Antidepressant effect of Perilla frutescens essential oil through monoamine neurotransmitters and BDNF/TrkB signal pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional Chinese medicine posits that affect-mind ill-being is the primary cause of depression, with Qi movement stagnation as its pathogenesis. As such, clinical treatment for depression should prioritize regulating Qi and relieving depressive symptoms. The pharmacological properties of traditional Chinese medicine indicate that Perilla frutescens may have potential therapeutic effects on depression and other neuropsychiatric diseases due to its ability to regulate Qi and alleviate depressive symptoms. Although previous studies have reported the antidepressant effects of Perilla frutescens, the mechanism underlying PFEO inhalation-mediated antidepressant effect remains unclear. AIM OF THE STUDY: The aim of this investigation is to elucidate the antidepressant mechanisms of PFEO by examining its effects on monoamine neurotransmitters and the BDNF/TrkB signaling pathway. MATERIALS AND METHODS: The CUMS rat model of depression was established, and the depressive state of the animals was assessed through sucrose preference and forced swim tests. ELISA assays were conducted to determine monoamine neurotransmitter levels in the hippocampus and cerebral cortex of rats. Immunohistochemistry, western blotting, and RT-PCR experiments were employed to investigate the BDNF/TrkB signaling pathway's regulation of depression via PFEO inhalation. RESULTS: It has been observed that inhalation administration of PFEO can significantly enhance the preference for sugar water in CUMS rats and reduce their immobility time during forced swimming. Additionally, there was an increase in the levels of monoamine transmitters in both the hippocampus and cerebral cortex of these rats. Furthermore, there was an upregulation in the expression levels of BDNF and TrkB positive cells as well as BDNF and TrkB proteins within both regions, along with increased BDNF mRNA and TrkB mRNA expression levels. CONCLUSION: The antidepressant effect of PFEO via inhalation administration is speculated to be mediated through the monoamine neurotransmitters and BDNF/TrkB signaling pathway.

Qinzhizhudan formula dampens inflammation in microglia polarization of vascular dementia rats by blocking MyD88/NF-κB signaling pathway: Through integrating network pharmacology and experimental validation.

ETHNOPHARMACOLOGICAL RELEVANCE: Qinzhizhudan Formula (QZZD) is composed of Scutellaria baicalensis Georgi (Huang Qin) extract, Gardenia jasminoides (Zhizi) extract and Suis Fellis Pulvis (Zhudanfen) (ratio of 4:5:6). This formula is optimized from Qingkailing (QKL) injection. Regarding brain injury, QZZD is protective. However, the mechanism by which QZZD treats vascular dementia (VD) has not been elucidated. AIM OF THE STUDY: To ascertain QZZD's effect on the treatment of VD and further investigate the molecular mechanisms. MATERIALS AND METHODS: In this study, we screened the possible components and targets of QZZD against VD and microglia polarization using network pharmacology (NP), then an animal model of bilateral common carotid artery ligation method (2VO) was induced. Afterward, The Morris water maze was employed to evaluate cognitive ability, and pathological alterations in the CA1 area of the hippocampus were detected using HE and Nissl staining. To confirm the affect of QZZD on VD and its molecular mechanism, the contents of inflammatory factors IL-1ß, TNF-α, IL-4, and IL-10 were performed to detect by ELISA, the phenotype polarization of microglia cells was detected by immunofluorescence staining, and the expressions of MyD88, p-IκBα and p-NF-κB p65 in brain tissue were detected by western blot. RESULTS: A total of 112 active compounds and 363 common targets of QZZD, microglia polarization, and VD were identified, according to the NP analysis. 38 hub targets were screened out from the PPI network. GO analysis and KEGG pathway analysis showed that QZZD may regulate microglia polarization through anti-inflammatory mechanism such as Toll-like receptor signaling pathway and NF-κB signaling pathway. The further results showed that QZZD can alleviate the memory impairment induced by 2VO. QZZD profoundly rescued brain hippocampus neuronal damage and increased the number of neurons. These advantageous outcomes were linked to the control of microglia polarization. QZZD decreased M1 phenotypic marker expression while increasing M2 phenotypic marker expression. QZZD may controll the polarization of the M1 microglia by blocking the core part of Toll-like receptor signaling pathway, that is the MyD88/NF-κB signaling pathway, which reduced the neurotoxic effects of the microglia. CONCLUSION: Here, we explored the anti-VD microglial polarization characteristic of QZZD for the first time and clarified its mechanisms. These findings will provide valuable clues for the discovery of anti-VD agents.

Astragalus saponins protect against extrahepatic and intrahepatic cholestatic liver fibrosis models by activation of farnesoid X receptor.

ETHNOPHARMACOLOGICAL RELEVANCE: Cholestatic Liver Fibrosis (CLF) is a hepatobiliary disease that typically arises as a late-stage complication of cholestasis, which can have multiple underlying causes. There are no satisfactory chemical or biological drugs for CLF. Total Astragalus saponins (TAS) are considered to be the main active constituents of the traditional Chinese herb Astragali Radix (AR), which has the obvious improvement effects for treating CLF. However, the mechanism of anti-CLF effects of TAS is still unclear. AIM OF THE STUDY: The present study was undertaken to investigate the therapeutic effects of TAS against bile duct ligation (BDL) and 3, 5-diethoxycarbonyl-1,4-dihydroxychollidine (DDC) -induced CLF models and to reveal the potential mechanism to support its clinic use with scientific evidence. MATERIALS AND METHODS: In this study, BDL-induced CLF rats were treated with TAS (20 mg/kg, 40 mg/kg) and DDC-induced CLF mice were treated with 56 mg/kg TAS. The therapeutic effects of TAS on extrahepatic and intrahepatic CLF models were evaluated by serum biochemical analysis, liver histopathology and hydroxyproline (Hyp). Thirty-nine individual bile acids (BAs) in serum and liver were quantified by using UHPLC-Q-Exactive Orbitrap HRMS. qRT-PCR, Western blot and immunohistochemistry analysis were used to measure the expression of liver fibrosis and ductular reaction markers, inflammatory factors and BAs related metabolic transporters, along with nuclear receptor farnesoid X receptor (FXR). RESULTS: The serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total bilirubin (TBiL), direct bilirubin (DBiL) and contents of liver Hyp were dose-dependently improved after treatment for TAS in BDL and DDC- induced CLF models. And the increased levels of ALT and AST were significantly improved by total extract from Astragali radix (ASE) in BDL model. The liver fibrosis and ductular reaction markers, α-smooth muscle actin (α-SMA) and cytokeratin 19 (CK19), were significantly ameliorated in TAS group. And the liver expression of inflammatory factors: interleukin 6 (IL-6), tumor necrosis factor-α (TNF-α) and interleukin 1ß (IL-1ß) were significantly decreased after TAS treatment. In addition, TAS significantly ameliorated taurine-conjugated BAs (tau-BAs) levels, particularly α-TMCA, ß-TMCA and TCA contents in serum and liver, which correlated with induced expressions of hepatic FXR and BAs secretion transporters. Furthermore, TAS significantly improved short heterodimer partner (SHP), cholesterol 7α-hydroxylase (Cyp7a1), Na+ taurocholate cotransport peptide (NTCP) and bile-salt export pump (BSEP) mRNA and protein expression. CONCLUSIONS: TAS exerted a hepatoprotective effect against CLF by ameliorating liver injury, inflammation and restoring the altered tau-BAs metabolism to produce a positive regulatory effect on FXR-related receptors and transporters.

Qinlian hongqu decoction ameliorates hyperlipidemia via the IRE1-α/IKKB-ß/NF-κb signaling pathway: Network pharmacology and experimental validation.

ETHNOPHARMACOLOGICAL RELEVANCE: Qinlian Hongqu decoction (QLHQD) is a traditional Chinese medicine (TCM) formula. It has previously been found to mitigate hyperlipidemia, although its mechanism requires further clarification. AIM OF THE STUDY: This study explored QLHQD's mechanism in treating hyperlipidemia based on network pharmacology and experimental validation. MATERIALS AND METHODS: The components of QLHQD were analyzed by means of ultrahigh performanceliquid chromatography-quadrupole/orbitrapmass spectrometry (UHPLC-Q-Orbitrap-HRMS) and the targets of hyperlipidemia were predicted using the Swiss ADME, GeneCards, OMIM, DrugBank, TTD, and PharmGKB databases. A drug-component-target-disease network was constructed using Cytoscape v3.7.1. Moreover, Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analyses were performed using the Bioinformatics platform. Based on the KEGG results, the non-alcoholic fatty liver disease signaling pathways were selected for experimental validation in an animal model. RESULTS: We identified 34 components of QLHQD, 94 targets of hyperlipidemia, and 18 lipid metabolism-related pathways from the KEGG analysis. The results of the animal experiment revealed that QLHQD alleviated lipid metabolism disorders, obesity, insulin resistance, and inflammation in rats with hyperlipidemia induced by high-fat diets. Additionally, it reduced the expression of IRE1-α, TRAF2, IKKB-ß, and NF-κB proteins in the liver of hyperlipidemic rats. CONCLUSION: QLHQD is able to significantly mitigate hyperlipidemia induced via high-fat diets in rats. The mechanism of action in this regard might involve regulating the IRE1-α/IKKB-ß/NF-κB signaling pathway in the liver, thereby attenuating inflammatory responses and insulin resistance.

Therapeutic potential of elema-1,3,7(11),8-tetraen-8,12-lactam from Curcuma wenyujin on diabetic retinopathy via anti-inflammatory and anti-angiogenic pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: In traditional Chinese medicine, the causes of diabetic retinopathy (DR) are blood stasis and heat. Curcuma wenyujin Y. H. Chen & C. Ling and its extracts have the effects of promoting blood circulation to remove blood stasis, clearing the heart, and cooling the blood, and have been used in the treatment of DR. Elema-1,3,7 (11),8-tetraen-8,12-lactam (Ele), an N-containing sesquiterpene isolated from this plant. However, the anti-inflammatory and anti-angiogenic effects of Ele and its therapeutic potential in DR are still unknown. AIM OF THE STUDY: To evaluate the anti-inflammatory and anti-angiogenic effects of Ele and its therapeutic potential in DR. MATERIALS AND METHODS: In vitro, anti-inflammatory and anti-angiogenic effects were assessed using TNF-α or VEGF-stimulated HUVECs. Protein expression was analyzed using Western blotting. ICAM-1 and TNF-α mRNA expressions were analyzed using real-time quantitative RT-PCR. The therapeutic potential in DR was assessed using both animal models of STZ-induced diabetes and oxygen-induced retinopathy. The retinal vascular permeability was measured using Evans blue, and the quantitation of retinal leukostasis using FITC-coupled Con A. The retinal neovascular tufts were analyzed using fluorescein angiography and counting pre-retinal vascular lumens. RESULTS: Ele inhibited NF-κB pathway, and ICAM-1, TNF-α mRNA expression in TNF-α- stimulated HUVECs. It also inhibits the multistep process of angiogenesis by inhibiting the phosphorylation of VEGFR2 and its downstream signaling kinases Src, Erk1/2, Akt, and mTOR in VEGF-stimulated HUVECs. Intravitreal injection of Ele can significantly reduce retinal microvascular leakage, leukostasis, and expression of ICAM-1, TNF-α in diabetic rats and inhibits oxygen-induced retinal neovascularization and VEGFR2 phosphorylation in OIR mice. CONCLUSIONS: Ele has anti-inflammatory and anti-angiogenic effects through inhibiting NF-κB and VEGFR2 signaling pathways, and it may be a potential drug candidate for DR.

Xiao-Ban-Xia decoction mitigates cisplatin-induced emesis via restoring PINK1/Parkin mediated mitophagy deficiency in a rat pica model.

ETHNOPHARMACOLOGICAL RELEVANCE: As a traditional Chinese anti-emetic formula, Xiao-Ban-Xia decoction (XBXD) was recorded in Golden Chamber, and has promising anti-emetic effect on chemotherapy-induced nausea and vomiting (CINV). AIM OF THE STUDY: This study aimed to determine whether the underlying mechanism of XBXD against CINV is correlated to the restoration of cisplatin-induced PINK1/Parkin mediated mitophagy deficiency and mitigation of gastrointestinal inflammation. MATERIALS AND METHODS: The rat pica model was established by intraperitoneal injection of cisplatin 6 mg/kg. The daily kaolin consumption, food intake and body weight were recorded every 24 h. The pathological damage of gastric antrum and ileum were observed by hematoxylin-eosin staining. The levels of serum reactive oxygen species (ROS), interleukin-1ß (IL-1ß) and interleukin-1ß (IL-18) were detected by ELISA. The expression of microtubule-associated protein 1 light chain 3 (LC3) in gastric antrum and ileum was detected by Immunofluorescence staining. The levels of LC3II, P62/SQSTM1, PTEN-induced putative protein kinases (PINK1), E3 ubiquitin ligase (Parkin), AMP-dependent protein kinases (AMPK), phosphorylated AMPK (p-AMPK), nuclear factor erythroid 2-related factor (Nrf2) and kelch like ECH Associated Protein 1 (Keap1) in gastric antrum and ileum were assayed by western blotting. RESULTS: At 24 h and 72 h following cisplatin challenge, XBXD inhibited cisplatin-induced elevation of kaolin consumption, and improved the daily food intake and body weight loss in rats. Cisplatin-induced gastrointestinal histopathological damages were alleviated, and serum levels of ROS, IL-1ß and IL-18 increases were mitigated following XBXD treatments. In gastric antrum and ileum, XBXD activated AMPK-Nrf2 signaling pathway and restored cisplatin-induced PINK1/Parkin mediated mitophagy deficiency. CONCLUSIONS: XBXD significantly ameliorated CINV in a cisplatin-induced rat pica model. The underlying anti-emetic mechanism of XBXD might be related to the activation of AMPK-Nrf2 signaling pathway and the restoration of cisplatin-induced PINK1/Parkin-mediated mitophagy deficiency in the gastrointestinal tract.

Danggui Shaoyao San: Chemical characterization and inhibition of oxidative stress and inflammation to treat CCl4-induced hepatic fibrosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Danggui Shaoyao San (DSS) has effective in treating hepatic ascites and liver disease. AIM OF THE STUDY: To explore the chemical characterization of DSS and protective effect on CCl4-induced hepatic fibrosis and its mechanism, especially its anti-oxidative stress and anti-inflammation. MATERIALS AND METHODS: The chemical characterization of DSS was determined by HPLC-Q-Exactive Orbitrap MS. And the antioxidant activity of DSS in vitro was determined. The hepatic fibrosis model was established using intragastric administration of 40% CCl4/soybean oil (v/v) twice weekly for 13 weeks. From 6th week, the DSS group and the positive control group were given DSS (2, 4, 8 g/kg/d) and silymarin (50 mg/kg/d), respectively. The livers of rats were examined histologically by H&E. The ALT, AST, ALB and TBIL were determined, and hepatic fibrosis markers (HA, LN, CIV, PIIINP), oxidative stress (SOD, MDA, GST, GSH) and inflammatory factor (IL-6, TNF-α) were tested using ELISA kits. In addition, the levels of TAC, TOS, LOOH and AOPP in the liver were also determined. RESULTS: The chemical characterization of DSS was determined by HPLC-Q-Exactive Orbitrap MS. The results show that DSS mainly includes triterpenoids, monoterpenes, phenols, sesquiterpenes, butyl phthalide, etc., and DSS has good antioxidant activity in vitro. In addition, the ALT, AST and TBIL of rats were remarkably reduced after treatment with DSS at three doses. Liver histopathological analysis showed that DSS alleviated the inflammatory infiltration, hepatocyte swelling, necrosis and hepatic fibrosis induced by CCl4. DSS significantly decreased HA, IV-C, PIIINP and LN. Further determination showed that DSS significantly increased TAC, OSI and decreased TOC, LOOH and MDA, indicating that DSS could regulate redox balance and reduce lipid peroxidation in vivo. DSS also increased the activity of GST, SOD and GSH concentration. In addition, DSS also reduced IL-6 and TNF-α. CONCLUSIONS: In this study, we described the chemical characterization of DSS and found that it has good antioxidant activity. We proved that DSS has the functions of reducing oxidative stress, anti-inflammatory, protecting liver cells and reducing hepatic fibrosis.

Beneficial biological effects of Flavokawain A, a chalcone constituent from kava, on surgically induced endometriosis rat model.

ETHNOPHARMACOLOGICAL RELEVANCE: Shrub kava has long been grown and utilized, primarily in the South Pacific region, for ceremonial, religious, and social occasions. It has been used as a pain reliever and muscle relaxant in medicinal practices from the eighteenth century. Interestingly, relatively low incidence of lung cancer may attribute to the high consumption of kava products in this region. AIM OF THE STUDY: Kava extracts were used to produce the kava chalcones Flavokawain A, B and C, which have a variety of bioactivities. In the present study, we show that Flavokawain A has positive effects on endometriosis. MATERIALS AND METHODS: The endometriosis rat model was surgically induced by the autologous transplantation of endometrial tissue. Rats were evaluated for clinical ratings and lesion volume following a 6-week Flavokawain A therapy. Peritoneal fluid and blood samples were taken and ELISA assay was used to measure the cytokines and chemokines levels. Transcriptional and expression levels of Akt, PI3K, NF-kB, iNOS, Bcl-2, Bax and caspase-3 were evaluated by Western blotting and RT-qPCR. Implanted tissue sections of the rats were also analyzed by immunofluorescent and histopathological staining. RESULTS: Lesion volumes and adhesion scores were successfully decreased. Blood and peritoneal fluid levels of associated cytokines and chemokines were markedly down-regulated. Besides, Flavokawain A also mediated cell apoptosis of endometrial implants. Additionally, VEGF expression was reduced, which inhibited the angiogenesis process. As for the expression of Akt, p-Akt, PI3K, p-PI3K, and NF-kB in endometriosis lesions, Flavokawain A significantly reduced them. CONCLUSION: Flavokawain A has beneficial effects on the surgically induced endometriosis rat model, by reducing inflammation, promoting apoptosis, and decreasing angiogenesis. Our findings suggest that these effects may be mediated through the regulation of PI3K/Akt and NF-κB signaling pathways.

Shenqisherong pill ameliorates neuronal apoptosis by inhibiting the JNK/caspase-3 signaling pathway in a rat model of cervical cord compression.

ETHNOPHARMACOLOGICAL RELEVANCE: The Shenqisherong (SQSR) pill is an empirical prescription of traditional Chinese medicine (TCM), which originated from the National Chinese Medical Science Master, Shi Qi. It has been widely used in the treatment of cervical spondylotic myelopathy (CSM) and promote the recovery of spinal cord function, but underlying molecular mechanism remains unclear. AIM OF THE STUDY: The objective of this study was to confirm the neuroprotective effects of the SQSR pill. MATERIALS AND METHODS: A rat model of chronic compression at double-level cervical cord was used in vivo. The protective role of SQSR pill on CSM rats was measured by Basso, Beattie, and Bresnahan (BBB) locomotor scale, inclined plane test, forelimb grip strength assessment, hindlimb pain threshold assessment, and gait analysis. The levels of reactive oxygen species (ROS) were examined by Dihydroethidium (DHE) staining and 2',7'-Dichlorofluorescein (DCF) assay, and apoptosis was detected by TdT-mediated dUTP nick-end labeling (TUNEL) assay. The expression of apoptosis proteins was evaluated by immunofluorescence staining and Western blot. RESULTS: SQSR pill could facilitate locomotor function recovery in rats with chronic cervical cord compression, reduce local ROS in the spinal cord and downregulate the c-Jun-N-terminal kinase (JNK)/caspase-3 signaling pathway. In addition, the SQSR pill could protect primary rat cortical neurons from glutamate-treated toxicity in vitro by reducing the ROS and downregulating the phosphorylation of JNK and its downstream factors related to neuronal apoptosis meditated by the caspase cascade. Then, the neuroprotective effect was counteracted by a JNK activator. CONCLUSIONS: Together, SQSR pill could ameliorate neuronal apoptosis by restraining ROS accumulation and inhibiting the JNK/caspase-3 signaling pathway, indicating that SQSR pill could be a candidate drug for CSM.

Ameliorative effect of traditional polyherbal formulation on TNF-α, IL-1ß and Caspase-3 expression in kidneys of wistar rats against sodium fluoride induced oxidative stress.

ETHNOPHARMACOLOGICAL RELEVANCE: Sharbat-e-bazoori Motadil (SBM) is a polyherbal formulation that have been used for centuries as a part of the Unani system of medicine for renal disease. AIM OF THE STUDY: The objective of this study was to explore and validate the nephroprotective potential of sugar-free SBM (SF-SBM) and its mechanisms of action against sodium fluoride (NaF)-induced nephrotoxicity in HEK-293 cells. Additionally, the study aimed to assess the quality control of SF-SBM and investigate its effects using an in vivo rat model with pattern recognition following oral administration of SF-SBM. MATERIALS AND METHODS: The nephroprotective effect of SF-SBM was investigated using both an HEK-293 cell line and Wistar rats. Nephrotoxicity was induced in these models by administering NaF at a concentration of 600 ppm (parts per million) for a duration of seven days. The SF-SBM formulation was standardized using high-performance thin-layer chromatography (HPTLC) to assess the presence of marker compounds, namely gallic acid, quercetin, and ferulic acid. Metabolite characterization of SF-SBM was carried out using ultra-high-performance liquid chromatography mass spectrometry (UPLC-MS) with a monolithic capillary silica-based C18 column. This analytical technique allowed for the identification of bioactive substances and verification of the identified markers. Acute toxicity of SF-SBM was evaluated in Wistar rats by administering a single oral dose of 2000 mg/kg of SF-SBM. The nephroprotective efficacy of SF-SBM was further assessed at low (LD), medium (MD) and high (HD) doses of 32.1, 64.2, and 128.4 mg/kg, respectively, administered orally. Nephrotoxicity was induced in Wistar rats by adding NaF to their drinking water for seven days. Biochemical and urine markers were analyzed to evaluate the antioxidant, inflammatory, and apoptotic potential of SF-SBM. Additionally, histopathological analysis and immunohistochemical alterations in the expression of caspase-3 and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-4 (NOX-4) in kidney tissue were performed to confirm the findings of the in vivo experiments. Furthermore, in vivo pattern recognition of SF-SBM metabolites, identified through GC-MS metabolomics, and in-silico docking analysis of major metabolites in plasma were conducted to gain further insights. RESULT: Phytochemical analysis using HPTLC, TLC-bioautography, and UPLC-MS revealed the presence of several bioactive constituents in SF-SBM, including ferulic acid, gallic acid (GA), ellagic acid, quercetin, and apigenin. These compounds exhibit diverse pharmacological properties. In vitro studies demonstrated the protective effect of SF-SBM on HEK-293 cell line against nephrotoxicity. The acute toxicity study of SF-SBM at a dose of 2000 mg/kg showed no mortality or signs of toxicity throughout the 14-day observation period. In the in vivo studies, administration of NaF resulted in significant elevation (P < 0.001) of biochemical and urine parameters, indicating oxidative, inflammatory, and apoptotic stress. Histopathological examination revealed severe depletion of Bowman's capsule, and immunohistochemistry demonstrated negative immunostaining for caspase-3 and reduced NOX-4 reactions. Pre-treatment with SF-SBM significantly attenuated the elevated biochemical and urine markers, restored the antioxidant enzyme levels (such as SOD, CAT, GSH, GPx and NO), and regulated the expression of inflammatory cytokines (TNF-α, IL-1ß, CASP-3) in kidney tissue at doses of SF-SBM-MD (64.2 mg/kg) and SF-SBM-HD (128.4 mg/kg), showing comparable results to those of α-Ketoanalogue. Histopathological assessment demonstrated improvements in tissue damage. Pattern recognition analysis of SF-SBM identified the presence of 56 metabolites at different time intervals. Additionally, in-silico studies revealed strong interactions of SF-SBM with a binding energy of -6.5 and -5.6 kcal for 4C2N. CONCLUSION: The phytoconstituents present in SF-SBM play a crucial role in its nephroprotective action by acting as potent antioxidants and reducing proinflammatory and apoptotic damage in rat cells. This indicates that SF-SBM has promising potential for the treatment of nephrotoxicity.

Modified Simiaowan prevents and treats gouty arthritis via the Nrf2/NLRP3 inflammasome signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Modified Simiaowan (MSM) is a six-herb formula that has been shown to be effective in gouty arthritis (GA) has been proven, but its regulatory mechanism has not been fully elucidated. AIM OF THE STUDY: To investigate the therapeutic effects and mechanism of MSM on gouty arthritis. MATERIALS AND METHODS: Mouse J774A.1 macrophages were induced with Lipopolysaccharide (LPS) and then stimulated with Adenosine 5'-triphosphate (ATP) or Nigericin (Nig.) in presence or absence of MSM. Expression of key indicators of pro-inflammatory cytokines and the NLRP3 inflammasome signaling pathway were investigated by western blot, ELISA and qRT-PCR. Fluorescence staining and flow cytometry were performed to detect intracellular reactive oxygen species (ROS) production. Another study, the anti-inflammatory and antioxidant activities of MSM were evaluated in rats with monosodium urate (MSU) -induced gouty arthritis using ELISA, hematoxylin-eosin staining (HE) staining, immunohistochemistry, and oxidative stress kits to measure relevant inflammatory markers and oxidative stress-related biomarkers. RESULTS: ELISA and qRT-PCR results demonstrated that MSM effectively reduced the secretion and the mRNA expression levels of pro-inflammatory cytokines. Western blot results indicated that MSM can suppress the expression of NLRP3, an inflamasomes-related protein. In addition, MSM regulated the transition from M1 to M2 macrophages and upregulated the protein expression of Nrf2 and HO-1. The flow cytometry results and the fluorescence staining result were consistent with hypothesis that a large amount of ROS could be effectively cleared by MSM. However, the anti-inflammatory effect of MSM was attenuated after the use of ML385. In vivo experiments demonstrated that joint swelling was significantly attenuated and knee neutrophil infiltration was alleviated in rats given MSM. SOD and GSH-px levels were elevated significantly, while COX-2 and MDA levels decreased. The immunohistochemical results suggested that MSM could effectively inhibit the activation of the NLRP3 inflammasome and the regulation of macrophage polarization in rat synovial tissue, and remarkably enhance the expression of Nrf2 and HO-1. CONCLUSION: MSM has potent anti-inflammatory and antioxidant effects on MSU-induced gouty arthritis. MSM alleviates GA through Nrf2/HO-1/ROS/NLRP3 signaling pathway.

The ameliorative effect of Piper trioicum in attenuating cognitive deficit in scopolamine induced neurotoxicity in experimental rats.

ETHNOPHARMACOLOGICAL RELEVANCE: In traditional system of medicine, Piper species, or its components are widely used to treat many diseases including memory improvement. One of the wild species Piper trioicum Roxb. (Piperaceae) is found in South Asian countries. The whole plant is used as folk medicine to improve memory. AIM OF THE STUDY: To our knowledge, no previous research has investigated the neuroprotective activities of P. trioicum. So, we studied the ameliorative effect of P. trioicum in attenuating cognitive deficit in scopolamine induced neurotoxicity in experimental rats. MATERIALS AND METHODS: Wistar rats were exposed to scopolamine (3 mg/kg, i. p.) for 14 consecutive days, and the effect of P. trioicum (HAPT; oral, 300, 400 mg/kg) on scopolamine-invoked neurotoxicity in brain were studied. During the experimental period, behaviour analyses of rats were observed 30 min post-drug administration. The role of antioxidants of HAPT in scavenging cellular oxygen/peroxyl radicals were studied. Acetylcholinesterase and butyrylcholinesterase inhibitions, and mode of inhibition kinetics of HAPT were studied. Pathogenic cellular oxidative (MDA, GSH, SOD, and CAT), DNA damage (8-oxodG), neurochemical (acetyl- and, butyryl-cholinesterase), ß-secretase (BACE-1 and 2), MAPτ, and neuroinflammation (IL-6, TNF-α) biomarkers in extension to the histopathological observation of brain cortex were studied. GC-MS/MS analysis was carried out to investigate the presence of bioactive constituents in HAPT. RESULTS: HAPT, a rich source of phenol and flavonoid type antioxidants were responsible in quenching oxygen/peroxyl radicals and protected the cellular membrane, and lipoproteins against ROS in DPPH, ORAC, and CAPe tests. HAPT inhibited acetylcholinesterase and butyrylcholinesterase activities, and showed competitive-inhibition (reversible) towards cholinesterase activities. HAPT-400 significantly improved the learning and memory-impairment by restoring oxidative MDA, GSH, SOD, CAT, and DNA damage (8-oxodG) markers of serum, and cortex. It also improved acetyl- and, butyryl-cholinesterase, ß-secretase, and MAPτ level in brain by restoring proinflammatory cytokines IL-6, and TNF-α indicators in neurotoxic rats. GC-MS/MS reported therapeutic significance active compounds were molecular-docked towards target proteins, found that proscillaridin showed the highest affinity towards AChE, BuChE, BACE1, and BACE2 with binding energy of ΔGb -9.1, ΔGb -10.2, ΔGb -11.4 and ΔGb -11.5 Kcal/mol, respectively. Cymarin and morphine-3-glucuronide showed the second highest binding affinity towards AChE (ΔGb -8.8) and BuChE (ΔGb -10.0), respectively. In BACE-1, betulin showed the second highest binding affinity ΔGb -10.7 Kcal/mol and in BACE-2, morphine-3-glucuronide showed the second highest binding affinity ΔGb -9.8 Kcal/mol. CONCLUSIONS: Synergistic impact of proscillaridin, Cymarin, morphine-3-glucuronide, betulin like compounds in HAPT improved memory impairment, healing of tissue architecture of cortex with the restoration of neurochemical, neuroinflammation, and oxidative indicators in neurotoxic rats.

Kuijieling decoction regulates the Treg/Th17 cell balance in ulcerative colitis through the RA/RARα signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Ulcerative colitis (UC) is an idiopathic intestinal disease characterized by chronic inflammation with unknown etiology. Kuijieling decoction is a traditional Chinese Medicine with unique therapeutic efficacy for UC. AIM OF THE STUDY: To validate the effects of Kuijieling decoction on vitamin A metabolism and retinoic acid (RA) production, along with evaluation of its immunomodulatory activity, and further clarify the upstream mechanisms underlying Treg/Th17 regulation that contribute to therapeutic effects against UC. MATERIALS AND METHODS: Network pharmacology and molecular docking analyses were employed to predict the potential anti-UC targets of Kuijieling and associated pathways. A rat model of UC was generated by treatment with trinitrobenzene sulfonic acid (TNBS) to induce inflammation. T lymphocytes were induced to differentiate into Th17 via combined stimulation with the cytokines TGF-ß1, IL-6 and IL-23. Expression levels of RA/RARα-related factors (RARα, CRABPII, Smad3, IL-6R and IL-23R) and Treg/Th17 cell-related factors (Foxp3, RORγt) were measured via western blot (WB), quantitative real-time PCR (RT-qPCR), and immunohistochemistry analyses. Components of the vitamin A metabolic pathway (vitamin A, retinol, retinoic acid) and Treg/Th17 cell-related cytokines (IL-10, IL-17, IL-21) were evaluated using ELISA. Flow cytometry was performed to determine the percentages of Treg and Th17 cells. RESULTS: The action targets of Kuijieling were significantly associated with T cell activation, Th17 cell differentiation and the immune response. IL-2, IL-6, STAT3 and RARα displayed strong binding affinities with the main components of Kuijieling, suggesting an important role in its therapeutic efficacy. Kuijieling promoted vitamin A metabolism and RA/RARα signaling in UC rats and T lymphocytes. Moreover, Kuijieling effectively regulated the Treg/Th17 cell balance in UC rats and T lymphocytes and relieved inflammation. The protective effect of Kuijieling was weakened after inhibition of the RA/RARα signaling pathway. CONCLUSIONS: Kuijieling promotes vitamin A metabolism and RA synthesis, enhances interactions between RA, intracellular binding protein CRABPII and nuclear receptor RARα, and upregulates Smad3 and Foxp3, thus promoting Treg differentiation. Simultaneously, Kuijieling inhibited expression of IL-6R and IL-23R genes and production of RORγt, leading to suppression of Th17 differentiation, and ultimately, reduction of the Th17/Treg cell ratio.

Tetrastigma hemsleyanum suppresses neuroinflammation in febrile seizures rats via regulating PKC-δ/caspase-1 signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Tetrastigma hemsleyanum Diels et Gilg (T. hemsleyanum, Sanyeqing) has been used in the prevention and treatment of repetitive Febrile seizures (FS) over the centuries in China. AIM OF THE STUDY: T. hemsleyanum exerts wide pharmacological action, which has been widely used for treating various diseases, including infantile febrile seizure. However, the systematic study on this herb's material basis and the functional mechanism is lacking. This study intended to systematically elucidate the mechanism of T. hemsleyanum against febrile seizures. MATERIALS AND METHODS: The efficacy of T. hemsleyanum was estimated by using a hot bath as a model of FS, the onset and duration of seizure, morphological structure changes of hippocampal neurons as well as magnetoencephalography were applied to evaluate the effects. Meanwhile, the bioactive components of T. hemsleyanum responsible for the therapeutic effect of T. hemsleyanum on FS were identified by UPLC-MS/MS. Then we systematically elucidated the mechanism of T. hemsleyanum based on metabonomics, transcriptomics, network pharmacological and experimental validation. RESULTS: In a hyperthermia-induced FS model of rats, T. hemsleyanum significantly increased the seizure latency and decreased seizure duration, alleviating the abnormal delta and gamma band activity during epileptic discharge. Furthermore, ten chemical components of ethanol extracts from T. hemsleyanum were identified by UPLC-MS/MS, including quercetin, kaempferol, and procyanidin B1 and so on, which was consistent with the network pharmacology prediction. The serum metabolomics indicated that T. hemsleyanum mainly acts on inflammation regulation and neuroprotection by the glycerophospholipid metabolism pathway. Ninety-two potential targets of T. hemsleyanum on FS were identified by network pharmacology, and TNF, IL-6, and IL-1ß were considered the pivotal targets. In the hippocampus transcriptomics, 17 KEGG pathways were identified after T. hemsleyanum treatment compared with the FS model group, among which 15 pathways overlapped with those identified by network pharmacology, and the PKC-δ/caspase-1 signaling pathway was a critical node. Finally, in vivo experiments also verified T. hemsleyanum inhibited the activation of microglia and resulted in a significant reduction in the level of PKCδ, NLRC4, caspase-1, IL-1ß, IL-6 and TNF-α in hippocampus of FS rats. CONCLUSIONS: Our study suggested that the therapeutic effect of T. hemsleyanum on FS might be regulated by inhibiting the neuroinflammation, thus exerting an anticonvulsant effect in vivo, and the mechanism might be related to regulating the PKC-δ/caspase-1 signaling pathway.

Quality control assessment, toxicity profiling, and experimental validation of network pharmacology-predicted anti-inflammatory potential of Natsiatum herpeticum Buch.-Ham. Ex Arn.

ETHNOPHARMACOLOGICAL RELEVANCE: Natsiatum herpticum Buch.-Ham. Ex Arn., a least-explored plant, is being considered a wild edible plant by the Bankariya community of Nepal and the Mishing, Sonowal Kachari, and several ethnic groups in the north-east region of India. It is also used as a traditional remedy for the treatment of pain and inflammation-associated conditions like cuts and wounds, stomach ache, backache, and headache as a practice of a folkloristic system of medicine. In spite of several previous publications suggesting its use by different tribes, no documentation or scientific approaches have been made hitherto to validate its ethnopharmacological claims. AIM OF STUDY: The study aimed at the botanical quality control assessment, toxicity profiling, and network pharmacology-assisted experimental validation of the anti-inflammatory potential of the aqueous extract of N. herpeticum to fill the lacunae in the current knowledge. MATERIAL AND METHOD: Plant material was authenticated using a classical taxonomical approach and DNA barcoding. The quality control methods, acute toxicity study, and repeated dose 28-day oral toxicity study were performed as per standard guidelines. QToF-MS analysis, drug-likeness properties, network pharmacology-based anti-inflammatory prediction, and in vitro assays were carried out. RESULTS: Quality control assessment was done for the plant. Toxicity studies revealed the aqueous extract to be non-toxic when consumed for short periods at low doses. Alterations in food and water intake, biochemical parameters, and alterations in liver histology (n = 2 female rats) implicate repeated exposure to high doses (2000 mg/kg) that may possess deleterious effects, particularly in hepatic tissues. 21 representative compounds (14 drug-like molecules) were detected by QToF-MS analysis and then subjected to network pharmacology to predict anti-inflammatory effects. It was found that an anti-inflammatory effect may be exerted by modulating inflammatory pathways involving genes such as TNF, PTGS2, EGFR, STAT3, PPARG, PTGER4, PPARA, NOS2, TRPV1, and JAK2. Further, in vitro studies demonstrated plant extract to possess a good anti-inflammatory effect with IC50 values of 98.76, 85.73, and 96.16 µg/ml in protein denaturation, proteinase inhibition, and haemolysis inhibition assays, respectively. CONCLUSION: The plant extract was found to be safer at acute dose but may cause potential liver toxicity on prolonged use. The anti-inflammatory property predicted by network pharmacology was further supported by the positive results of in vitro experiments. In summary, to further establish the toxicity profile of this edible plant and its anti-inflammatory properties, chronic toxicity study and in vivo experiments are required.

Toxicological safety evaluation of zengye granule through acute and 30-day toxicity studies in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Zengye granule (ZYG), a traditional Chinese medicine formula composed of Radix Scrophulariae, Radix Ophiopogonis, and Radix Rehmanniae in the ratio of 1.0:0.8:0.8, is listed in the Chinese Pharmacopoeia for treating diseases associated with yin deficiency, such as inner heat, dry mouth and pharynx, and dry bound stool. However, little information is available on its toxicological safety. AIM OF THE STUDY: To evaluate the acute and subacute toxicity of ZYG after oral administration in rats. MATERIALS AND METHODS: In the acute toxicity study, ZYG was orally administered to rats at a single dose of 10 g/kg/day. In the subacute toxicity study, ZYG was administered orally to rats at repeated daily doses of 2.5, 5.0, or 10 g/kg/day for 30 days. The toxicological effects were evaluated by assessing the rats' general behavior, body weight, food intake, water consumption, blood biochemical and hematological parameters, organ coefficients, and organ histopathology. RESULTS: No obvious adverse reactions were found in the rats in the acute toxicity study, indicating that ZYG was non-toxic. In the subacute toxicity study, ZYG had no toxic effect on the rats at a dose of 2.5 g/kg/day but showed slight toxicity in the kidneys, and spleens of the rats at doses of 5 and 10 g/kg/day. Significant drug toxicity was observed in male and female rats at 5 and 10/kg/day; however, elevated WBCs counts, ALT, and LYMs levels were found in female rats. CONCLUSIONS: The oral administration of ZYG at a dose of less than 10 g/kg/day for 1 day or 2.5 g/kg/day for 30 consecutive days can be considered safe, as these doses showed no distinct toxicity or side effects in the rats in this study. Therefore, the dosage should be set according to the clinically recommended dosage to ensure its safety.

Urtica cannabina L. water extract exhibits anti-inflammatory activity by regulating inflammatory cytokines: In vitro and in vivo evidence.

ETHNOPHARMACOLOGICAL RELEVANCE: Urtica cannabina L. (U. cannabina) is a medicinal plant used in traditional Chinese and Kazakh medicine for treatment of various ailments such as rheumatoid arthritis, rheumatic pain, high blood pressure, and snake bites. However, very few studies have focused on the anti-inflammatory effects of U. cannabina and the mechanisms underlying these effects. AIM OF THE STUDY: This study to investigate the in vitro and in vivo anti-inflammatory effect of U. cannabina, the underlying mechanisms, and its phytochemical profile. MATERIALS AND METHODS: We investigated the anti-inflammatory effects of the U. cannabina water extract on lipopolysaccharide-stimulated RAW264.7 macrophages and paw edema in rats and analyzed its chemical components using ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). RESULTS: U. cannabina water extract effectively inhibited the secretion of multiple inflammatory factors, and its corresponding mRNA expression in LPS-induced RAW264.7 cells (p < 0.05). Tincture of U. cannabina water extract significantly reduced carrageenan-induced rat paw edema and levels of inflammatory factors (p < 0.05). A total of 31 compounds, which mainly include organic acids, were tentatively identified based on the comparison of their mass spectrum profiles with those recorded in a mass spectra database. CONCLUSIONS: The results of this study elucidated the anti-inflammatory effect of U. cannabina water extract in vitro and in vivo and showed that the extract elicits the anti-inflammatory effects by regulating the activity of inflammatory cytokines. The results prove that U. cannabina is a valuable source of active compounds with anti-inflammatory activity.

Qingjie decoction attenuated E.coli-induced diarrhea by regulating energy metabolism and alleviating inflammation based on network analysis and metabolomics.

ETHNOPHARMACOLOGICAL RELEVANCE: Diarrhea is a frequently encountered gastrointestinal complication in clinical practice, and E. coli is one of the main causative agents. Although Qingjie decoction (QJD) has been shown to be highly effective in treating diarrhea by eliminating heat-toxin, the underlying molecular mechanisms and pathways of QJD remain unclear. AIM OF REVIEW: The aim of this research was to explore the effects and fundamental mechanism of QJD on diarrhea induced by E.coli in rats. MATERIALS AND METHODS: Initially, we used UHPLC-MS/MS analysis to identify the chemical composition of QJD. Then, we constructed a visualization network using network pharmacology. Next, we utilized metabolomics to identify differentially expressed metabolites of QJD that are effective in treating diarrhea. RESULTS: The chemical composition of QJD was analyzed using UHPLC-MS/MS, which identified a total of 292 components. Using a network pharmacology approach, 127 bioactive compounds of QJD were screened, targeting 171 potential diarrhea treatment targets. TNF-α, IL-6, IL-1ß, and CAT were identified as important targets through visualizing the PPI network. Enrichment analysis demonstrated significant enrichment in the TNF signaling pathway, IL-17 signaling pathway, and PI3K-Akt signaling pathway. QJD showed beneficial effects, such as increased body weight, decreased fecal water content, and reduced inflammatory cell infiltration in the duodenum and colon, as well as maintaining the structure of the duodenum and colon. Metabolomic analysis revealed 32 differentially expressed metabolites in the control, model and QJD-H groups, including glucose, valine, and cysteine. Functional analysis indicated that differential metabolites were related to energy metabolism, including glucose metabolism, TCA cycle, and amino acid metabolism. CONCLUSION: QJD significantly increased body weight, decreased water content in feces, relieved inflammatory cell infiltration, maintained the structure of duodenum and colon. Combining network analysis and metabolomics, QJD exerted therapeutic effects by inhibiting inflammation and oxidative stress, regulating glucose metabolism, tricarboxylic acid metabolism, and amino acid metabolism.

Sijunzi decoction ameliorates gastric precancerous lesions via regulating oxidative phosphorylation based on proteomics and metabolomics.

ETHNOPHARMACOLOGICAL RELEVANCE: Sijunzi decoction (SJZD), a traditional Chinese medicine formula, is commonly used in clinical practice for the treatment of gastric precancerous lesions (GPL). However, the mechanism of gastric protection is not fully understood. AIMS OF THE STUDY: The purpose of this study was to systematically evaluate the efficacy of SJZD in blocking the development of GPL and to reveal the underlying mechanism. METHODS: First, we established a rat model of GPL, which was induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) combined with an irregular diet and 40% ethanol. The efficacy of SJZD was evaluated based on pathological sections and serum biochemical indices. Then, the pharmacodynamic mechanism of SJZD was revealed by quantitative proteomics based on stable isotope dimethyl labeling. At the same time, the pharmacodynamic mechanism was verified by quantitative metabolomics. In addition, the anti-gastritis effect of SJZD was confirmed by a serum pharmacology method in a cell model, and the functional mechanism was further verified. RESULTS: We demonstrated that SJZD could block the development of GPL in the animal model. Proteomics and metabolomics revealed that SJZD blocks GPL development by regulating oxidative phosphorylation (OXPHOS). In addition, the serum pharmacology results showed that SJZD-containing serum (SJZD-CS) could inhibit apoptosis in MNNG-induced GES-1 cells. OXPHOS inhibitors could significantly reduce the protective effect of SJZD-CS. CONCLUSION: SJZD effectively ameliorates GPL, and proteomics and metabolomics revealed that its protective effects are closely related to OXPHOS.

Sanpian decoction ameliorates cerebral ischemia-reperfusion injury by regulating SIRT1/ERK/HIF-1α pathway through in silico analysis and experimental validation.

ETHNOPHARMACOLOGICAL RELEVANCE: Cerebral ischemia-reperfusion injury (CIRI) is a complex pathophysiological process involving multiple factors, and becomes the footstone of rehabilitation after ischemic stroke. Sanpian decoction (SPD) has exhibited protective effects against CIRI, migraine, and other cerebral vascular diseases. However, the underlying mechanisms have not been completely elucidated. AIM OF THE STUDY: This study sought to explore the potential mechanisms underlying the effect of SPD against CIRI. MATERIALS AND METHODS: High-performance liquid chromatography (HPLC) and ultra-high-performance liquid chromatography (UPLC) were carried out to determine the chemical constituents of SPD. A network pharmacology approach combined with experimental verification was conducted to elucidate SPD's multi-component, multi-target, and multi-pathway mechanisms in CIRI occurrence. The pharmacodynamics of the decoction was evaluated by establishing the rat model of middle cerebral artery occlusion/reperfusion (MCAO/R). In vivo and in vitro experiments were carried out, and the therapeutic effects of SPD were performed using 2,3,5-triphenyltetrazolium chloride (TTC) staining, hematoxylin-eosin (HE) staining, and Nissl staining. We used terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining and flow cytometry to evaluate cortex apoptosis. The quantification of mRNA and corresponding proteins were performed using real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) and Western blot respectively. RESULTS: Our research showed that pretreatment with SPD improved neurological function and inhibited CIRI. Network pharmacology revealed that the hypoxia-inducible factor-1 (HIF-1) signaling pathway and mitogen-activated protein kinase (MAPK) signaling pathway-mediated apoptosis may be associated with CIRI. In vivo and in vitro experiments, we confirmed that SPD increased cerebral blood flow, improved neural function, and reduced neural apoptosis via up-regulating the expression of sirtuin 1 (SIRT1) and down-regulating phospho-extracellular regulated protein kinases (p-ERK)/ERK and HIF-1α levels in CIRI rats. CONCLUSION: Taken together, the present study systematically revealed the potential targets and signaling pathways of SPD in the treatment of CIRI using in silico prediction and verified the therapeutic effects of SPD against CIRI via ameliorating apoptosis by regulating SIRT1/ERK/HIF-1α.