand-foot-mouth disease and verified an effective drug for the treatment of EV71 infections.

[Proposal of Living Evidence-based Guideline for Combination of Traditional Chinese and Western Medicine for Treatment of COVID-19].

In order to standardize the clinical diagnosis and treatment decision-making with traditional Chinese medicine for pa-tients of coronavirus disease 2019(COVID-19) and put the latest clinical study evidence into clinical practice, the international trust-worthy traditional Chinese medicine recommendations( TCM Recs) working group started the compilation of Living Evidence-based Guideline for Combination of Traditional Chinese and Western Medicine for Treatment of COVID-19 on the basis of the standards and re-quirements of WHO handbook, GRADE and RIGHT. This proposal mainly introduces the formulation methods and processes of the living guidelines in details, such as the composition of the working group, the collection and identification of clinical issues and out-comes, the production of the living systematic review and the consensus of recommendations. The guidelines will continue to monitor the clinical study evidences of TCM in the prevention and treatment of COVID-19, and conduct regular evidence updating, retrieval and screening. When there is new study evidence, the steering committee will evaluate the possibility of the evidence to change clinical practice or previous recommendations, so as to decide whether the recommendations for the guidelines shall be implemented or upda-ted. The main criteria considered in the guideline updating are as follows:(1) There are new high-quality randomized controlled trial(RCT) evidences for TCM uninvolved in the previous edition of the guidelines;(2) as for the TCM involved in the guidelines, living sys-tematic review shows that new evidence may change the direction or strength of the existing recommendations. The specific implementation of the living evidence-based guidelines will take this proposal as the study basis and framework, in order to ensure the standardization of the formulation process and methods. This will be the first exploration of the methodology for living guidelines in the field of TCM.

[Study on effect of "Trichosanthis Fructus-Allii Macrostemonis Bulbus" on atherosclerosis in ApoE~(-/-) mice based on liver metabonomics].

In this study, ultra-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry(UPLC-Q-TOF-MS)-based liver metabolomics approach was used to explore the mechanism of "Trichosanthis Fructus-Allii Macrostemonis Bulbus" in improving atherosclerosis(AS) of mice with apolipoprotein E gene knockout(ApoE~(-/-)). AS mouse model was induced by high-fat diet. The pathological and biochemical indexes such as the histopathological changes, body weight, liver weight, blood lipid level and inflammatory factors in the liver of mice were determined. The metabolic profiling of mice liver samples was performed with UPLC-Q-TOF-MS. Multiple statistical analysis methods including partial least squares discriminant analysis(PLS-DA) and orthogonal partial least squares discriminant analysis(OPLS-DA) were employed to screen and identify biomarkers. The levels of related enzymes including LCAT, sPLA2, EPT1 and ACER1 were detected. The results showed that "Trichosanthis Fructus-Allii Macrostemonis Bulbus" significantly reduced the areas of aortic plaque and fat vacuoles of liver in AS mice and decreased the accumulation of lipid droplets and liver coefficient. "Trichosanthis Fructus-Allii Macrostemonis Bulbus" also regulated the levels of blood lipid and inflammatory injury in the liver. The metabolites of the control group, the model group and the "Trichosanthis Fructus-Allii Macrostemonis Bulbus" group could be distinguished significantly. Fifteen potential biomarkers related to AS were discovered and preliminarily identified, seven of which could be regulated by "Trichosanthis Fructus-Allii Macrostemonis Bulbus" in a trend of returning to normal. Metabolic pathway analysis screened out two major metabolic pathways. "Trichosanthis Fructus-Allii Macrostemonis Bulbus" obviously regulated the levels of LCAT, sPLA2, EPT1 and ACER1. It was inferred that "Trichosanthis Fructus-Allii Macrostemonis Bulbus" could play a major role in AS treatment by regulating glycerophospholipid and sphingolipid metabolism disorders in the liver, with the mechanism probably relating to the intervention of the expression of LCAT, sPLA2, EPT1 and ACER1.

Immune Mechanism, Gene Module, and Molecular Subtype Identification of Astragalus Membranaceus in the Treatment of Dilated Cardiomyopathy: An Integrated Bioinformatics Study.

Astragalus membranaceus has complex components as a natural drug and has multilevel, multitarget, and multichannel effects on dilated cardiomyopathy (DCM). However, the immune mechanism, gene module, and molecular subtype of astragalus membranaceus in the treatment of DCM are still not revealed. Microarray information of GSE84796 was downloaded from the GEO database, including RNA sequencing data of seven normal cardiac tissues and ten DCM cardiac tissues. A total of 4029 DCM differentially expressed genes were obtained, including 1855 upregulated genes and 2174 downregulated genes. GO/KEGG/GSEA analysis suggested that the activation of T cells and B cells was the primary cause of DCM. WGCNA was used to obtain blue module genes. The blue module genes are primarily ADCY7, BANK1, CD1E, CD19, CD38, CD300LF, CLEC4E, FLT3, GPR18, HCAR3, IRF4, LAMP3, MRC1, SYK, and TLR8, which successfully divided DCM into three molecular subtypes. Based on the CIBERSORT algorithm, the immune infiltration profile of DCM was analyzed. Many immune cell subtypes, including the abovementioned immune cells, showed different levels of increased infiltration in the myocardial tissue of DCM. However, this infiltration pattern was not obviously correlated with clinical characteristics, such as age, EF, and sex. Based on network pharmacology and ClueGO, 20 active components of Astragalus membranaceus and 40 components of DMCTGS were obtained from TCMSP. Through analysis of the immune regulatory network, we found that Astragalus membranaceus effectively regulates the activation of immune cells, such as B cells and T cells, cytokine secretion, and other processes and can intervene in DCM at multiple components, targets, and levels. The above mechanisms were verified by molecular docking results, which confirmed that AKT1, VEGFA, MMP9, and RELA are promising potential targets of DCM.

Target identification of hepatic fibrosis using Pien Tze Huang based on mRNA and lncRNA.

Hepatic fibrosis is a spontaneous wound-healing response triggered by chronic liver injury. Pien Tze Huang (PZH), a traditional Chinese herbal medicine, has been widely used to treat various hepatic diseases in Asia. We used a CCl4-induced mouse model to establish a PZH group of hepatic fibrosis mice treated with PZH and a control group of hepatic fibrosis mice without any treatment. We performed RNA-seq and mass spectrometry sequencing to investigate the mechanism of the PZH response in hepatic fibrosis and identified multiple differentially expressed transcripts (DETs) and proteins (DEPs) that may be drug targets of PZH. Liver functional indices, including serum albumin (ALB), alanine aminotransferase (ALT) and aspartate aminotransferase (AST), were significantly decreased in the PZH treatment group (P < 0.05) in the eighth week. Hematoxylin-eosin (HE), Masson and Sirius red staining demonstrated that PZH significantly inhibited infiltration of inflammatory cells and collagen deposition. A total of 928 transcripts and 138 proteins were differentially expressed in PZH-treated mice compared to the control group. Gene Ontology (GO) enrichment analysis suggested that PZH may alleviate liver injury and fibrosis by enhancing the immune process. Taken together, our results revealed that multiple DETs and DEPs may serve as drug targets of PZH in hepatic fibrosis patient in future clinical practice.