Integrating UPLC-Q-Exactive Orbitrap/MS, network pharmacology and experimental validation to reveal the potential mechanism of Tibetan medicine Rhodiola granules in improving myocardial ischemia-reperfusion injury.

ETHNOPHARMACOLOGY RELEVANCE: Rhodiola granules (RG) is a traditional Tibetan medicine prescription that can be used to improve the symptoms of ischemia and hypoxia in cardiovascular and cerebrovascular diseases. However, there is no report on its use to improve myocardial ischemia/reperfusion (I/R) injury, and its potential active ingredients and mechanism against myocardial ischemia/reperfusion (I/R) injury remain unclear. AIM OF THE STUDY: This study aimed to reveal the potential bioactive components and underlying pharmacological mechanisms of RG in improving myocardial I/R injury through a comprehensive strategy. MATERIALS AND METHODS: UPLC-Q-Exactive Orbitrap/MS technology was used to analyze the chemical components of RG, the potential bioactive components and targets were tracked and predicted by the SwissADME and SwissTargetPrediction databases, and the core targets were predicted through the PPI network, as well the functions and pathways were determined by GO and KEGG analysis. In addition, the molecular docking and ligation of the anterior descending coronary artery-induced rat I/R models were experimentally validated. RESULTS: A total of 37 ingredients were detected from RG, including nine flavones, ten flavonoid glycosides, one glycoside, eight organic acids, four amides, two nucleosides, one amino acid, and two other components. Among them, 15 chemical components, such as salidroside, morin, diosmetin, and gallic acid were identified as key active compounds. Ten core targets, including AKT1, VEGF, PTGS2, and STAT3, were discovered through the analysis of the PPI network constructed from 124 common potential targets. These possible targets were involved in the regulation of oxidative stress and HIF-1/VEGF/PI3K-Akt signaling pathways. Furthermore, molecular docking confirmed that the potential bioactive compounds in RG have good potential binding abilities to AKT1, VEGFA, PTGS2, STAT3, and HIF-1α proteins. Then, the animal experiments showed that RG could significantly improve the cardiac function of I/R rats, reduce the size of myocardial infarction, improve the myocardial structure, and reduce the degree of myocardial fibrosis, inflammatory cell infiltration, and myocardial cell apoptosis rate in I/R rats. In addition, we also found that RG could decrease the concentration of AGE, Ox-LDL, MDA, MPO, XOD, SDH, Ca2+, and ROS, and increase the concentration of Trx, TrxR1, SOD, T-AOC, NO, ATP, Na+k+-ATPase, Ca2+-ATPase, and CCO. Moreover, RG could significantly down-regulate the expressions of Bax, Cleaved-caspase3, HIF-1α, and PTGS2, as well up-regulate the expressions of Bcl-2, VEGFA, p-AKT1, and p-STAT3. CONCLUSION: In summary, we revealed for the first time the potential active ingredients and mechanisms of RG for myocardial I/R injury therapy through a comprehensive research strategy. RG may synergistically improve myocardial I/R injury through anti-inflammatory, regulating energy metabolism, and oxidative stress, improving I/R-induced myocardial apoptosis, which may be related to the HIF-1/VEGF/PI3K-Akt signaling pathway. Our study provides new insights into the clinical application of RG and also provides a reference for the development and mechanism research of other Tibetan medicine compound preparations.

Preclinical anti-apoptotic properties of salidroside for hypoxic-ischemic cerebral damage: a systematic review and meta-analysis / 数字中医药（英文）

@#【Objective】   As the main active ingredient of Tibetan medicine Hongjingtian (Rhodiolae Crenulatae Radix et Rhizoma), salidroside (Sal) has a good anti-apoptotic potential. Currently, there are some conflicting results on the anti-apoptotic mechanisms of Sal. Here we conducted a systematic review and meta-analysis to provide the preclinical evidence of its anti-apoptotic properties in preventing and treating hypoxic-ischemic cerebral damage(HICD). 【Methods】   The literature on the anti-apoptotic potential of Sal in the treatment of HICD from January 1, 1980 to November 9, 2021 was searched online using Chinese databases including Chinese National Knowledge Infrastructure (CNKI), China Science and Technology Journal Database (VIP), and Wanfang Database, and English databases including PubMed and Web of Science. The quality of the included articles was evaluated by the Cochrane Collaboration network bias risk assessment criteria, and meta-analysis was performed using RevMan 5.3 software. 【Results】  A total of 40 articles were finally included. Among the 40 articles, 30 were about in vivo animal experiments and 17 about in vitro cell experiments, and 7 of them included both animal and cell experiments. After analysis, it was found that Sal had significant effects on disease-related indicators of HICD (P < 0.05), such as cerebral infarctsize and brain water content. As to in vivo studies, Sal mainly affects the expressions of apoptotic factors through antiinflammation, anti-oxidation, activation of complement pathway, and regulation of signal transduction and autophagy, thus exerting anti-apoptotic potential in treating HICD. While for in vitro studies, Sal plays the anti-apoptotic role in HICD models mainly through anti-oxidation, anti-inflammation, reduction of Ca2+ overload, regulation of mitochondrial function, signal transduction, and C3 complement. 【Conclusion】  Sal can take anti-apoptotic effects to prevent and treat HICD through mechanisms such as anti-inflammation, anti-oxidation, enhanced autophagy, complement and signal transduction, regulation of mitochondrial membrane potential, and reduction of Ca2 + overload.

Targeting P2 receptors in purinergic signaling: a new strategy of active ingredients in traditional Chinese herbals for diseases treatment.

Adenosine triphosphate (ATP) and its metabolites adenosine diphosphate, adenosine monophosphate, and adenosine in purinergic signaling pathway play important roles in many diseases. Activation of P2 receptors (P2R) channels and subsequent membrane depolarization can induce accumulation of extracellular ATP, and furtherly cause kinds of diseases, such as pain- and immune-related diseases, cardiac dysfunction, and tumorigenesis. Active ingredients of traditional Chinese herbals which exhibit superior pharmacological activities on diversified P2R channels have been considered as an alternative strategy of disease treatment. Experimental evidence of potential ingredients in Chinese herbs targeting P2R and their pharmacological activities were outlined in the study.

Salvia miltiorrhiza improves type 2 diabetes: A protocol for systematic review and meta-analysis.

BACKGROUND: Diabetes refers to any group of metabolic diseases characterized by high blood sugar and generally thought to be caused by insufficient production of insulin, impaired response to insulin. Globally, patients with type 2 diabetes account for more than 85% of the total diabetic patients, and due to factors, such as obesity, aging, environment and lifestyle, the incidence of diabetes is rising. Salvia miltiorrhiza (SM) is a medicine used to treat diabetes in China. In recent years, it has been reported that SM has the effect of improving type 2 diabetes. However, there is no systematic review of its efficacy and safety yet. Therefore, we propose a systematic review to evaluate the efficacy and safety of SM for T2D. METHODS: Six databases will be searched: China National Knowledge Infrastructure (CNKI), China Biological Medicine (CBM), China Scientific Journals Database (CSJD), Wanfang database, PubMed, and EMBASE. The information is searched from January 2010 to July 2020. Languages are limited to English and Chinese. The primary outcomes include 2 hour plasma glucose, fasting plasma glucose, hemoglobin A1c, homeostasis model assessment of insulin resistance, and fasting plasma insulin. The secondary outcomes include clinical efficacy and adverse events. RESULTS: This systematic review will evaluate the efficacy and safety of Salvia miltiorrhiza in the treatment of type 2 diabetes. CONCLUSION: This systematic review provides evidence as to whether Salvia miltiorrhiza is effective and safe for type 2 diabetes. ETHICS: Ethical approval is not necessary as this protocol is only for systematic review and does not involve in privacy data or an animal experiment. SYSTEMATIC REVIEW REGISTRATION: INPLASY2020110046.

Is gastrointestinal motility related to alkaloids of Charred Semen Arecae?

ETHNOPHARMACOLOGICAL RELEVANCE: Semen Arecae (SA) is one of the most commonly used Traditional Chinese Medicine. Charred Semen Arecae (CSA) is the processed product of SA. Alkaloids are considered as pharmacological mechanisms of SA and CSA on gastrointestinal motility. Recent studies have shown alkaloids decreased quickly after procession. However, the promoting on gastrointestinal motility were not decreased. Is gastrointestinal motility related to alkaloids of CSA? This study explored the effects of SA, CSA, Semen Arecae-Removal (SA-R), and Charred Semen Arecae-Removal (CSA-R) on gastrointestinal motility, Gastric Inhibitory Polypeptide (GIP), Glucagon Like Peptide-1 (GLP-1), gastric juice and bile in rats. MATERIAL AND METHODS: Rats were randomly divided into six groups, including the Control group, SA group, CSA group, SA-R group, CSA-R group, and Positive drug group (Mosapride). Alkaloids of samples were knocked out by using the "target constituent removal" strategy. Gastric residue and intestinal propulsion rate were evaluated in rats. Serum levels of GIP and GLP-1 were measured by Enzyme-Linked Immunosorbent Assay (ELISA). Gastric juice and bile were examined, respectively. RESULTS: CSA-R and SA-R have been investigated by the Preparative Thin-layer Chromatography (PTLC) method. Intestinal propulsion and gastric residue assessments confirmed the effectiveness of CSA and CSA-R. CSA-R was higher than SA-R in the GLP-1, pepsin activity, the secretion of bile, Bilirubin (BIL), and Cholesterol (CHO). The statistical comparison demonstrated that there is no difference between the CSA group and CSA-R group. CONCLUSIONS: After processing, the promoting gastrointestinal motility might be not related to alkaloids. Maillard reaction could be produced to promote the secretion of GLP-1, bile, and CHO for gastrointestinal motility. Our findings provide a pharmacological reference for the clinical application of SA and CSA in the treatment of digestive diseases.

The efficacy and safety of acupuncture for perimenopause symptom compared with different sham acupuncture control groups: A protocol of systematic review and meta-analysis.

BACKGROUND: Perimenopause is a period that every woman must go through, most people are more or less affected by perimenopausal symptoms, it to affect women's health, work, life, and economy. As acupuncture treatment is more and more increasing in perimenopausal symptoms, there have also been many clinical trials about it. But the results of the trials are inconsistent. Therefore, we will conduct a systematic review and meta-analysis of the safety and efficacy of perimenopausal symptoms treated with acupuncture. METHODS: The protocol followed Preferred Reporting Items for Systematic Reviews and Meta-Analyses Protocols. RCT study on different acupuncture interventions for perimenopausal symptoms will be searched in 8 databases (PubMed, EMBASE, the Cochrane Library, the web of science, CBM, CNKI, WAN FANG, and VIP). Besides, the search will also be performed on the clinical trial research platform if necessary. The primary outcome that will be extracted: the Flushes per 24 hours, the Frequency of hot flashes, the severity of hot flashes, the menopause-related symptom score, the treatment efficacy, the adverse event. Endnote software X8 will be used for study selection, STATA 13.0 and Review Manager software 5.3 will be used for analysis and synthesis. These studies selection, data extraction, and risk of bias assessment will be conducted by 2 independent reviewers. RESULTS: This study will provide the results: 1. the primary and secondary outcome indicators of different acupuncture intervention measures (traditional hand acupuncture, moxibustion, ear acupuncture, laser, acupressure points) for perimenopausal symptoms. 2. The effects of different control groups (medicine control, routine care, waiting, and sham acupuncture control) on the analysis results will be reported, especially the effects of different sham acupuncture control (invasive/noninvasive) on the analysis results. CONCLUSION: This systematic review and meta-analysis study hopes to provide useful evidence for better use of different types of acupuncture in treat perimenopausal symptoms and better design of control groups in related clinical trials. In addition, the research conclusion will be published in peer journals.OSF REGISTRATION NUMBER DOI 10.17605/OSF.IO/VZCKU Ethics and dissemination This conclusion of the study will be published in peer journals. The ethical approval is not required because there is no direct involvement of human.

Patchouli alcohol isolated from Pogostemon cablin mediates endothelium-independent vasorelaxation by blockade of Ca2+ channels in rat isolated thoracic aorta.

ETHNOPHARMACOLOGICAL RELEVANCE: The aerial parts of Pogostemon cablin (Blanco) Benth. for the treatment of cardiodynia have been documented in Mingyi Bielu of late Han Dynasty, in addition to that the Ca2+ antagonized activities of P. cablin and its critically pharmacological ingredient patchouli alcohol (PA) were reported previously. AIM OF THE STUDY: To investigate the relaxant effects of PA on rat isolated thoracic aortas and further explore its potential mechanisms of actions. MATERIALS AND METHODS: The aortas with endothelium and without endothelium were prepared and suspended in the organ bath for isometric tension recordings. The responses to accumulative concentrations of PA in endothelium-intact (E + ) aortas with basal tension and in different treated aortas pre-contracted with potassium chloride (KCl) or phenylephrine (PHE) were observed; the effects of L-NAME and indomethacin in aortas with intact endothelium, and of L-NAME, propranolol, tetraethtylamine (TEA), 4-aminopyridine (4-AP), barium chloride (BaCl2), glyburide in aortas with endothelial denudation on PA-produced vasorelaxation were assessed; the influences of PA on extracellular Ca2+ influx and intracellular Ca2+ release were measured in Ca2+-free medium. Finally, the abilities of PA to inhibit KCl- and PHE-induced contractions were compared to that of verapamil in E- aortas. RESULTS: PA produced vasorelaxant effects in KCl- and PHE-precontracted E + aortas in a concentration-dependent manner, which had no statistically different from that in KCl- and PHE-precontracted E- aortas. PA (10 µM) significantly reduced KCl-induced contractions while PHE-induced contractions were significantly reduced by 100 µM of PA instead of 10 µM and 30 µM in aortas with endothelium. Pre-incubation of E + aortas with L-NAME as well as indomethacin and of E- aortas with L-NAME, propranolol, TEA, 4-AP, BaCl2 and glyburide had no obvious effects on vasorelaxation of PA. In endothelium-removed aortas around Ca2+-free solution, PA remarkably lowered the contractions stimulated with Ca2+ and PHE, and application of ruthenium red and heparin further enhanced the abilities of PA to reduce PHE-caused contractions. In aortas without endothelium, 100 µM of PA markedly attenuated KCl-induced contractions but not affect PHE-induced contractions. Verapamil at the equal dose markedly attenuated KCl- and PHE-induced contractions, and the inhibitory effects on KCl-induced contractions were more forceful than that on PHE-induced contractions. In combined usage, the inhibitory effects on the contractions elicited by KCl were evidently weaker than that of verapamil alone and indifferently stronger than that of PA alone, and the inhibitory effects on the contractions elicited by PHE were evidently weaker than that of single verapamil but stronger than that of single PA. CONCLUSION: PA may act as a Ca2+ antagonist to exert an intensively vasorelaxant effects through endothelium-independent pathway, and its mechanisms underlying the vasoactivities seem to be associated with the blockade of extracellular Ca2+ influx through VDCCs and ROCCs in vascular smooth muscle cells (VSMCs) membrane and intracellular Ca2+ releases through IP3R- and RYR-mediated Ca2+ channels in sarcolemma.

Danggui Buxue decoction promotes angiogenesis by up-regulation of VEGFR1/2 expressions and down-regulation of sVEGFR1/2 expression in myocardial infarction rat.

BACKGROUND: The traditional herbal compound Danggui Buxue decoction (DBD), has long been used for the prevention and treatment of cardiovascular diseases, however, the underlying molecular mechanism for its effect remains still unknown. So this study would to investigate the effect of DBD on cardiac damage induced by myocardial infarction (MI) challenge. METHODS: SD Rats with ligation of left anterior descending (LAD) coronary artery were randomly divided into MI, MI plus Betaloc Zok, MI plus DBD high dose, and MI plus DBD low dose group, together with sham-operated group. After corresponding treatment for consecutive 4 weeks, cardiac function was evaluated by hemodynamics with the method of pressure-volume conduit system. Cardiac histological morphology, microvascular density and the expressions of VEGF and VEGFR1/2 mRNA and their relative protein including VEGF, membranous VEGFR1 (VEGFR1), soluble VEGFR1 (sVEGFR1), VEGFR2, and sVEGFR2 were examined by hematoxylin & eosin staining, immunohistochemical staining and quantitative polymerase chain reaction and western blot assay, respectively. RESULTS: It showed that a significant impaired cardiac function and a remarkably inducible increase in fibrotic scar formation, microvascular density and VEGF mRNA expressions in MI rats. While DBD treatment could markedly boost cardiac angiogenesis further, hinder fibrotic scar formation, and improve declined cardiac function. Apart from the up-regulation of VEGF mRNA and VEGF and the down-regulation of sVEGFR1/2, high dose of DBD dedicated to increasing VEGFR1 mRNA and VEGFR1 expression, while low dose to elevating VEGFR2 mRNA and VEGFR2 expression. CONCLUSION: The present study demonstrated that DBD could accelerate cardiac angiogenesis, restrain fibrous scar formation and thus ameliorate cardiac function in post-MI, via the active regulation of VEGF/VEGFRs signaling pathway.

Availability, Pharmaceutics, Security, Pharmacokinetics, and Pharmacological Activities of Patchouli Alcohol.

Patchouli alcohol (PA), a tricyclic sesquiterpene, is one of the critical bioactive ingredients and is mainly isolated from aerial part of Pogostemon cablin (known as guanghuoxiang in China) belonging to Labiatae. So far, PA has been widely applied in perfume industries. This review was written with the use of reliable information published between 1974 and 2016 from libraries and electronic researches including NCKI, PubMed, Reaxys, ACS, ScienceDirect, Springer, and Wiley-Blackwell, aiming at presenting comprehensive outline of security, pharmacokinetics, and bioactivities of PA and at further providing a potential guide in exploring the PA and its use in various medical fields. We found that PA maybe was a low toxic drug that was acquired numerously through vegetable oil isolation and chemical synthesis and its stability and low water dissolution were improved in pharmaceutics. It also possessed specific pharmacokinetic characteristics, such as two-compartment open model, first-order kinetic elimination, and certain biometabolism and biotransformation process, and was shown to have multiple biological activities, that is, immunomodulatory, anti-inflammatory, antioxidative, antitumor, antimicrobial, insecticidal, antiatherogenic, antiemetic, whitening, and sedative activity. However, the systematic evaluations of preparation, pharmaceutics, toxicology, pharmacokinetics, and bioactivities underlying molecular mechanisms of action also required further investigation prior to practices of PA in clinic.

Comparative Researches of Semen Arecae and Charred Semen Arecae on Gastrointestinal Motility, Motilin, Substance P, and CCK in Chronically Stressed Rats.

AIMS: To compare the effects of Semen Arecae (SA) and Charred Semen Arecae (CSA) on gastrointestinal motility, motilin, substance P (SP), and cholecystokinin (CCK) in chronically stressed rats. METHODS: Rats were randomly divided into control group and stress group. Rats in stress group were randomly exposed to a variety of unpredictable stimulations for 21 days. Then, the rats were treated orally with distilled water, SA, CSA, and mosapride for 7 days. Gastric residue rate and intestinal propulsion rate were evaluated. Serum levels of motilin and SP were measured by enzyme-linked immunosorbent assay (ELISA). CCK mRNA was quantified by using quantitative real-time PCR (qRT-PCR). RESULTS: Both SA and CSA improved the intestinal propulsion and reduced the gastric residue in chronically stressed rats. Furthermore, the serum levels of motilin and SP were significantly higher and the CCK mRNA expressions in intestine and hypothalamus were downregulated in SA and CSA groups. Furthermore, it was found that CSA is more effective. CONCLUSION: Both SA and CSA enhanced gastrointestinal motility and increased serum levels of motilin and SP in chronically stressed rats via downregulating CCK mRNA expressions in intestine and hypothalamus. Importantly, CSA possessed more effective promoting effects.

Research on choleretic effect of menthol, menthone, pluegone, isomenthone, and limonene in DanShu capsule.

Danshu capsule (DSC) is a medicinal compound in traditional Chinese medicine (TCM). It is commonly used for the treatment of acute & chronic cholecystitis as well as choleithiasis. To study its choleretic effect, healthy rats were randomly divided into DSC high (DSCH, 900mg/kg), medium (DSCM, 450mg/kg), and low (DSCL, 225mg/kg) group, Xiaoyan Lidan tablet (XYLDT, 750mg/kg), and saline group. The bile was collected for 1h after 20-minute stabilization as the base level, and at 1h, 2h, 3h, and 4h after drug administration, respectively. Bile volume, total cholesterol, and total bile acid were measured at each time point. The results revealed that DSC significantly stimulated bile secretion, decreased total cholesterol level and increased total bile acid level. Therefore, it had choleretic effects. To identify the active components contributing to its choleretic effects, five major constituents which are menthol (39.33mg/kg), menthone (18.02mg/kg), isomenthone (8.18mg/kg), pluegone (3.31mg/kg), and limonene (4.39mg/kg) were tested on our rat model. The results showed that menthol and limonene could promote bile secretion when compared to DSC treatment (p > 0.05); Menthol, menthol and limonene could significantly decrease total cholesterol level (p<0.05 or p<0.01) as well as increase total bile acid level (p<0.05 or p<0.01); Isomenthone, as a isomer of menthone, existed slightly choleretic effects; Pluegone had no obvious role in bile acid efflux. These findings indicated that the choleretic effects of DSC may be attributed mainly to its three major constituents: menthol, menthone and limonene.

The effect of puerarin on serum nitric oxide concentration and myocardial eNOS expression in rats with myocardial infarction.

Puerarin (1) is a major effective ingredient extracted from the traditional Chinese medicine Ge-gen (Radix Puerariae, RP). Recently, puerarin has been used to treat patients with coronary artery diseases (CAD). However, the mechanisms of puerarin on CAD are still not very clear. In this study, we investigated the role of puerarin on serum nitric oxide (NO) concentration, myocardial endothelial nitric oxide synthase (eNOS) gene expression, the protein expression of eNOS and inducible nitric oxide synthase (iNOS), as well as the level of protein kinase B (Akt/PKB) phosphorylation in rats with myocardial infarction. We found that puerarin (120 mg/kg/day, i.p.) could increase serum nitrite concentration in rat with myocardial ischemia (MI). It also induced the gene expression or activation of eNOS, protein expression of eNOS, and the Akt/PKB phosphorylation. From these results, we suggested that puerarin could increase serum nitric oxide level of rat with myocardial infarction, which should be one of the mechanisms of the therapeutic effect of puerarin on CAD. The increased expression of eNOS and the Akt/PKB pathway may be the underlying mechanism by which puerarin stimulates NO production.

Puerarin induces angiogenesis in myocardium of rat with myocardial infarction.

Puerarin is a major effective ingredient extracted from the traditional Chinese medicine ge-gen (radix puerariae, RP). Recently, puerarin has been used to treat patients with coronary artery diseases (CAD). However, the mechanisms of puerarin on coronary artery diseases are still not very clear. In this study, we investigated the role of puerarin on angiogenesis in the non-ischemic and ischemic myocardium. We found that puerarin (120, 60 mg/kg, i.p.) could reduce infarct area in the heart of rat with myocardial infarction (MI). Puerarin (120 mg/kg) induced angiogenesis in the non-ischemic and ischemic myocardium, which was one of the mechanisms of curing coronary artery diseases. The gene expression or activation of vascular endothelial growth factor (VEGF), hypoxia-inducible factor 1alpha (HIF-1alpha) and endothelial nitric oxide synthase (eNOS) that correlated with angiogenesis were also induced by puerarin. From these results, we suggested that puerarin may induce therapeutic angiogenesis in myocardium of rat with MI. The mechanism may be that puerarin can induce VEGF and eNOS expression.