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Aim To predict the targets of Modified Danggui Shaoyao San ( MDSS) in the treatment of chronic atrophic gastritis ( CAG) based on network pharmacology and vertify the results based on experim-ention. Methods TCMSP, SWISS TARGETS, GENE CARDS and OMIM databases were used to screen the therapeutic targets of MDSS for CAG. STRING database and Cytoscape software were used to construct the protein interaction network and screen the core targets. Metascape database was used for GO analysis and KEGG enrichment pathways. And molecular docking was used for target validation. CAG rat model was pre¬pared by N-methyl-N'-nitroso-N-nitroguanidine free drink combined with sodium salicylate gastric lavage. The pathology of rat gastric mucosa was observed by hematoxylin-eosin staining,and the ultrastructure of ep¬ithelial cells was observed by transmission electron mi-croscopy. The serum IL-6 and IL-10 content was detected by enzyme-linked immunosorbent assay, and the expression of JAK2, STAT3 , p-STAT3 , c-MYC mRNA and protein in rats was detected by qPCR and Western blot. Results MDSS acted on 189 targets, mainly involved in response to oxidative stress and apoptotic signaling pathway. KEGG analysis related to pathways in cancer and JAK-STAT signaling pathway. The experimental results showed that the MDSS could improve the degree of atrophy of gastric mucosa in CAG rats and improve the status of epithelial cells, down-regulate the serum IL-6 content of CAG rats, up-regulate the IL-10 content, and reduce the expression of JAK2, STAT3 , p-STAT3 , c-MYC mRNA and protein in gastric mucosa with statistical significance. Conclusions MDSS treats CAG through multiple active ingredients, targets, and pathways, the mechanism of which may be related to the inhibition of the JAK2/STAT3 signaling pathway.
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Objective:To investigate the effects of Danggui Shaoyao San(DSS) on cognitive function and neuronal apoptosis in vascular dementia (VD) rats.Methods:Fifty SPF grade male SD rats aged 6-7 weeks were randomly divided into sham operation group, model group, positive drug group (nimodipine group, 9.45 mg·kg -1), DSS low-dose group (1.6 g·kg -1), DSS high-dose group (6.4 g·kg -1) according to random number table, with 10 rats in each group. The VD rat model was established by permanent ligation of bilateral common carotid arteries. Seven days after modeling, the rats in different groups were administrated by gavage according to corresponding interventions, once a day, for 28 days. Morris water maze test was used to evaluate the learning and memory ability of rats.The levels of malondialdehyde (MDA), superoxide dismutase (SOD) and reactive oxygen species (ROS) in hippocampal area of rat brain were detected by ELISA.The protein expressions of apoptosis-related proteins Bcl-2, Bax, cleaved Caspase-3 and leptin receptor/glycogen synthase kinase 3β microtubule-associated protein tau(LEP-R/GSK-3β/tau) signaling pathway were detected by Western blot. GraphPad Prism 9 software was used for statistical analysis of data, repeated measure ANOVA and one-way ANOVA were used for comparison between multiple groups, and SNK- q test was used for further pairwise comparison. Results:The results of water maze experiment showed that the time and group interaction of escape latency of the five groups were not significant ( F=1.223, P>0.05), the main effect of group and time were significant ( F=74.65, 18.32, both P<0.05). On the 5th day, the escape latency of nimodipine group, DSS low-dose group and DSS high-dose group were lower than that of model group ( q=14.425, 7.477, 21.392, all P<0.05), and that of DSS high-dose group was lower than that of nimodipine group ((15.28±2.46)s, (22.78±3.31)s, q=6.966, P<0.05). There was statistically significant difference in the number of crossing platforms of rats in 5 groups ( F=17.331, P<0.05). The numbers of platform crossing in nimodipine group and DSS high-dose group were higher than that in model group ( q=6.789, 10.635, 5.270, all P<0.05), and the number of platform crossing in DSS high-dose group was higher than that in nimodipine group ((6.84±1.63), (5.22±1.75), q=3.846, P<0.05). ELISA results showed that the levels of MDA, ROS and SOD in hippocampal tissues of rats in 5 groups were significantly different ( F=49.338, 38.518, 15.440, all P<0.05). The levels of MDA and ROS in hippocampus of DSS high-dose group were lower than those of model group ( q=16.061, 13.541, both P<0.05) and nimodipine group ( q=4.317, 5.162, both P<0.05), SOD level of DSS high-dose group was higher than those of model group ( q=8.179, P<0.05) and nimodipine group ( q=4.135, P<0.05). Western blot results showed that the levels of apoptosis-related proteins Bcl-2/Bax and Caspase-3 were significantly different in the 5 groups ( F=30.692, 43.384, both P<0.01). The level of Bcl-2/Bax in DSS high-dose group was higher than that in model group ( q=10.562, P<0.05) and nimodipine group ( q=3.820, P<0.05), the level of Caspase-3 was lower than those of model group ( q=12.139, P<0.05) and nimodipine group ( q=7.734, P<0.05). The levels of LEP-R, p-GSK-3β, p-S404 tau and p-S202 tau expression level in hippocampal tissues of the 5 group were significantly different ( F=80.927, 59.230, 159.784, 105.923, all P<0.01). The levels of LEP-R and p-GSK-3β protein in nimodpine group and DSS high-dose group were higher than those in model group ( q=16.275, 20.104, both P<0.05; q=12.942, 17.257, both P<0.05), the levels of p-S404 Tau and p-S202 Tau in the two groups were lower than those in model group ( q=19.121, 27.456, both P<0.05; q=17.559, 22.780, both P<0.05). The levels of LEP-R(0.98±0.15), (0.86±0.14)) and p-GSK-3β((0.95±0.16)s, (0.82±0.13)) in DSS high-dose group were higher than those in nimodipine group ( q=3.829, 4.314, both P<0.05), the levels of p-S404 Tau((0.41±0.03)s, (0.58±0.07)) and p-S202 Tau((0.48±0.05)s, (0.59±0.06)) in DSS high-dose group were lower than those of nimodipine group ( q=8.335, 5.220, both P<0.05). Conclusion:DSS can improve the cognitive function of VD rats, and the mechanism may be related with reducing oxidative stress level, inhibiting neuronal apoptosis, and upregulating LEP-R/GSK-3β/Tau signaling pathway.
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Objective: To explore the effect of modified Danggui Shaoyao San on matrix metalloproteinase-2(MMP-2), intercellular adhesion molecule-1 (ICAM-1), emorheology and inflammation in patients with chronic pelvic inflammatory disease. Method: Patients with chronic pelvic inflammatory disease during May 2017 to June 2018 were randomly divided into treatment group and control group, with 37 cases in each group. The control group was orally treated with levofloxacin tablets and metronidazole tablets. In addition to the therapy of the control group, the treatment group was also given modified Danggui Shaoyao San. Traditional Chinese medicine(TCM) symptom scores, MMP-2 and ICAM-1, interleukin-1β(IL-1β), IL-6, IL-4, IL-10, and transforming growth factor-β (TGF-β), grain-megakaryocyte colony stimulating factor (GM-CSF), tumor necrosis factor-alpha (TNF-α), whole blood viscosity (ηb), plasma viscosity (ηp), erythrocyte aggregation index (AI), fibrinogen (Fib) before and after treatment were compared between two groups. The efficacy, adverse reactions and recurrence were observed in two groups. Result: The clinical efficacy of treatment group was better than that of the control group (Z=2.791, PPα, IL-1β, IL-6, GM-CSF,ηb,ηp, AI, Fib levels in the treatment group after treatment were significantly lower than those in control group (Pβ1 levels in treatment group after treatment were significantly higher than those in control group (Pχ2=6.198, PConclusion: Modified Danggui Shaoyao San has a significant clinical efficacy in the treatment of CPID, and can effectively relieve clinical symptoms and greatly reduce the recurrence rate, which may be related to the improvement of the regulation of MMP-2 and ICAM-1 levels, the inhibition of inflammatory reactions and the improvement of hemorheology.
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Objective:To investigate the effect of endothelin-1 (ET-1) on the expression of phosphorylated myosin light chain Ⅱ(p-MLCⅡ)and myosin light chain Ⅱ(MLCⅡ)protein in rat hepatic stellate cells HSC-T6 and explore the intervention effect of Danggui Shaoyao San(DSS)drug-containing serum. Method:After HSC-T6 cells were seeded, DMEM and blank rat serum with final concentrations of 2.5%, 5%, 10%, 15% and 20% were added to each well. The viability of HSC-T6 cells was determined by methyl thiazolyl tetrazolium(MTT) assay to screen the suitable serum concentration range. The cells were divided into blank serum control group (5%, 10%, 15%) and DSS drug-containing serum group (5%, 10%, 15%). ELISA was used to detect the content of ET-1 in cell culture supernatant under basic state. The cells were divided into blank serum control group (10%), DSS drug-containing serum low (5%), medium (10%) and high dose (15%) groups. Real-time fluorescent quantitative polymerase chain reaction (Real-time PCR) was used to detect the level of ET-1 mRNA in cell culture supernatant under basic state. The cells were divided into blank serum control group (10%), model group (10%), DSS drug-containing serum low (5%), medium (10%), high dose (15%) groups and Y-27632 inhibitor group (100 μmol·L-1). Except the blank serum control group, the other groups all received 10 nmol·L-1 ET-1 to induce HSC-T6 cells. Western blot was used to detect the expression of p-MLCⅡ and MLCⅡ in HSC-T6 cells induced by ET-1. Result:Serum concentrations of 5%, 10% and 15% were used as drug-containing serum concentrations. As compared with the blank serum control group, the DSS drug-containing serum group significantly reduced the relative content of ET-1 and ET-1 mRNA in the basic state (PPPPPConclusion:DSS drug-containing serum may down-regulate the expression of p-MLCⅡ and MLCⅡ by down-regulating the content of ET-1 and inhibiting the autocrine of ET-1.
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Aim To investigate the Na+,k +-ATPase, cyclc-AMP (cAMP), protein kinase A (PKA), cAMP-responsive element binding protein (CREB) and aquaporin 2 (AQP2) of the inner medullary col-lecting duct cell (IMCD3) induced by adramycin (ADR) , and to study on the protection mechanism of Danggui-Shaoyao-San (DSS) from the perspective of water-liquid balance. Methods IMCD3 cells were used as the research object. The effects of different concentrations of ADR on the proliferation of IMCD3 cells were determined by MIT assay. There were six groups in the cell experiment, namely, control group, model group, low-, medium-, and high-dose DSS extract (concentrations of 0. 8, 1.6, 3.2 g L"1) and H-89 inhibitor group. ELISA was used to detect the content of cAMP in cells. Changes of Na+ , K +-ATPase activity in cells were detected by Na+,k +-ATPase assay kit. The level of PKA, CREB and AQP2 mRNA in cells were detected by real-time PCR. The protein expression of PKA, CREB and AQP2 in IM-CD3 cells was assessed by Western blot. Results 1 x 10~8 mol L"1 ADR was the optimum concentration in IMCD3 cells. Different concentrations of DSS extract could effectively inhibit the injuiy of IMCD3 cells induced by ADR, and increase the activity of Na+,k +-ATPase and the content of cAMP in cell supernatant. DSS (1.6, 3.2 g L"1) extract could up-regulate the expressions of PKA, CREB and AQP2 mRNA and pro-tein expression (P < 0. 05 , P < 0. 01). Conclusion DSS extract can increase Na+,k +-ATPase activity and activate the cAMP-PKA-CREB pathway, thus inhibiting IMCD3 cell contraction mediated by ADR.
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Objective: To study the mechanism of action of Danggui Shaoyao San (DSS) in the treatment of Alzheimer'sdisease (AD) from a systematic network level using network pharmacology. Methods: The active ingredients of DSS intreating AD was screened from the Traditional Chinese Medicine System Pharmacology Database (TCSMP), and thedrugs-targets-disease network was built. Target organ localization network, GO analysis and Pathway enrichmentanalysis were used for bioinformatics analysis. Results: 51 kinds of DSS active ingredients were screened by networkpharmacology, showing 377 predicted targets of active compounds. Among them, 197 targets were associated with AD, and the pharmacodynamic targets of DSS active compounds were highly enriched with AD-related signals. The pathwaywas closely related to the reduction of Aβ aggregation and inhibition of tau hyperphosphorylation and biological processessuch as anti-inflammatory and anti-apoptosis, and the mRNA expression of the targeted gene was mainly enriched in theliver, heart, and brain. Conclusion: DSS active compounds interact with multiple targets in multiple ways in a synergisticmanner, mainly to produce important therapeutic effects on AD with anti-inflammatory, anti-apoptotic, enhancingmetabolism and immune system.
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Objective: To study the constituents in Danggui-Shaoyao-San (DSS, a powder prescription with Angelicae Sinensis Radix, Paeoniae Radix Alba, Atractylodis Macrocephalae Rhizoma, Poria, Alismatis Rhizome, and Chuanxiong Rhizome). Methods: To analyze the 70% ethanol-water extract from DDS by LC-Q-TOF-MS and LC-IT-MS. The analysis was performed on an Agilent Zorbax SB-C18 column and Rapid Resolution HT column (50 mm × 4.6 mm, 1.8 μm) with gradient elution of 0.05% formic acid (A)-acetonitrile (B) at the flow rate of 0.6 mL/min. The constituents from DSS were identified according to the relative molecule weight by LC-Q-TOF-MS and multistage MS informatics by LC-IT-MS. All the peaks were assigned to the corresponding traditional Chinese medicinal materials. Results: Thirty compounds from DDS could be identified, and 14 of them were identified for the first time in DSS including six organic acids, such as chlorogenic acid (8) and galloylsucrose (5-7); two protostane trierpenoids, such as 16-oxoalisol A 23-acetate (22); four cyclopeptides, such as cyclo tetraleucyl (or isoleucyl) (14) and other types of compounds, such as N-(1-deoxy-D-fructos-1-yl) pyroglutamic acid (1) and adenosine (3). Conclusion: This work investigates the compounds from DSS, and the results could provide the evidence for the research on active constituents and quality control of DSS.