[Anti-depression mechanism of Zuojin Pills:based on UHPLC-TOF-MS, network pharmacology, and experimental verification].

This study aims to explore the anti-depression mechanism of Zuojin Pills based on the plasma constituents, network pharmacology, and experimental verification. UHPLC-TOF-MS was used for qualitative analysis of Zuojin Pills-containing serum. Targets of the plasma constituents and the disease were retrieved from PharmMapper and GeneCards. Then the protein-protein interaction(PPI) network was constructed and core targets were screened for GO term enrichment and KEGG pathway enrichment. Cytoscape 3.7.2 was employed construct the "compound-target-pathway" network and the targets and signaling pathways of Zuojin Pills against depression were predicted. CUMS-induced depression mouse model was established to verify the key targets. The results showed that a total of 21 constituents migrating to blood of Zuojin Pills were identified, which were mainly alkaloids. A total of 155 common targets of the constituents and the disease and 67 core targets were screened out. KEGG enrichment and PPI network analysis showed that Zuojin Pills may play a role in the treatment of depression through AMPK/SIRT1, NLRP3, insulin and other targets and pathways. Furthermore, the results of animal experiments showed that Zuojin Pills could significantly improve the depression behaviors of depression, reduce the levels of IL-1ß, IL-6 and TNF-α in hippocampus and serum, activate AMPK/SIRT1 signaling, and reduce the protein expression of NLRP3. In conclusion, Zuojin Pills may play a role in the treatment of depression by activating AMPK/SIRT1 signaling pathway, and inhibiting NLRP3 activation and neuroinflammation in the hippocampus of mice.

[Mechanism of Jiaotai Pills in treatment of depression based on quantitative proteomics].

This study aimed to investigate the effect of Jiaotai Pills on protein expression in the hippocampus of the rat model of chronic unpredictable mild stress(CUMS)-induced depression by quantitative proteomics and explore the anti-depression mechanism of Jiaotai Pills. The SD rats were randomized into control, model, Jiaotai Pills, and fluoxetine groups(n=8). Other groups except the control group were subjected to CUMS modeling for 4 weeks. After 4 weeks of continuous administration, the changes of behavior and pathological morphology of the hippocampal tissue were observed. Proteins were extracted from the hippocampal tissue, and bioinformatics analysis was performed for the differentially expressed proteins(DEPs) identified by quantitative proteomics. Western blot was employed to verify the key DEPs. The results showed that Jiaotai Pills significantly alleviated the depression behaviors and hippocampal histopathological changes in the rat model of CUMS-induced depression. A total of 5 412 proteins were identified in the hippocampus of rats, including 65 DEPs between the control group and the model group and 35 DEPs between the Jiaotai Pills group and the model group. There were 16 DEPs with the same trend in the Jiaotai Pills group and the control group, which were mainly involved in sphingolipid, AMPK, and dopaminergic synapse signaling pathways. The Western blot results of Ppp2r2b, Cers1, and Ndufv3 in the hippocampus were consistent with the results of proteomics. In conclusion, Jiaotai Pills may play an anti-depression role by modulating the levels of Ppp2r2b, Cers1, Ndufv3 and other proteins and regulating sphingolipid, AMPK, and dopaminergic synapse signaling pathways.

Integrated analysis of comprehensive metabolomics and network pharmacology to reveal the mechanisms of abelmoschus manihot (L.) medik. in the treatment of cisplatin-induced chronic kidney disease.

Background: Abelmoschus manihot (L.) Medik ("Huangkui" in Chinese, HK) has been widely used for the treatment of kidney diseases. Nephrotoxicity is the side effect of cisplatin (CDDP), which greatly limits its clinical application. Therefore, CDDP could be used to establish the chronic kidney disease (CKD) model. However, the protective effects of HK on CDDP-induced CKD have not been investigated. Purpose: To explore the protective effect and underlying mechanisms of HK on multiple low-dose CDDP-induced CKD in rats by the integrated analysis of serum, kidney, and urine metabolomics and network pharmacology. Methods: The CKD model was induced by multiple low-dose CDDP. Body weight, organ index, serum biochemical, and kidney histology were examined to evaluate the effect of HK. Serum, kidney, and urine were collected and profiled by HILIC/RPLC-Q-TOF/MS-based metabolomics. Potential biomarkers (PBs) were screened according to the criteria of VIP >1, p < 0.01, and FC > 2, and then identified or assigned. The pathway analysis and PBs enrichment were conducted by MetaboAnalyst and ChemRICH. Furthermore, network pharmacology was adopted to dig out the active components and targets. Finally, the results from metabolomics and network pharmacology were integrated to confirm each other. Results: HK could recover the CDDP-induced abnormal pharmacological and metabolic profile changes. A total of 187 PBs were screened and identified from the serum, kidney, and urine metabolomics. Pathway analysis showed that multiple metabolic pathways, mainly related to amino acid and lipid metabolisms, were involved in the nephroprotective effect of HK, and especially, HK could significantly alleviate the disorder of tryptophan metabolism pathway in serum, kidney, and urine. Meanwhile, network pharmacology analysis revealed that 5 components in HK and 4 key genes could be responsible for the nephroprotection of HK, which also indicated that the metabolism of tryptophan played an important role in HK against CKD. Conclusion: HK has a nephroprotection on CDDP-induced CKD, mainly by restoring the dysregulation of tryptophan metabolism. Integrated analysis of serum, kidney, and urine metabolomics and network pharmacology was a powerful method for exploring pharmacological mechanisms and screening active components and targets of traditional Chinese medicine.

[Pharmacokinetic interaction of Jiaotai Pills and Fluoxetine in rats with CUMS-induced depression].

A high-performance liquid chromatography-tandem mass spectrometry(LC-MS/MS) was developed for simultaneously determining the components(magnoflorine, jatrorrhizine, berberrubine, coptisine, berberine) of Jiaotai Pills and Fluoxetine in plasma of rats with chronic unpredictable mild stress(CUMS)-induced depression to investigate the pharmacokinetic herb-drug interaction of Jiaotai Pills and Fluoxetine in the rats. The six components showed good linear relationship within the corresponding concentration ranges, and the method showed high specificity, accuracy, precision, and stability. Their pharmacokinetic parameters were calculated by DAS 3.2.2, and the results showed that the in vivo metabolic processes of the six components accorded with the characteristics of non-compartmental model. When Jiaotai Pills and Fluoxetine were used together, the AUC_(0-t), AUC_(0-∞), C_(max), and C_(av) of magnoflorine all significantly increased(P<0.05), while the pharmacokinetic trend of berberrubine was opposite to that of magnoflorine, as manifested by the decrease in AUC_(0-t), AUC_(0-∞), T_(max), C_(max), and C_(av)(P<0.01, P<0.05). The pharmacokinetic characteristics of jatrorrhizine, coptisine, and berberine followed the trend of berberrubine. There was no significant difference in the pharmacokinetic characteristics of Fluoxetine in the single or combination groups. This study suggests that the enhanced antidepressant efficacy of Jiaotai Pills and Fluo-xetine may be attributed to the pharmacokinetic interaction.

ACSL4 promotes colorectal cancer and is a potential therapeutic target of emodin.

BACKGROUND: Colorectal cancer (CRC) is an important death-related disease in the world and new therapeutic strategies are urgently needed to reduce mortality. Several studies have demonstrated that emodin, the main ingredient of Rheum palmatum, fights cancer but its potential anti-tumor effect on CRC is still unknown. PURPOSE: The present study is aimed to explore the potential anti-tumor effects of emodin against CRC and the underlying molecular mechanism. METHODS: CRC-related datasets were screened according to filter criteria in the GEO database and TCGA database. By using screened differentially expressed genes, GO, KEGG and survival analysis were carried out. The expressions of ACSL4, VEGFR1, and VEGFR2 were examined by immunohistochemistry and western blot. Then, pcDNA-ACSL4, pcDNA-VEGFR1, and pcDNA-VEGFR2 were used to overexpress ACSL4, VEGFR1, and VEGFR2, while ACSL4 siRNA was used to silence ACSL4 expression in HCT116 cells. CCK-8 assay and transwell migration assay were used to detect the cell proliferation and invasion. A docking simulation assay and an MST assay were performed to explore the potential mode of emodin binding to ACSL4. The HCT116 cells and CRC mouse model were established to investigate the effects of emodin on CRC. RESULTS: The ACSL4, VEGFR1, and VEGFR2 expression were upregulated in CRC tissues and ACSL4 was associated with a shorter survival time in CRC patients. ACSL4 downregulation reduced cell proliferation and invasion, while ACSL4 exhibited a positive correlation with the levels of VEGFR1, VEGFR2, and VEGF. In HCT116 cells, emodin reduced cell proliferation and invasion by inhibiting ACSL4, VEGFR1, and VEGFR2 expression and VEGF secretion. Docking simulation and MST assay confirmed that emodin can directly bind to ACSL4 target. Moreover, ACSL4 overexpression abolished the inhibitory effect of emodin on VEGF secretion and VEGFR1 and VEGFR2 expression, but VEGFR1 and VEGFR2 overexpression did not affect the inhibitory effect of emodin on ACSL4 expression and VEGF secretion. Furthermore, emodin reduced the mortality and tumorigenesis of CRC mice and reduced ACSL4, VEGFR1, VEGFR2 expression, and VEGF content. CONCLUSION: Our findings indicate that emodin inhibits proliferation and invasion of CRC cells and reduces VEGF secretion and VEGFR1 and VEGFR2 expression by inhibiting ACSL4. This emodin-induced pathway offers insights into the molecular mechanism of its antitumor effect and provides a potential therapeutic strategy for CRC.

Integrated Network Pharmacology and GC-MS-Based Metabolomics to Investigate the Effect of Xiang-Su Volatile Oil Against Menopausal Depression.

Menopausal depression perplexes a great number of women in later life. Xiangfu-Zisu (Xiang-Su), a traditional Chinese herbal pair composed of rhizomes of Cyperus rotundus L. (Xiangfu) and leaves of Perilla frutescens (L.) Britt. (Zisu), is frequently reported with antidepressant-like effects. The volatile oil from Xiangfu and Zisu has shown good antidepressant action, but its mechanism is still unclear. This study aimed to investigate the pharmacological mechanism of Xiang-Su (XS) volatile oil against menopausal depression through gas chromatography-mass spectrometry (GC-MS)-based network pharmacology and metabolomics. First, ADME screening was performed on actual detected components of XS volatile oil to obtain active constituents, and then duplicates of active constituent-related targets and menopausal depression-related targets were collected. These duplicates were considered as targets for XS volatile oil against menopausal depression, followed by GO and KEGG enrichment analyses. It showed that a total of 64 compounds were identified in XS volatile oil, and 38 active compounds were screened out. 42 overlapping genes between 144 compound-related genes and 780 menopausal depression-related genes were obtained. Results showed that targets of SLC6A4 and SLC6A3, regulation of serotonergic and dopaminergic synapses, were involved in the antidepressant mechanism of XS volatile oil. Next, antidepressant-like effect of XS volatile oil was validated in menopausal rats by ovariectomy (OVX) combined with chronic unpredictable mild stress (CUMS). Behavioral tests, biochemical analysis, and GC-MS-based non-targeted plasma metabolomics were employed to validate the antidepressant effect of XS volatile oil. Experimental evidence demonstrated that XS volatile oil reversed behavioral parameters in the sucrose preference test (SPT), open-field test (OFT), forced swim test (FST), and serum estradiol levels in OVX rats. Furthermore, results of metabolomics indicated that XS volatile oil mainly acts on regulating metabolic pathways of phenylalanine, tyrosine and tryptophan biosynthesis, tyrosine metabolism, and tryptophan metabolism, which were corresponding with the above-predicted results. These data suggest that network pharmacology combined with metabolomics provides deep insight into the antidepressant effect of XS volatile oil, which includes regulating key targets like SLC6A4 and SLC6A3, and pathways of serotonergic and dopaminergic synapses.

Integrated Network Pharmacology Analysis and Experimental Validation to Investigate the Mechanism of Zhi-Zi-Hou-Po Decoction in Depression.

Zhi-Zi-Hou-Po Decoction (ZZHPD) is a well-known traditional Chinese medicine (TCM) that has been widely used in depression. However, the antidepressant mechanism of ZZHPD has not yet been fully elucidated. The purpose of this study was to explore the pharmacological mechanisms of ZZHPD acting on depression by combining ultra flow liquid chromatography with quadrupole time-of-flight mass spectrometry (UFLC-Q-TOF/MS) and network pharmacology strategy. The chemical components of ZZHPD were identified using UFLC-Q-TOF/MS, while the potential drug targets and depression-related targets were collected from databases on the basis of the identified compounds of ZZHPD. Protein-protein interaction (PPI) network, gene ontology (GO), and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analyses were used to unravel potential antidepressant mechanisms. The predicted antidepressant targets from the pharmacology-based analysis were further verified in vivo. As a result, a total of 31 chemical compounds were identified by UFLC-Q-TOF/MS; 514 promising drug targets were mined by using the Swiss Target Prediction; and 527 depression-related target genes were pinpointed by the GeneCards and OMIM databases. STRING database and Cytoscape's topological analysis revealed 80 potential targets related to the antidepressant mechanism of ZZHPD. The KEGG pathway analysis revealed that the antidepressant targets of ZZHPD were mainly involved in dopaminergic synapse, serotonin synapse, cAMP, and mTOR signaling pathways. Furthermore, based on the animal model of depression induced by chronic corticosterone, the regulatory effects of ZZHPD on the expression of MAOA, MAOB, DRD2, CREBBP, AKT1, MAPK1, HTR1A, and GRIN2B mRNA levels as well as the cAMP signaling pathway and monoaminergic metabolism were experimentally verified in rats. Our study revealed that ZZHPD is expounded to target various genes and pathways to perform its antidepressant effect.

[Comparison of in vivo pharmacokinetics of five constituents in Zhishi Zhizi Chi Decoction in normal rats and CUMS-induced depressive rats].

A LC-MS/MS method was developed for the rapid and simultaneous determination of genipin-1-ß-D-gentiobioside,geniposide,naringin,hesperidin and neohesperidin in SD rat plasma.The linear relationships of these five constituents in rats were validated,and the specificity,accuracy,precision and stability met the requirements.Their pharmacokinetic parameters were calculated by DAS 3.2.2,and the results showed that the metabolic process in vivo of the five constituents accorded with the characteristics of noncompartmental model.Their main pharmacokinetic parameters were listed as follows:(1) genipin-1-ß-D-gentiobioside:t_(1/2)(3.20±0.51)h,C_(max)(403.15±96.93)µg·L~(-1)and AUC_(0-t)(612.56±148.50)µg·L~(-1)·h for the model group,while t_(1/2)(3.07±0.75) h,C_(max)(229.50±60.63)µg·L~(-1)and AUC_(0-t)(413.14±76.37)µg·L~(-1)·h for the normal group;(2) geniposide:t_(1/2)(3.24±0.68) h,C_(max)(2 961.40±688.02)µg·L~(-1),and AUC_(0-t)(10 972.87±1 992.96)µg·L~(-1)·h for the model group,while t_(1/2)(4.56±0.96) h,C_(max)(1 833.27±558.13)µg·L~(-1),and AUC_(0-t)(8 996.27±3 053.48)µg·L~(-1)·h for the normal group;(3) naringin:t_(1/2)(1.64±0.59) h,C_(max)(415.13±259.54)µg·L~(-1),and AUC_(0-t)(608.62±289.05)µg·L~(-1)·h for the model group,while t_(1/2)(1.02±0.25) h,C_(max)(355.08±180.00)µg·L~(-1),and AUC_(0-t)(501.07±242.68)µg·L~(-1)·h for the normal group;(4) hesperidin:t_(1/2)(0.86±0.29) h,C_(max)(95.17±22.80)µg·L~(-1)and AUC_(0-t)(141.19±54.63)µg·L~(-1)·h for the model group,while t_(1/2)(0.95±0.31) h,C_(max)(46.48±18.33)µg·L~(-1)and AUC_(0-t)(69.51±14.73)µg·L~(-1)·h for the normal group;(5) neohesperidin:t_(1/2)(0.89±0.29) h,C_(max)(828.78±361.56)µg·L~(-1)and AUC_(0-t)(1 292.29±553.73)µg·L~(-1)·h for the model group,while t_(1/2)(0.90±0.31) h,C_(max)(314.68±172.45)µg·L~(-1)and AUC_(0-t)(385.99±138.55)µg·L~(-1)·h for the normal group.

[Effects of Jiaotai Pills on CUMS-induced depression model in mice based on changes of SIRT1 expression in hippocampus].

The present study investigated the effects and mechanisms of Jiaotai Pills on depressed mice induced by chronic unpredictable mild stress(CUMS). The CUMS-induced depression model mice were established and the depression behaviors of mice were evaluated by sucrose preference test, open field test, tail suspension test, and forced swimming test. Molecular docking was employed to simulate the interaction of six main active ingredients in Jiaotai Pills with SIRT1. Immunohistochemical staining was used to detect the level of SIRT1 in the hippocampus of mice. Western blot was used to detect the protein expression levels of SIRT1, p-NF-κB p65, NF-κB p65, and FoxO1 in the hippocampus of mice. Enzyme-linked immunosorbent assay(ELISA) kits were used to detect the levels of interleukin(IL)-1ß, IL-6, tumor necrosis factor-α(TNF-α), and brain-derived neurotrophic factor(BDNF) in the hippocampus and serum of mice. Biochemical kits were used to detect superoxide dismutase(SOD) activity and malondialdehyde(MDA) and glutathione(GSH) levels in the hippocampus and serum of mice. Liquid chromatography-tandem mass spectrometry(LC-MS/MS) was used to detect the levels of dopamine(DA), 5-hydroxytryptamine(5-HT), and norepinephrine(NE) in the hippocampus and serum of mice. The results showed that the sucrose preference rate, movement distance, and the number of crossing centers were reduced in the model group(P<0.01), and the tail suspension time and swimming immobility time were increased(P<0.01). Molecular docking results indicated good binding of six main active ingredients in Jiaotai Pills to SIRT1. In the hippocampus, the expression level of SIRT1 was reduced(P<0.01), and the levels of p-NF-κB p65/NF-κB p65 and FoxO1 were increased(P<0.01). In the hippocampus and serum, the levels of IL-1ß, IL-6, TNF-α, and MDA were increased(P<0.01), and the activity of SOD and the levels of GSH, DA, 5-HT, NE, and BDNF were reduced(P<0.01). The treatment with high-dose Jiaotai Pills increased the sucrose preference rate, movement distance, and the number of crossing centers(P<0.05), reduced tail suspension time and swimming immobility time(P<0.01), elevated hippocampal SIRT1 expression level(P<0.01), decreased hippocampal and serum IL-1ß, IL-6, TNF-α, and MDA levels(P<0.01), potentiated SOD activity, and up-regulated GSH, DA, 5-HT, NE, and BDNF levels in the hippocampus and serum(P<0.05, P<0.01) in model mice. In conclusion, the results showed that Jiaotai Pills could improve the depression behaviors of model mice with CUMS-induced depression, and the underlying mechanism was related to the up-regulation of SIRT1 in the hippocampus of mice to exert anti-inflammatory and anti-oxidative stress effects.

[A systematic pharmacological investigation of pharmacologically active ingredients in Toujie Quwen granules for treatment of COVID-19].

OBJECTIVE: To explore the pharmacologically active ingredients in Toujie Quwen granules (TJQW) for treatment of coronavirus disease 2019 (COVID-19) in light of systemic pharmacology. METHODS: We performed database search, literature mining and drug-like index screening to identify the bioactive components in TJQW, the positive drugs for disease treatment and their therapeutic targets. The core disease target was investigated based on the cross-linking interaction of the bioactive components, positive drug and potential disease target, and the target proteins at the key nodes were analyzed by GO and KEGG analyses. Based on the therapeutic targets for COVID-19, virtual screening was conducted to screen the compounds in TJQW and construct the network cross-linking the key bioactive molecules in TJQW, key node targets of the disease, and the related biological pathways. RESULTS: We identified 159 compounds in TJQW and obtained 18 core proteins based on the cross-linking of the bioactive components, positive drugs and disease targets. The key node targets consisted of 22 targets including the latest 4 COVID-19 proteins. Virtual screening results showed that at least 14 compounds could bind with the core disease target proteins. The material basis of TJQW for COVID-19 treatment was explained in multi-pathway, multi-component and multi-target perspectives. In terms of the structural characteristics of the compounds, we screened the top 30 molecules with strong binding with the target proteins, among which flavonoids were the predominant components. CONCLUSIONS: This investigation reveals the therapeutic mechanism of TJQW for COVID-19 involving multiple components, targets and pathways from the perspective of key bioactive molecules, disease key node targets and related biological pathways. We screened 30 active precursors from TJQW, which provides reference for the clinical application and further development of TJQW.

Mahuang-Fuzi-Xixin Decoction Reverses Depression-Like Behavior in LPS-Induced Mice by Regulating NLRP3 Inflammasome and Neurogenesis.

Evidence suggests that inflammation and neurogenesis play an important role in major depressive disorder (MDD). Mahuang-Fuzi-Xixin decoction (MFX), as the traditional Chinese prescription, has been widely applied for asthma, migraine, and MDD in clinics. However, the effects of MFX on the potential mechanism in MDD are still unclear. Hence, the present study is aimed at exploring whether the antidepressive effect of MFX is connected to the anti-inflammatory and promoting neurogenesis. Besides, lipopolysaccharide (LPS) of Gram-negative bacteria can induce depressive-like behaviors. We demonstrated that administration of MFX corrected the depressive-like behaviors in LPS-induced mice and significantly decreased the expression of IL-1ß in the hippocampus. LPS injection induced a significant increase in the levels of NLRP3, cleaved caspase-1 p20, and ASC in the hippocampus, as well as Trx-interacting protein (TXNIP), and MFX could reverse this change. What is more, treatment of MFX increased the level of doublecortin (DCX), brain-derived neurotrophic factor (BDNF), and tropomyosin-related kinase receptor B (TrkB) in the hippocampus which means that MFX could promote the neurogenesis. In conclusion, the study indicates that MFX relieves a depressive-like state in LPS-induced mice through the inhibition of the NLRP3 inflammasome and the enhancement of the neurogenesis pathway.

Comparative pharmacokinetic study of four major bioactive components after oral administration of Zhi-Zi-Hou-Po decoction in normal and corticosterone-induced depressive rats.

A highly selective and efficient LC-MS/MS method was developed to determine the plasma concentration of magnolol, hesperidin, neohesperidin and geniposide following oral administration of Zhi-Zi-Hou-Po decoction in normal and depressed rats. Plasma samples were pretreated by protein precipitation with methanol. Chromatographic separation was performed on an XTerra® MS C18 column using a gradient elution with a mobile phase composed of acetonitrile-0.1% aqueous formic acid. The proposed method was validated to be specific, accurate and precise for the analytes determination in plasma samples. The calibration curves displayed good linearity over definite concentration ranges for the analytes. The intra- and inter-day precision of the proposed method at three different levels were all within <11.13% and the relative errors ranged from -8.46 to 8.93%. The recovery of the four compounds ranged from 82.72 to 89.08% and no apparent matrix effect was observed during sample analysis. After full validation, the established method was successfully applied for comparing the pharmacokinetics of four components between normal and depressed rats. The results showed that the AUC and Cmax of four analytes in depressed rats were significantly different from those in normal rats and might provide helpful information to guide the clinical use of Zhi-Zi-Hou-Po to treat depression.

Ginsenoside Rb2 inhibits epithelial-mesenchymal transition of colorectal cancer cells by suppressing TGF-ß/Smad signaling.

BACKGROUND: Treating colorectal cancer (CRC) continues to be a clinical challenge. Studies have shown that epithelial-mesenchymal transition (EMT) is a critical step in tumor progression and transforming growth factor-ß1 (TGF-ß1) signaling has been shown to play a crucial role in EMT. Here, we investigate the inhibition effect of Ginsenoside Rb2, main bioactive component of ginseng, in human colorectal cancer cells via TGF-ß1. PURPOSE: The current study aims to study the inhibitory effect of Ginsenoside Rb2 on HCT116 and SW620 cells and its anti-tumor mechanism. METHODS: Histomorphological analysis and western blot analysis were performed to evaluate expression of TGF-ß1 in human cancerous colon samples and the adjacent normal samples. The docking simulation assay were performed to explore the potential mode of binding of Ginsenoside Rb2 to the TGF-ß1 protein. CCK8, adhesion and invasion assay were used to assess the effects of Ginsenoside Rb2 in HCT116 and SW620 cells. RT-PCR, Western blot and Immunohistochemical staining were employed to detect the TGF-ß1-related signaling pathways in the colon cancer cells and/or xenograft mice. RESULTS: The expression of TGF-ß1 in human cancerous colon samples was significantly increased compared with the adjacent normal samples. Ginsenoside Rb2 inhibit the growth, adhesion, EMT and metastasis of human colorectal cancer cells. The docking simulation assay confirmed that Ginsenoside Rb2 bound to the hydrophobic pocket of TGF-ß1, which partially overlaps with the binding sites on TGF-ß1, and thus disrupted TGF-ß1 dimerization. Western Blot analysis further confirmed that Ginsenoside Rb2 could inhibit the expression of TGF-ß1 in vitro and in vivo. Furthermore, Ginsenoside Rb2 could inhibit the expression of Smad4 and phosphorylated Smad2/3. CONCLUSION: Ginsenoside Rb2 could inhibit EMT of colorectal cancer cells through the TGF-ß1/Smad signaling, and might be a potential candidate for the treatment of colorectal cancer.

Si-Ni-San Prevents Reserpine-Induced Depression by Inhibiting Inflammation and Regulating CYP450 Enzymatic Activity.

Depression is becoming a major public health concern worldwide. Si-Ni-San (SNS) is a famous formula in Traditional Chinese Medicine (TCM) with potent antidepressant effects. However, the antidepressant mechanism of SNS has not been clearly elucidated. This study was performed to verify whether the antidepressant effects of SNS were related to its anti-inflammatory effects, the levels of brain-derived neurotrophic factor (BDNF) and Cytochrome P450 (CYP450) enzymatic activity. In our study, behavioral tests such as the forced swim test, sucrose preference test and open-field test were evaluated to ensure the establishment of depressive rats. The levels of IL-1ß, IL-6, and TNF-α in the serum, liver, and hippocampus of rats were measured by enzyme-linked immunosorbent assays (ELISA). Furthermore, the key proteins NF-κB, BDNF, and TrkB were analyzed by Western blot (WB) analysis in the hippocampus. In addition, CYP450 enzymatic activity analysis was performed using LC-MS/MS in conjunction with drug and statistics (DAS 3.0) after oral administration of six probe drugs. Our results showed that SNS attenuated reserpine-induced increases in IL-1ß, IL-6, and TNF-α expression in the serum, liver, and hippocampus. The levels of NF-κB, BDNF, and TrkB in the hippocampus of depressive rats were also altered. According to the pharmacokinetic parameters, SNS had moderate inhibitory effects in the reserpine-induced depression model on CYP1A2, CYP2D1, CYP2E1, and CYP3A2, but no significant metabolic changes to CYP2C6 and CYP2D2. These findings suggested that SNS has a protective effect on reserpine-induced depressive rats, which may be related to the improvement of the inflammatory factors, the level of BDNF and the activity of CYP450 enzymes.

Comparison of the Pharmacokinetic Profiles of Ceftriaxone Used Alone and Combined with Danhong Injection in Old Rats.

BACKGROUND AND OBJECTIVES: Danhong injection is the most commonly prescribed adjuvant drug applied for the treatment of cardiovascular and cerebrovascular diseases in China. Ceftriaxone is usually prescribed along with Danhong injection to elderly patients with complications. However, the pharmacokinetic interactions between these two medications have not been investigated. The aim of this study was to investigate whether Danhong injection influences the pharmacokinetic profile of ceftriaxone in old rats when these two medications are used in combination. METHODS: The animal experiment protocol was designed according to the clinical data. Ten-month-old male Sprague-Dawley (SD) rats were dosed with ceftriaxone through intravenous administration for 1 or 7 days in the presence or absence of Danhong injection. The combinations were divided into 1-day, 7-day, and 14-day combined-treatment groups in which Danhong injection was administered for 1, 7, or 14 days and ceftriaxone was given for 1, 7, or 7 days, respectively. The plasma concentration of ceftriaxone was determined by ultrahigh performance liquid chromatography coupled with triple-quadrupole mass spectrometry (UHPLC-TQ-MS) on a BEH C18 column with a mobile phase consisting of acetonitrile and 0.4% formic acid-water. The chromatographic method was validated and found to be simple, rapid, and stable. RESULTS: Danhong injection significantly increased the plasma clearance of and decreased systemic exposure to ceftriaxone. In the 1-day combined-treatment group, the plasma clearance of ceftriaxone increased by 52.69%, and the area under the concentration-time curve (AUC) of ceftriaxone was decreased by 32.54% (P < 0.01). In the 7-day combined-treatment group, the rate of plasma clearance increased by 52.49% and the area under the concentration-time curve decreased by 31.15% (P < 0.01). For the 14-day combined-treatment group, the plasma clearance of ceftriaxone increased by 26.73%, and the area under the concentration-time curve decreased by 21.44% (P < 0.05). CONCLUSIONS: In old male rats, systemic exposure to ceftriaxone decreased when used concomitantly with Danhong injection, which may be because Danhong injection increased the plasma clearance of ceftriaxone. Further investigations should be carried out to clarify the mechanism for the influence of Danhong injection on the pharmacokinetics of ceftriaxone.

Comparative pharmacokinetics of three alkaloids in normal and acute hepatitis rats after oral administration of Yanhuanglian total alkaloids extract.

Acute hepatitis is a severe inflammatory disease with high mortality rates. Dehydrocavidine (DC), berberine (BB) and palmatine (PT), the bioactive components of Yanhuanglian total alkaloids extract (YTAE), may play important roles in its protective effect against acute hepatitis. The present study was designed to compare the pharmacokinetic properties of DC, BB and PT after oral administration of YTAE in normal and acute hepatitis rats. The plasma pharmacokinetic profiles of three major bioactive alkaloids from YTAE were analyzed using an LC-MS/MS method. It was found that the pharmacokinetic parameters of DC, BB and PT represented a statistically significant difference (p ˂ 0.05) between the normal rats and the acute hepatitis rats after administration of YTAE. The results indicated that the Cmax , T1/2ß and AUC of DC, BB and PT in acute hepatitis rats were significantly increased, whereas the CL and Vd values were remarkably decreased (p ˂ 0.05) over those of the normal rats. The results suggested that the pharmacokinetic behavior of the active constituents from YTAE could be changed when it was used in acute hepatitis animals, which provide a meaningful basis for developing a clinical dosage regimen in the treatment of acute hepatitis by YTAE.

Inhibitory Effects of Danshen components on CYP2C8 and CYP2J2.

The use of Chinese herbal medicines and natural products has become increasingly popular in both China and Western societies as an alternative medicine for the treatment of diseases or as a health supplement. Danshen, the dried root of Salvia miltiorrhiza (Fam.Labiatae), which is rich in phenolic acids and tanshinones, is a widely used herbal medicine for the treatment of cardio-cerebrovascular diseases. The goal of this study was to examine the inhibitory effects of fifteen components derived from Danshen on CYP2C8 and CYP2J2, which are expressed both in human liver and cardiovascular systems. Recombinant CYP2C8 and CYP2J2 were used, and the mechanism, kinetics, and type of inhibition were determined. Taxol 6-hydroxylation and astemizole O-desmethyastemizole were determined as probe activities for CYP2C8 and CYP2J2, respectively. Metabolites formations were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The results demonstrated that salvianolic acid A was a competitive inhibitor of CYP2C8 (Ki = 2.5 µM) and mixed-type inhibitor of CYP2J2 (Ki = 7.44 µM). Salvianolic acid C had moderate noncompetitive and mixed-type inhibitions on CYP2C8 (Ki = 4.82 µM) and CYP2J2 (Ki = 5.75 µM), respectively. Tanshinone IIA was a moderate competitive inhibitor of CYP2C8 (Ki = 1.18 µM). Dihydrotanshinone I had moderate noncompetitive inhibition on CYP2J2 (Ki = 6.59 µM), but mechanism-based inhibition on CYP2C8 (KI = 0.43 µM, kinact = 0.097 min-1). Tanshinone I was a moderate competitive inhibitor of CYP2C8 (Ki = 4.20 µM). These findings suggested that Danshen preparations appear not likely to pose a significant risk of drug interactions mediated by CYP2C8 after oral administration; but their inhibitory effects on intestinal CYP2J2 mediated drug metabolism should not be neglected when they are given orally in combination with other drugs. Additionally, this study provided novel insights into the underling pharmacological mechanisms of Danshen components from the perspective of CYP2C8 and CYP2J2 inhibition.

Antidepressant effects of magnolol in a mouse model of depression induced by chronic corticosterone injection.

Evidence showed that the stress hormone corticosterone (CORT) injection resulted in dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis implicated in major depressive disorder. Magnolol, main constituent identified in the barks of Magnolia officinalis, exerted antidepressant effects in a rat model of depression induced by chronic unpredictable mild stress in previous studies. However, its antidepressant-like effects and mechanisms have never been studied in depression model induced by CORT administration in rodents. This study aimed to investigate the antidepressant-like effects and possible mechanisms of magnolol in CORT-treated mice by utilizing a combination of behavioral and biochemical analysis. The depressive model was developed by subcutaneous injection of CORT for 21 days at a dose of 20 mg/kg. CORT administration formed depressive-like behaviors in mice, as indicated by increased immobility time in the forced swim test (FST) and tail suspension test (TST), as well as decreased sucrose intake in sucrose preference test (SPT). Moreover, we also found that CORT levels in serum were significantly increased, along with the decrease of brain-derived neurotrophic factor (BDNF) mRNA, BDNF protein, 5-hydroxytryptamine (5-HT) and norepinephrine (NE) levels in the hippocampus. Treatment with magnolol alleviated depressive-like behaviors, reduced the levels of CORT, and improved the levels of BDNF protein, 5-HT, and NE compared with those in CORT-treated mice. These findings indicated that magnolol possessed antidepressant effects in mice exposed to CORT, which might be partially related to modulate HPA axis, up-regulate BDNF expression and increase neurotransmitters levels in the hippocampus.

[Screen astragalosides from Huangqi injections by LC-TOF-MS-based mass defect filtering approach].

The samples of Huangqi injection (HI) were analyzed by liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (LC-TOF-MS), and both positive and negative ion modes were employed to obtain the LC-TOF-MS analysis information of chemical compounds in HI. Then the mass defect filtering (MDF) approach, which was developed based on the previously published articles, was utilized to rapidly screen the astragalosides from the obtained LC-TOF-MS data. Each screened astragaloside was confirmed by the presence of no less than 2 quasi-molecular ions. All the screened astragalosides were then tentatively assigned according to the parent ion and daughter ion information. Finally, a total of 62 astragalosides were screened and characterized from the HI samples, including 15 new detected ones. The identification results indicated that acetylation, hydrogenation, dehydrogenation, methoxylation and hydration might be the major conversion reactions involved in the formation of the astragalosides. The LC-TOF-MS-based MDF approach was proved to be a feasible and efficient tool to screen the chemical constituents in complex matrices such as herbal medicines.

Urinary metabolomics reveals the therapeutic effect of HuangQi Injections in cisplatin-induced nephrotoxic rats.

The side effects of cisplatin (CDDP), notably nephrotoxicity, greatly limited its use in clinical chemotherapy. HuangQi Injections (HI), a commonly used preparation of the well-known Chinese herbal medicine Astragali radix, appeared to be promising treatment for nephrotoxicity without compromising the anti-tumor activity of CDDP. In this study, the urinary metabolomics approach using liquid chromatography time of flight mass spectrometry (LC-TOF/MS) was developed to assess the toxicity-attenuation effects and corresponding mechanisms of HI on CDDP-exposed rats. As a result, successive administration of HI significantly recovered the decline of body weight and downregulated the abnormal increase of serum creatinine and urea. HI partly restored the CDDP-induced alteration of metabolic profiling back into normal condition. Totally 43 toxicity-attenuation potential biomarkers were screened and tentatively identified, which were involved in important metabolic pathways such as amino acid metabolism, TCA cycle, fatty acid metabolism, vitamin B6 metabolism and purine metabolism. The results clearly revealed that HI could alleviate CDDP-induced nephrotoxicity and improve the disturbed metabolic balance induced by repeated CDDP exposure. The present study provided reliable evidence for the protective effect of HI on CDDP-induced toxicity with the multi-target pharmacological characteristics.