Anti-depression targets and mechanism study of Kaixin San / 中国中药杂志

This study identified the anti-depression targets of Kaixin San(KXS) in the brain tissue with "target fishing" strategy, and explored the target-associated pharmacological signaling pathways to reveal the anti-depression molecular mechanism of KXS. The Balb/c mouse model of depression was established by chronic unpredictable mild stress(CUMS) and the anti-depression effect of KXS was evaluated by forced swimming test and sucrose preference test. KXS active components were bonded to the benzophenone-modified magnetic nanoparticles by photocrosslinking reaction for capturing target proteins from cortex, thalamus and hippocampus of depressive mice. The target proteins were identified by liquid chromatography-mass spectrometry/mass spectrometry(LC-MS/MS). The enrichment analysis on signaling pathways was performed by Cytoscape. The potential biological functions of targets were verified by immunohistochemistry and Western blot assay. The results showed that KXS significantly improved the behavioral indexes. There were 64, 91, and 44 potential targets of KXS identified in cortex, thalamus, and hippocampus, respectively, according to the target identification experiment. The functions of these targets were mainly associated with vasopressin-regulated water reabsorption, salmonella infection, thyroid hormone synthesis, and other signaling pathways. Besides, the results of immunohistochemistry and Western blot showed that KXS up-regulated the expressions of argipressine(AVP) in the cortex, heat shock protein 60(HSP60), cytochrome C oxidase 4(COX4), and thyrotropin-releasing hormone(TRH) in the thalamus, and down-regulated the expressions of tumor necrosis factor-α(TNF-α) and nuclear factor kappa B(NF-κB) p65 in the thalamus. Therefore, KXS may exert anti-depression effect through regulating vasopressin signaling pathway in the cortex and inflammation, energy metabolism, and thyroid hormone signaling pathways in the thalamus, and the effect of KXS on hippocampus is not significant.

Anti-infectious pneumonia target discovery and molecular mechanism study of Jingfang Granules / 中国中药杂志

This study aimed to identify the direct pharmacological targets of Jingfang Granules in treating infectious pneumonia via "target fishing" strategy. Moreover, the molecular mechanism of Jingfang Granules in treating infectious pneumonia was also investigated based on target-related pharmacological signaling pathways. First, the Jingfang Granules extract-bound magnetic nanoparticles were prepared, which were incubated with lipopolysaccharide(LPS)-induced mouse pneumonia tissue lysates. The captured proteins were analyzed by high-resolution mass spectrometry(HRMS), and the target groups with specific binding to the Jingfang Granules extract were screened out. Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment analysis was used to identify the target protein-associated signaling pathways. On this basis, the LPS-induced mouse model of infectious pneumonia was established. The possible biological functions of target proteins were verified by hematoxylin-eosin(HE) staining and immunohistochemical assay. A total of 186 Jingfang Granules-specific binding proteins were identified from lung tissues. KEGG pathway enrichment analysis showed that the target protein-associated signaling pathways mainly included Salmonella infection, vascular and pulmonary epithelial adherens junction, ribosomal viral replication, viral endocytosis, and fatty acid degradation. The target functions of Jingfang Granules were related to pulmonary inflammation and immunity, pulmonary energy metabolism, pulmonary microcirculation, and viral infection. Based on the in vivo inflammation model, Jingfang Granules significantly improved the alveolar structure of the LPS-induced mouse model of infectious pneumonia and down-regulated the expressions of tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6). Meanwhile, Jingfang Gra-nules significantly up-regulated the expressions of key proteins of mitochondrial function COX Ⅳ and ATP, microcirculation-related proteins CD31 and Occludin, and proteins associated with viral infection DDX21 and DDX3. These results suggest that Jingfang Gra-nules can inhibit lung inflammation, improve lung energy metabolism and pulmonary microcirculation, resist virus infection, thus playing a protective role in the lung. This study systematically explains the molecular mechanism of Jingfang Granules in the treatment of respiratory inflammation from the perspective of target-signaling pathway-pharmacological efficacy, thereby providing key information for clinical rational use of Jingfang Granules and expanding potential pharmacological application.

Potentiating effect and mechanism of extract of Jingfang Granules on activation of macrophages / 中国中药杂志

This study aimed to explore the potentiating effect and mechanism of the extract of Jingfang Granules(JFG) on the activation of macrophages. The RAW264.7 cells were treated with JFG extract and then stimulated by multiple agents. Subsequently, mRNA was extracted, and reverse transcription-polymerase chain reaction(RT-PCR) was used to measure the mRNA transcription of multiple cytokines in RAW264.7 cells. The levels of cytokines in the cell supernatant were detected by enzyme-linked immunosorbent assay(ELISA). In addition, the intracellular proteins were extracted and the activation of signaling pathways was determined by Western blot. The results showed that JFG extract alone could not promote or slightly promote the mRNA transcription of TNF-α, IL-6, IL-1β, MIP-1α, MCP-1, CCL5, IP-10, and IFN-β, and significantly enhance the mRNA transcription of these cytokines in RAW264.7 cells induced by R848 and CpG in a dose-dependent manner. Furthermore, JFG extract also potentiated the secretion of TNF-α, IL-6, MCP-1, and IFN-β by RAW264.7 cells stimulated with R848 and CpG. As revealed by mechanism analysis, JFG extract enhanced the phosphorylation of p38, ERK1/2, IRF3, STAT1, and STAT3 in RAW264.7 cells induced by CpG. The findings of this study indicate that JFG extract can selectively potentiate the activation of macrophages induced by R848 and CpG, which may be attributed to the promotion of the activation of MAPKs, IRF3, and STAT1/3 signaling pathways.

Therapeutic effect and mechanism of Mailuo Shutong Pills on posterior limb swelling caused by femur fracture in rats based on intestinal flora and intestinal metabolism / 中国中药杂志

This study aimed to investigate the protective effect and underlying mechanism of Mailuo Shutong Pills(MLST) on posterior limb swelling caused by femur fracture in rats. The rats were randomly divided into a sham operation group, a model group, a low-dose MLST group(1.8 g·kg~(-1)·d~(-1)), a high-dose MLST group(3.6 g·kg~(-1)·d~(-1)), and a positive drug group(60 mg·kg~(-1)·d~(-1) Maizhiling Tablets). The femur in the sham operation group was exposed and the wound was sutured, while the other four groups underwent mechanical damage to cause femur fracture. The rats were treated with corresponding drugs by gavage 7 days before modeling and 5 days after modeling, while those in the sham operation group and the model group were given an equivalent dose of distilled water by gavage. Hematoxylin-eosin(HE) staining was used to detect the pathological injury of the posterior limb muscle tissues in rats, and the degree of hind limb swelling was measured. The enzyme-linked immunosorbent assay(ELISA) kit was used to detect the expression levels of interleukin-6(IL-6), interleukin-1β(IL-1β), and tumor necrosis factor-α(TNF-α) in the serum of rats in each group. The activity of superoxide dismutase(SOD), malondialdehyde(MDA), catalase(CAT), and glutathione peroxidase(GSH-Px) in rat serum was also measured. Western blot was used to detect the protein expression levels of heme oxygenase 1(HO-1), NAD(P)H quinone oxidoreductase 1(NQO1), and nuclear transcription factor E2-related factor 2(Nrf2) in rat posterior limb muscle tissues. The changes in the intestinal flora and intestinal metabolites in rats were detected by 16S rDNA sequencing and ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS), respectively, to explore the underlying mechanism of MLST in treating posterior limb swelling caused by femur fracture in rats. Compared with the model group, MLST significantly improved the degree of posterior limb swelling in rats, reduced the levels of serum inflammatory factors, and alleviated oxidative stress injury. The HE staining results showed that the inflammatory infiltration in the posterior limb muscle tissues of rats in the MLST groups was significantly improved. Western blot results showed that MLST significantly increased the protein expression of HO-1, NQO1, and Nrf2 in rat posterior limb muscle tissues compared with the model group. The 16S rDNA sequencing results showed that MLST improved the disorder of intestinal flora in rats after femur fracture. The UPLC-MS/MS results showed that MLST significantly affected the bile acid biosynthesis and metabolism pathway in the intestine after femur fracture, and the Spearman analysis confirmed that the metabolite deoxycholic acid involved in bile acid biosynthesis was positively correlated with the abundance of Turicibacter. The metabolite cholic acid was positively correlated with the abundance of Papilibacter, Staphylococcus, and Intestinimonas. The metabolite lithocholic acid was positively correlated with Papilibacter and Intestinimonas. The above results indicated that MLST could protect against the posterior limb swelling caused by femur fracture in rats. This protective effect may be achieved by improving the pathological injury of the posterior limb muscle, reducing the expression levels of inflammatory and oxidative stress-related factors in serum, reducing the oxidative injury of the posterior limb muscle, improving intestinal flora, and balancing the biosynthesis of bile acids in the intestine.

Molecular mechanism study of Shouhui Tongbian Capsules on promoting energy metabolism of gastrointestinal stromal cells / 中国中药杂志

Mechanism study was performed to explore how Shouhui Tongbian Capsules promotes energy metabolism of gastrointestinal stromal cells. In this study, gastrointestinal stromal cells line GIST-882 was used as the model to explore energy metabolism regulation effects of Shouhui Tongbian Capsules extract(10, 20, 50 and 100 μg·mL~(-1)) by measuring the cell proliferation, ATP level, mitochondrial membrane potential, and mitochondrial isocitrate dehydrogenase activity. Meanwhile, Western blot was used to detect the proteins expression of SCF/c-Kit and CDK2/cyclin A signaling pathways. Our results showed that Shouhui Tongbian Capsules promoted cell proliferation and increased ATP level of gastrointestinal stromal cells. In addition, Shouhui Tongbian Capsules obviously improved mitochondrial structural integrity, and increased mitochondrial membrane potential in GIST-882 cells. Mechanism study revealed that Shouhui Tongbian Capsules increased mitochondrial isocitrate dehydrogenase activity and up-regulated the proteins expression of SCF/c-Kit and CDK2/cyclin A signaling pathways. Collectively, our study indicated that Shouhui Tongbian Capsules promoted the energy metabolism for gastrointestinal stromal cells proliferation by activating mitochondrial isocitrate dehydrogenase to induce ATP production, as well as activating SCF/c-Kit and CDK2/cyclin A signaling pathways.

Metabonomics study of Shouhui Tongbian Capsules in slow transit constipation based on UPLC-ESI-QE-Orbitrap-MS / 中国中药杂志

The effect of Shouhui Tongbian Capsules(SHTB) on the endogenous metabolites of colon tissue in mice with slow transit constipation was analyzed by metabolomics methods to explore its mechanism in the treatment of constipation. ICR mice were randomly divided into normal group, model group and SHTB group according to the body weight. The mice were given diphenoxylate to establish the slow transit constipation model. Mouse carbon ink pushing rate, first defecation time and the number of defecation particles in 12 h were observed. The mouse colon tissue was separated and the mucous cells were detected by Periodic acid Schiff and Alcian blue(AB-PAS) staining. Ultra-high-performance liquid chromatography electrospray ionization orbitrap tandem mass spectrometry(UPLC-ESI-Orbitrap-MS/MS) technology was used to characterize the differences in tissue metabolism to screen out the potential different metabolites and possible metabolic pathways in colon tissue. The results indicated that SHTB could significantly shorten the first defecation time and the number of defecations, and increase the number of intestinal peristalsis and mucous cells in the colonic mucosa compared to the model mice. Metabolomics results showed that, compared with the normal group, a total of 17 potential biomarkers, including L-kynurenine, N6,N6,N6-trimethyl-L-lysine, L-formylkynurenine, N6-acetyl-L-lysine, L-phenylalanine, phenylacetaldehyde, xanthoxin, thymidine, glycyl-L-leucine, cystathionine,(R)-1-aminopropan-2-ol, deoxycytidine, gamma-glutamyl-gamma-aminobutyraldehyde, D-galactose, L-arginine, L-proline and pyruvate, were found and identified in colon tissue. Treated with SHTB, these metabolic differences tended to return to normal levels. Therefore, it could be made a conclusion that the therapeutic effect of SHTB on chronic transit constipation may be related to regulating phenylalanine metabolism, phenylalanine, tyrosine and tryptophan biosynthesis, arginine and proline metabolism, cysteine and methionine metabolism, tyrosine metabolism, arginine biosynthesis, pyruvate metabolism, glycolysis, pyrimidine metabolism, tricarboxylic acid cycle and galactose metabolism.

Protective effect of icaritin on focal cerebral ischemic–reperfusion mice / 中草药·英文版

Objective: To investigate the protective effects of icaritin (ICT), one of the active ingredients in Epimedii Folium, on mouse model of cerebral ischemia-reperfusion (I/R) in vivo. Methods: ICR mice were subjected to an 1 h transient middle cerebral artery occlusion (MCAO) and followed by 24 h of reperfusion. Neurological deficits, infarct volume, brain edema and survive rate were measured, respectively. The levels of brain IL-1β TNF-α ROS and DNA-binding activity of NF-κB p65 were measured by ELISA kits. The levels of malondialdehyde (MDA) and activities of superoxide dismutase (SOD) were detected by spectrophotometry, and the release of nitric oxide (NO) were detected by Griess kit. Results: ICT markedly reduced the neurological deficit scores, brain edema, infarct volume and increased the survival rate of the cerebral I/R mice. The expression of IL-1β TNF-α NO, MDA and DNA-binding activity of NF-κB p65 were significantly inhibited by ICT, while the activity of SOD were up-regulated at the same time. Conclusion: ICT possessed significant neuroprotective effects in cerebral I/R mice, which might be related to prevent neuroinflammatory and oxidative damage.

Efficient discovery and capturing of nNOS-PSD-95 uncouplers from Trifolium pratense / 中国中药杂志

Magnetic molecularly imprinted polymers(MMIPs) were prepared with ZL006 as template, acrylamide(AA) as the functional monomer, and acetonitrile as pore-forming agent; then Fourier transform infrared spectroscopy(FT-IR) and scanning electron microscopy(SEM) were used to characterize their forms and structures. Simultaneously, the MMIPs prepared previously were used as sorbents for dispersive magnetic solid phase extraction(DSPE) to capture and identify potential nNOS-PSD-95 uncouplers from extracts of Trifolium pratense and the the activities of the screened compounds were evaluated by the neuroprotective effect and co-immunoprecipitation test. The experiment revealed that the successfully synthesized MMIPs showed good dispersiveness, suitable particle size and good adsorption properties. Formononetin, prunetin and biochanin A were separated and enriched from Trifolium pratense by using the MMIPs as artificial antibodies and finally biochanin A was found to have higher cytoprotective action and uncoupling action according to the neuroprotective effect and co-immunoprecipitation test.

Efficacy comparison of different points combination in the treatment of menopausal insomnia: a randomized controlled trial / 中国针灸

<p><b>OBJECTIVE</b>To compare the efficacy of different points combination in the treatment of menopausal insomnia.</p><p><b>METHODS</b>Ninety-six cases of menopausal insomnia were randomized into 3 groups, Xinshu (BL 15), Shenshu (BL 23), Sishencong (EX-HN 1), Shenmen (HT 7), Sanyinjiao (SP 6) were chosen in the restore interaction between the heart and the kidney group (group A, 32 cases); Zhaohai (KI 6), Jiaoxin (KI 8), Shenmai (BL 62), Pucan (BL 61) were chosen in the acupuncturing qiao mai group (group B, 32 cases); auricular Shenmen (TF4) and sensitive spot at the distribution area of auricular vagus nervus were chosen in the ear acupuncture group (group C, 32 cases). Six days made one session and the treatments were finished after 4 courses. The polysomnography (PSG) and Pittsburgh sleep quality index (PSQI) were employed before and after treatment to evaluate the alleviation of insomnia.</p><p><b>RESULTS</b>The parameters of the sleep latency (SL), rapid wave sleep latency (RL) and sleep efficiency (SE) were significantly improved in the three groups, and the differences were statistically significant (P < 0.05, P < 0.01). The SL and awaking time (AT) in group C [SL (401.08 +/- 16.54) min and AT (4.87 +/- 2.64) times] were significantly superior to those in the other two groups [SL (50.36 +/- 18.47) min, (54.87 +/- 20.92) min, AT (5.98 +/- 2.11) times, (6.13 +/- 3.04) times, all P < 0.05]. The S(3+4) (%) in group C was also significantly higher than those in the other two groups (both P < 0.05). It was indicated by PSQI that the sleep quality of group C (0.78 +/- 0.12) was significantly superior to that in group B (1.32 +/- 0.29), the total score and cured and markedly effective rate in group C [(4.34 +/- 1.43), 68.8% (22/32)] were superior to those in group A [(7.48 +/- 3.09), 53.1% (17/32), both P < 0.05].</p><p><b>CONCLUSION</b>Ear acupuncture has a better curative effect than the restore interaction between the heart and the kidney group and acupuncturing qiao mai group, it is worth of being promoted.</p>

Inhibitory effect of arctigenin on lymphocyte activation stimulated with PMA/ionomycin / 药学学报

This study investigated the effect of arctigenin (Arc) on the cell activation, cytokines expression, proliferation, and cell-cycle distribution of mouse T lymphocytes. Mouse lymphocytes were prepared from lymph node and treated with Phorbol-12-myristate-13-acetate (PMA)/Ionimycin (Ion) and/or Arc. CD69, CD25, cytokines, proliferation and cell cycle were assayed by flow cytometry. The results showed that, at concentrations of less than 1.00 micromol x L(-1), Arc expressed non-obvious cell damage to cultured lymphocytes, however, it could significantly down-regulate the expression of CD69 and CD25, as well as TNF-alpha, IFN-gamma, IL-2, IL-4, IL-6 and IL-10 on PMA/Ion stimulated lymphocytes. At the same time, Arc could also inhibit the proliferation of PMA/Ion-activated lymphocytes and exhibited lymphocyte G 0/G1 phase cycle arrest. These results suggest that Arc possesses significant anti-inflammatory effects that may be mediated through the regulation of cell activation, cytokines expression and cell proliferation.

Treatment of peripheral facial paralysis with acupuncture at Renying (ST 9) mainly cooperated with stellate ganglion block: a randomized controlled trial / 中国针灸

<p><b>OBJECTIVE</b>To explore the better therapy for peripheral facial paralysis.</p><p><b>METHODS</b>One hundred and twenty patients were randomized into three groups: a common acupuncture group: acupuncture at Yangbai (GB 14), Sibai (ST 2) and Yingxiang (LI 20) as main acupoints, a ST 9 group: acupuncture at Renying (ST 9) as main and a ST 9 plus SGB group: acupuncture at Renying (ST 9) as main cooperated with stellate ganglion block (SGB). Once daily, 7 treatments made one session. After three sessions of treatment, the latency period and amplitude of evoked potential in ENoG, R1 value and R2 value of blink reflex were compared before and after the treatment in different groups separately. The total therapeutic effect was evaluated after treatment.</p><p><b>RESULTS</b>All the treatments shortened the latency period of ENoG, and elevated the amplitude evoked potential significantly. After treatment, the latency period in ST 9 plus SGB group was reduced significantly as compared with common acupuncture group (P < 0.05). The amplitude of evoked potential in ST 9 group was increased significantly as compared with the other two groups (both P < 0.05). After treatment, in each group, R1 and R2 values were shortened significantly. The difference values of R1 and R2 in ST 9 group and ST 9 plus SGB group were all significantly higher as compared with common acupuncture group (both P < 0.05). Additionally, the difference value of R1 in ST 9 plus SGB group was higher significantly than that in ST 9 group (P < 0.05). The clinical cured and remarkably effective rate was 87.5% (35/40) in ST9 plus SGB group, which was higher than 77.5% (31/40) in ST 9 group, and 65.0% (26/40) in common acupuncture group (P < 0.05).</p><p><b>CONCLUSION</b>As compared with common acupuncture group, ST 9 group and ST 9 plus SGB group achieve the much superior efficacy on peripheral facial paralysis. The treatment with ST 9 acupuncture and SGB can better repair the early reflex induced by the injury of facial nerve.</p>