Application of clinical value assessment of treatment protocols in guideline development: taking the WFAS Clinical Practice Guideline of Acupuncture and Moxibustion for Migraine as an example. / 治疗方案临床价值评估在指南制定中的应用.

To enhance the clinical applicability of guidelines and provide more effective guidance for clinical practice, a clinical value assessment was conducted during the development of the World Federation of Acupuncture-Moxibustion Societies (WFAS) Clinical Practice Guideline of Acupuncture and Moxibustion for Migraine, which involved the evaluation of 59 acupuncture and moxibustion treatment protocols from randomized controlled trials (RCTs). This article introduced the methodology, content and results of the clinical value assessment of RCT-based acupuncture and moxibustion treatment protocols, which involved the integration of historical and contemporary medical evidence and expert consensus. It served as a methodological reference for the future development of acupuncture and moxibustion clinical practice guidelines.

Effectiveness of Acupuncture for Prophylactic Treatment of Migraine: A Systematic Review and Bayesian Network Meta-Analysis.

It is aimed to evaluate the effectiveness and provide evidence-based medical support for acupuncture as a prophylactic treatment for migraines. Randomized controlled trials (RCTs) from inception to April 2022 are included in 14 databases. Pairwise meta-analysis is conducted using STATA software V14.0, while Windows Bayesian Inference Using Gibbs Sampling (WinBUGS V.1.4.3) is applied to generate Bayesian Network Meta-analysis (NMA) using Markov chain Monte Carlo algorithm. Forty RCTs are included, with 4405 participants. The effectiveness of six acupuncture techniques, three types of prophylactic drugs, and psychotherapy are compared and ranked. Acupuncture outperformed prophylactic drugs in terms of diminishing visual analog scale (VAS) score, migraine attack frequency, and days during the treatment and at the 12-week follow-up. At the 12-week follow-up, the effectiveness of various interventions is ranked as follows: manual acupuncture (MA) > electroacupuncture (EA) > calcium antagonists (CA) in reducing VAS score; MA > EA > CA in reducing migraine attack frequency; MA > EA > ß-receptor blocker and CA in reducing headache attack days. Acupuncture is a promising treatment for migraine prevention. The best option of acupuncture for improving various migraine outcomes has changed over time. However, the quality of included trials and NMA inconsistency limited the credibility of the conclusion.

Steroidal alkaloid with unprecedented triheterocyclic architecture.

Veratrazine A (1), a steroidal alkaloid with a unique 6/5/5 triheterocyclic scaffold as the side chain, was isolated from Veratrum stenophyllum, and its structure was established via spectroscopic analyses and X-ray diffraction. A plausible biosynthetic pathway for 1 is proposed. Bioassy exhibits moderate anti-inflammatory activities in vitro and in vivo.

Dimeric benzylisoquinoline alkaloids from Thalictrum delavayi and their biological activities.

A phytochemical investigation of the whole plants of T. delavayi led to the isolation of five new dimeric benzylisoquinoline alkaloids, thalidelavines A-E (1-5), together with six known congeners (6-11). The structures and absolute configurations of new compounds were established based on analyses of spectroscopic data, ECD calculations, and single crystal X-ray crystallography. Thalidelavines A-E (1-5) were structurally complex bisbenzylisoquinoline alkaloids with various configurations. These isolated alkaloids were evaluated for their cytotoxic and immunosuppressive effects. Among them, both 9 and 10 displayed significant cytotoxicities against T98G cell lines with an IC50 value of 2.1 µM, compared with the positive CPT-11 (IC50 = 3.0 µM). In addition, 5-7 showed remarkable immunosuppressive effects. These findings not only enrich the structural diversity of bisbenzylisoquinoline alkaloids, but also provide potential candidates for the further development of the antitumor and immunosuppressive agents.

[Study on key outcome indexes in treatment of migraine with acupuncture and moxibustion].

OBJECTIVE: To identify the key outcome indexes in treatment of migraine with acupuncture and moxibustion. METHODS: Using literature research, questionnaire survey and consensus conference, the key outcome indexes in treatment of migraine with acupuncture and moxibustion were screened and prioritized. RESULTS: The critical outcome indexes for the treatment in attack stage of migraine included 6 effectiveness outcome indexes (headache intensity, headache duration, headache relieve time, effectiveness and level of headache relief within 2 h, headache-related quality of life, level of headache relief within 24 h) and 1 safety outcome index (incidence of serious adverse reactions). The critical outcome indexes for prophylactic treatment included 6 effectiveness outcome indexes (headache day, headache frequency, headache intensity, effective rate, headache-related quality of life, health-related quality of life) and 1 safety outcome index (incidence of serious adverse reactions). CONCLUSION: In terms of the attack stage treatment and prophylactic treatment with acupuncture and moxibustion, the outcome indexes are different, among which, those can directly reflect the conditions of migraine should be optioned in priority. To assess the effectiveness of attack stage, the headache intensity is preferred, using the visual analogue scale (VAS) score, and the preferred time is 2 hours after treatment. Regarding the effectiveness of prophylactic treatment, the headache day, headache frequency and headache intensity should be firstly considered in the assessment, in which, the preferred time for assessment is 12 weeks into treatment, while, the best time for follow-up should be 12 weeks after treatment completion. When the quality of life is considered, the migraine-specific quality of life questionnaire (MSQ) is the top option. For either the attack stage treatment or the prophylactic treatment, the high attention should be laid on the outcome indexes for safety and medical economics evaluation.

Discovery of potent immune-modulating molecule taccaoside A against cancers from structures-active relationships of natural steroidal saponins.

BACKGROUND: In recent years, the T-cell therapy and immune checkpoint inhibitors toward CTLA-4 and PD-1/PD-L1 axis antibody therapy have acquired encouraging success. However, most of patients were still not benefited with lots of troubles, such as low penetration of tissues/cells, strong immunogenicity and cytokine release syndrome, and long manufacturing process and expensive costs. By contrast, the immune-modulating small molecules possessed natural advantages to overcome these obstacles and might achieve greater success. PURPOSE: Exploring the potent immune-modulating natural small molecules and revealing what kinds of molecules or structures with the immunomodulatory activity against cancers. METHODS: A novel non-cytotoxic T-cell immunomodulating screening model was used to identify the cytotoxic/selective/immunomodulatory bioactivity for 148 natural steroidal saponins. The structure-activity relationships (SARs) research was used to reveal the key groups for immunomodulation/cytotoxicity/selectivity. The negative selection was used to isolate and purify the T-cell. The cell viability assay was used to measure the anti-cancer effect in vitro. The ELISA assay was used to detect the cytokines for IL-1ß, IL-6, TNF-α, IFN-γ, IL-12, perforin and granzyme B (GZMB). The western blotting assay was used to research the immunomodulatory mechanism. The siRNA knockdown was used to generate the IFN-γ resistant melanoma cells. The NOG immune-deficient mice were used to evaluate the anti-tumor efficacy in vivo. The peripheral blood samples from 10 cancer patients were used to detect the broad population anti-tumor efficacy. RESULTS: It was reported that the correlation among structures and immunomodulation/ cytotoxicity/selectivity, in which opening ring-F with 26-O-glucopyranosyl, disaccharide and trisaccharide chains at C-3, steric hindrance and polarity of C-22 were key immunomodulatory groups. Moreover, taccaoside A was identified as the most potent candidate against cancer cells, including non-small cell lung cancer, triple negative breast cancer, and the IFN-γ resistant melanoma, partly through enhancing T lymphocyte mTORC1-Blimp-1 signal to secrete GZMB. Besides, 10 patients derived T-cell also would be modulated against cancer cells in vitro. Moreover, the overall survival was great extended (>140 days vs 93 days) with nearly 100% tumor burden disappearance (0 mm3vs 1006 ± 79.5 mm3) in mice. CONCLUSION: This work demonstrated one possibility for this concerned purpose, and identified a potent immune-modulating natural molecule taccaoside A, which might contribute to cancer immunotherapy in future.

Que Zui tea ameliorates hepatic lipid accumulation and oxidative stress in high fat diet induced nonalcoholic fatty liver disease.

In this study, the protective effects of hot water (QW) and aqueous-ethanol extracts (QA) from Que Zui tea on non-alcoholic fatty liver disease (NAFLD) were investigated. Quantitative and qualitative analysis revealed that QW and QA were rich in polyphenols, especially 6'-O-caffeoylarbutin. Both QW and QA significantly reduced body weight and liver index, increased serum levels of high density lipoprotein cholesterol (HDL-C), and decreased the levels of total cholesterol (TC), triglyceride (TG), nonesterified free fatty acids (NEFA) and low density lipoprotein cholesterol (LDL-C) in NAFLD rats induced high fat diet. Furthermore, the contents of TC, TG, NEFA, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the liver tissues were inhibited after QW and QA administration. Histopathological examination showed that QW and QA significantly reduced liver lipid accumulation of NAFLD rats. In addition, QW and QA could enhance increase the activity of antioxidant (glutathione, superoxide dismutase and catalase) in the liver by regulation Nrf2 signaling pathway, thereby alleviating liver damage caused by lipid peroxidation. QW and QA activated AMPK/PPAR-α signaling pathway by increasing the expression of adiponectin and its receptor AdipoR2, thereby reducing fat production and enhancing fatty acid ß oxidation. These data suggested that QW and QA had the potential to in the prevention and treatment of NAFLD.

Phenolic Constituents, Antioxidant and Cytoprotective Activities, Enzyme Inhibition Abilities of Five Fractions from Vaccinium dunalianum Wight.

The bud of Vaccinium dunalianum Wight has been traditionally consumed as health herbal tea by "Yi" people in Yunnan Province, China, which was locally named "Que Zui tea". This paper studied the chemical constituents of five fractions from Vaccinium dunalianum, and their enzyme inhibitory effects of α-glucosidase and pancreatic lipase, antioxidant activity, and cytoprotective effects on H2O2-induced oxidative damage in HepG2 cells. The methanol extract of V. dunalianum was successively partitioned with petroleum ether (PF), chloroform (CF), ethyl acetate (EF), n-butanol (BF), and aqueous (WF) to obtain five fractions. The chemical profiling of the five fractions was analyzed by ultra-high-performance liquid chromatography coupled with a tandem mass spectrometry (UHPLC-MS/MS), and 18 compounds were tentatively identified. Compared to PF, CF, BF and WF, the EF revealed the highest total phenols (TPC) and total flavonoids (TFC), and displayed the strongest enzyme inhibition ability (α-glucosidase and pancreatic lipase) and antioxidant capacity (DPPH, ABTS and FRAP). Furthermore, these five fractions, especially EF, could effectively inhibit reactive oxygen species (ROS) production and cell apoptosis on H2O2-induced oxidative damage protection in HepG2 cells. This inhibitory effect might be caused by the up-regulation of intracellular antioxidant enzyme activity (CAT, SOD, and GSH). The flavonoids and phenolic acids of V. dunalianum might be the bioactive substances responsible for enzyme inhibitory, antioxidant, and cytoprotective activities.

Glutamine Maintains Satellite Glial Cells Growth and Survival in Culture.

Satellite glial cells (SGCs) tightly surround neurons and modulate sensory transmission in dorsal root ganglion (DRG). At present, the biological property of primary SGCs in culture deserves further investigation. To reveal the key factor for SGCs growth and survival, we examined the effects of different culture supplementations containing Dulbecco's Modified Eagle Medium (DMEM)/F12, DMEM high glucose (HG) or Neurobasal-A (NB). CCK-8 proliferation assay showed an increased proliferation of SGCs in DMEM/F12 and DMEM/HG, but not in NB medium. Bax, AnnexinV, and propidium iodide (PI) staining results showed that NB medium caused cell death and apoptosis. We showed that glutamine was over 2.5 mM in DMEM/F12 and DMEM/HG, whereas it was absence in NB medium. Interestingly, exogenous glutamine application significantly reversed the poor proliferation and cell death of SGCs in NB medium. These findings demonstrated that DMEM/F12 medium was optimal to get high-purity SGCs. Glutamine was the key molecule to maintain SGCs growth and survival in culture. Here, we provided a novel approach to get high-purity SGCs by changing the key component of culture medium. Our study shed a new light on understanding the biological property and modulation of glial cells of primary sensory ganglia.

6'-O-Caffeoylarbutin from Que Zui tea ameliorates acetaminophen-induced liver injury via enhancing antioxidant ability and regulating the PI3K signaling pathway.

Que Zui tea (QT), a traditional herbal tea in China, has a significant hepatoprotective effect. 6'-O-Caffeoylarbutin (CA) is the most abundant chemical compound in the QT. However, the hepatoprotective effect of CA has not been investigated. This study is aimed to evaluate the protective effect of CA on acetaminophen (APAP) induced hepatotoxicity in vivo and in vitro and its possible underlying mechanism. In APAP-induced HepG-2 cells, CA inhibited intracellular ROS accumulation and cell apoptosis, and improved the expression of antioxidants including SOD, CAT and GSH. In APAP-administrated mice, CA pretreatment remarkably ameliorated the histopathological damage and inflammatory response, and antioxidant enzyme activity in the serum and liver tissues. Moreover, the immunohistochemistry and immunofluorescence assay results revealed that the CA markedly reduced ROS production and apoptosis, and activated antioxidant transcription factor Nrf2 in the liver. Meanwhile, molecular docking results showed that the strong binding force of CA and PI3K was due to the higher number of hydrogen- and π-bonds with active site residues. Notably, CA pretreatment significantly regulated the expression of PI3K, Akt, Nrf2, NQO1, HO-1, Bcl-2, Bax, caspase-3, and caspase-9 proteins in APAP-treated liver tissues. These data demonstrated that CA had a protective effect against APAP-induced hepatotoxicity via regulating the PI3K/Akt and Nrf2 signaling pathway.

Hypoglycemic and hypolipidemic effects of Epigynum auritum in high fat diet and streptozotocin-induced diabetic rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Epigynum auritum is mainly distributed in Southwest China, and has been used as a "dai" folk medicine with promising Besides, the leaves and barks of E. auritum have detoxifying, analgesic and relieving swelling effects. Previous studies evidenced that E. auritum was rich in pregnanes and their glycosides. However, the hypoglycemic and hypolipidemic effects of the extract from E. auritum (EAE) and its molecular mechanism are still not studied. AIM OF THE STUDY: The aim of this study is to investigate the hypoglycemic and hypolipidemic effects of EAE on high-fat diet and streptozocin-induced type 2 diabetic rats. MATERIALS AND METHODS: The high-fat diet and streptozocin induced type 2 diabetic model was established. The diabetic rats were treated with 70% ethanol extract of E. auritum (100 and 300 mg/kg/d) or metformin (DMBG, 100 mg/kg/d) every day for 4 weeks. Fasting blood glucose was recorded weekly. The phenotypic changes were evaluated by the measurement of biochemical indexes and immunohistochemical. The expressions of signaling-related proteins were explored by western blotting. RESULTS: EAE could effectively regulate the metabolism of glucose and lipids in diabetic rats by increasing insulin sensitivity. In addition, EAE ameliorated the oxidative stress damage and further mitigated the liver, kidney, and pancreatic damage. Mechanism research results show that EAE treatment increased the phosphorylation of Akt, AMPK and GSK-3ß, up-regulated the expression of GLUT-2, GLUT-4 and PPAR-α, and reduced PPAR-γ and FAS expressions. CONCLUSION: EAE exhibited significant hypoglycemic and hypolipidemic effects in HFD/STZ-induced diabetes rats. The mechanism may be related to the effective upregulation of AMPK/Akt/GSK-3ß pathway and the decreased expression of PPAR-γ and FAS. It could be a promising natural product with potential value for the development of drugs to prevent or treat type 2 diabetic.

Anti-leukemic effect and molecular mechanism of 11-methoxytabersonine from Melodinus cochinchinensis via network pharmacology, ROS-mediated mitochondrial dysfunction and PI3K/Akt signaling pathway.

Melodinus cochinchinensis (Lour.) Merr. is a Yunnan endemic folk medicine. Our previous study showed that 11-methoxytabersonine (11-MT) isolated from M. cochinchinensis has strong cytotoxicity on human T-ALL cells, but its molecular mechanism has not been studied. In current study, the cytotoxicity and possible mechanism of 11-MT on T-cell acute lymphoblastic leukemia was explored using network pharmacology and molecular biology techniques. 11-MT significantly inhibited the cell proliferations on different four human T-ALL cells (MOLT-4, Jurkat, CCRF-CEM, and CEM/C1 cells). 11-MT triggered ROS accumulation, calcium concentration and cell apoptosis, and decreased the mitochondrial membrane potential (MMP) in human T-ALL cells, especially MOLT-4 cells. Western blot analysis showed that it can induce MOLT-4 cell apoptosis by up-regulating PI3K/Akt signaling pathway. Therefore, 11-MT induces human T-ALL cells apoptosis via up-regulation of ROS-mediated mitochondrial dysfunction and down-regulation of PI3K/Akt/mTOR signaling pathway.

The effect of ultra-high pretreatment on free, esterified and insoluble-bound phenolics from mango leaves and their antioxidant and cytoprotective activities.

Ultra-high pressure (UHP) is a novel non-thermal pretreatment method in food processing for improving the extraction yield of polyphenols and functional properties. The present work investigated the phenolic profiles, antioxidant activities, and cytoprotective effects of the free, esterified, and insoluble-bound phenolic fractions from mango leaves before and after ultra-high pressure (UHP) treatment. UHPLC-Q-Orbitrap-MS/MS analysis resulted in the identification of 42 phenolic compounds in the different phenolic forms. UHP pretreatment could significantly influence the contents of total phenols, total flavonoids and individual compounds in the different phenolic fractions (p < 0.05). After UHP pretreatment, these phenolic fractions exhibited greater antioxidant activity, and inhibited reactive oxygen species production and cell apoptosis (p < 0.05). Meanwhile, IBP were the most potential antioxidative and cytoprotective ingredients. Therefore, UHP pretreated mango leaves with enhanced bioactivity could be used as biological agents in the health food industry to improve its application and economic values.

Phenolic constituents, antioxidant activity and neuroprotective effects of ethanol extracts of fruits, leaves and flower buds from Vaccinium dunalianum Wight.

Vaccinium dunalianum Wight is an important healthy tea resource in China with health benefits. The chemical compositions and the possible bioactive substances in its fruits, leaves and flower buds extracts (FE, LE and FBE) were identified and characterized by UHPLC-HRMS/MS. Consequently, FE, LE and FBE were rich in phenolic and flavonoid compounds. Among them, 21 compounds were identified, and the main components were chlorogenic acid, quinic acid and 6'-O-caffeoylarbutin. Furthermore, their neuroprotection and mechanism on H2O2-induced neurotoxicity in PC12 cells were investigated. All the different concentrations of FE, LE and FBE were apparently inhibited the H2O2-induced ROS generation and apoptosis on PC12 cells. FBE showed stronger neuroprotective activity against H2O2-induced PC12 cell damage than those of FE and LE. The mechanism of neuroprotective effect might be related to the upregulation of endogenous antioxidant enzymes expressions and activation of the Nrf2/HO-1 signaling pathway.

Anti-inflammatory and analgesic monoterpenoid indole alkaloids of Kopsia officinalis.

ETHNOPHARMACOLOGICAL RELEVANCE: "Ya gai", an important part of Dai medical theory, is traditionally recognized as an antidote. Kopsia officinalis Tsiang et P. T. Li is a "Ya gai" related medicine and has been widely used by Dai people for the treatment of pain and inflammation. Previous literature on title species suggested that monoterpenoid indole alkaloids (MIAs) could be its main bioactive components. However, the specific bioactive ingredients for inflammation-related treatment are still unrevealed, which inspired us to conduct a phytochemical and pharmacological investigation related to its traditional use. AIM OF THE STUDY: To support the traditional use of K. officinalis by assessing the anti-inflammatory and analgesic effects of its purified MIAs. MATERIAL AND METHODS: Compounds were isolated and purified from the barks and leaves of K. officinalis using diverse chromatographic methods. The structures were established by means of extensive spectroscopic analyses and quantum computational technique. The anti-inflammatory activities of the purified MIAs were evaluated in vitro based on the suppression of lipopolysaccharide-activated inflammatory mediators (COX-2, IL-1ß, and TNF-α) in RAW 264.7 macrophage cells. Anti-inflammatory and analgesic activities in vivo were assessed with carrageenan-induced paw edema and acetic acid-stimulated writhing in mice models. RESULTS: 23 MIAs including four new compounds were obtained and structurally established. Most of isolates showed significant anti-inflammatory effects in vitro by inhibiting inflammatory mediators (COX-2, IL-1ß, and TNF-α). Further pharmacological evaluation in vivo revealed that 12-hydroxy-19(R)-hydroxy-ibophyllidine (1) and 11,12-methylenedioxykopsinaline N4-oxide (5) remarkably decreased the number of writhing, while kopsinic acid (8), (-)-kopsinilam (12), and normavacurine-21-one (20) significantly relieved paw edema, respectively, even better than the positive control aspirin. CONCLUSIONS: The in vitro and in vivo findings supported the traditional use of K. officinalis with respect to its anti-inflammatory and analgesic effect, as well as provided potent bioactive MIAs for further chemical modification and pharmacological investigation.

Phytochemicals and Allelopathy of Induced Water Hyacinth against Microcystis aeruginosa.

Induced water hyacinth with purple roots (PRWH) exerts a significant inhibitory effect on the growth of blue-green algae. Interestingly, its chemical constituents differ from those of wild-type water hyacinth and have not yet been reported. This study aimed to explore the chemical constituents of PRWH and its bioactive components serving as allelopathic agents against blue-green algae. Phytochemical investigation of the bioactive ethyl acetate fraction of a crude methanol extract from PRWH led to the isolation of 56 compounds, including 11 new phenylphenalene derivatives. The structures of these compounds were elucidated by comprehensive analyses through NMR, HRMS, and X-ray techniques. Bioactivity evaluation against Microcystis aeruginosa indicated that compounds 7, 12, 15, 37, 39, 45, and 47 potently inhibited blue-green algae growth.

Antioxidant and Cytoprotective Effects of New Diarylheptanoids from Rhynchanthus beesianus.

Rhynchanthus beesianus (Zingiberaceae) has been an important food spice and vegetable in southern China. Fifteen phenolic compounds (1-15) including three new diarylheptanoids, rhynchanines A-C (1-3) and one new phenylpropanoid, 4-O-methylstroside B (9), were isolated from R. beesianus rhizomes. The structures of new compounds were elucidated by comprehensive analyses through NMR, HRMS technique, acid hydrolysis, and Mosher's reaction. Among them, compound 5 is the first isolated natural product and its NMR data are reported. Most of the isolated compounds, especially 3-6 and 8, showed significant antioxidant activities on DPPH, ABTS+ radical scavenging, and FRAP assays. Furthermore, the antioxidant phenolic compounds were evaluated for their cytoprotective capacity against H2O2-induced oxidative stress in HepG-2 cells. Compounds 3 and 5 could significantly inhibit reactive oxygen species production, and compounds 3, 5, and 6 could remarkably prevent the cell apoptosis. Then, the R. beesianus rhizome, which contained phenolic compounds, might serve as a functional food for potential application on preventing oxidative stress-connected diseases.

4,21-Secovincanol, a Novel Immunosuppressive Monoterpenoid Indole Alkaloid from Kopsia Hainanensis.

4,21-Secovincanol (1), a novel C-21/N-4 cleavage monoterpenoid indole alkaloid, along with four analogs (2-5), were obtained from the aerial parts of Kopsia hainanensis. Structurally, compound 1 might be a derivative of epivincanol (2) via C-21/N-4 cleavage. Their structures were confirmed by means of comprehensive spectroscopic data analysis and comparison with the reported data. All isolates significantly inhibited Con A-stimulated mice splenocytes proliferation at 10-40 µM in a dose-dependent manner in vitro. Especially, compound 3 exhibited potent activities comparable to positive control (Dexamethasone, DXM).

Diverse isoquinolines with anti-inflammatory and analgesic bioactivities from Hypecoum erectum.

ETHNOPHARMACOLOGICAL RELEVANCE: Hypecoum erectum has been used extensively in folk medicine to treat inflammation, fever, and pain. However, few investigations have been carried out on the biological activities related to its traditional use. The chemical constituents of this plant along with their anti-inflammatory and analgesic effects have yet to be revealed. AIM OF THE STUDY: This study aimed to support the traditional use of H. erectum by first assessing its anti-inflammatory and analgesic effects and then investigating its chemical constituents to identify any anti-inflammatory and/or analgesic compounds. MATERIAL AND METHODS: The in vivo anti-inflammatory and analgesic activities of the MeOH extract (ME), total alkaloid (AL), and non-alkaloid (Non-AL) fractions of H. erectum at doses of 200, 100, and 50 mg/kg and four major constituents (20, 21, 22, and 27) at doses of 100 and 50 mg/kg delivered via intragastrical administration were evaluated using carrageenan-induced paw edema and acetic acid-stimulated writhing animal models. A phytochemical study of the bioactive (AL) fraction was conducted using various chromatographic techniques, and the structures of the obtained isoquinolines were identified by multiple spectroscopic analyses and quantum chemical computations. Moreover, the anti-inflammatory activities of all the isolates were assessed in vitro based on the suppression of lipopolysaccharide-activated inflammatory mediators (COX-2, IL-1ß, and TNF-α) in RAW 264.7 macrophage cells. RESULTS: At the dose of 200 mg/kg, the three fractions (ME, AL, and Non-AL) of H. erectum ameliorated the paw edema by carrageenan-stimulated and reduced the number of writhing by acetic acid-induced in mice compared to the model group, with the AL fraction showing the most potent effects. Subsequent phytochemical investigation of the AL fraction led to the isolation of six new isoquinoline alkaloids (1-6) as well as 23 known analogues (7-29). However, compared to common isoquinolines, compounds 1-4 possess an additional nitrogen atom, while compound 5 has two additional nitrogen atoms. These additional atoms enrich the diversity of natural isoquinoline alkaloids. Further pharmacological evaluation in vivo revealed that the four major constituents (20, 21, 22, and 27) significantly relieved paw edema at 100 mg/kg, while protopine (20) and oxyhydrastinin (27) remarkably decreased the number of writhing at 100 mg/kg. In addition, most of the isolates displayed anti-inflammatory effects, as indicated by the inhibition of inflammatory mediators (COX-2, IL-1ß, and/or TNF-α) in vitro at a treatment concentration of 5 µg/mL. trans-benzindenoazepines (13), protopine (20), and 1,3,6,6-tetramethyl-5,6,7,8-tetrahyboisoquiolin-8-one (25) showed comparable anti-inflammatory activity to dexamethasone by inhibiting the secretion of IL-1ß. CONCLUSIONS: This investigation validated the traditional use of H. erectum by assessing its anti-inflammatory and analgesic effects. Phytochemical investigation revealed the diversity and novelty of the natural isoquinoline alkaloids in H. erectum. Four major isoquinolines were identified as the bioactive constituents of H. erectum. The findings provide scientific justification to support the traditional application of H. erectum for treating inflammatory and pain disorders.

Pharmacological effects of indole alkaloids from Alstonia scholaris (L.) R. Br. on pulmonary fibrosis in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Alstonia scholaris (L.) R. Br. (Apocynaceae) is a Dai folk medicine for the treatment of lung diseases in China. AIM OF THE STUDY: The present study investigated the anti-pulmonary fibrosis effects of total alkaloids (TA) and the potential active ingredients and its possible mechanism. MATERIALS AND METHODS: After intratracheal instillation of bleomycin (BLM, 5 mg/kg), mice were divided into ten groups, and orally treated with the corresponding samples once daily for 28 days. The effect of indole alkaloids was determined through analysis of cytokines, as well as histopathological examinations and gene expressions. RESULTS: Severe lung fibrosis was observed in the BLM-treated mice on day 28. However, the administration of TA significantly ameliorated the pathological changes in the lungs, decreased the content of Krebs von den Lungen-6, lactate dehydrogenase, transforming growth factor-ß (TGF-ß), hydroxyproline, type I collagen, and malonaldehyde, and enhanced the activity of superoxide dismutase in the serum and lung tissues. In addition, the enhanced TGF-ß and matrix metalloproteinase-1 (MMP-1) expressions in BLM-induced mice were obviously weakened by indole alkaloids, as well as the ratio of matrix metalloproteinase-1 to tissue inhibitor of metalloproteinase-1 was decreased. Moreover, picrinine and scholaricine yielded markedly better values in the aforementioned indices than those in other samples, indicating that they may be the active ingredients of alkaloids. CONCLUSIONS: TA exerted protective effects against BLM-induced pulmonary fibrosis by reducing collagen deposition through TGF-ß/MMP-1 pathway.