Mechanism of Schisandra Chinensis-mediated microglia phenotypic transformation by regulation of the TLR4 pathway based on miR-124 / 药学学报

To investigate the mechanism by which Schisandra Chinensis mediates the phenotypic transformation of microglia via microRNA-124 (miR-124)-based regulation of the Toll-like receptor 4 (TLR4) pathway, a model was established using lipopolysaccharide (LPS) stimulation of BV2 cells. Cells were treated with different doses of Schisandra Chinensis extract (SCE). MiR-124 inhibitors and negative control sequences (NC inhibitor) were transfected into LPS-induced BV2 cells and treated with SCE. The MTT assay was used for cell activity detection; an NO kit was used to measure NO release; ELISA kits were used to measure the levels of interleukin-10 (IL-10) and tumor necrosis factor-α (TNF-α). Microglia markers, including ionized calcium binding adapter molecule-1 (IBA-1) and arginase-1 (Arg-1), and the nuclear translocation of nuclear factor-kappa B (NF-κB) were evaluated by immunofluorescent staining. NF-κB p65, IBA-1, Arg-1, TLR4, myeloid differentiation primary factor 88 (MyD88), inhibitor of nuclear factor-kappa B kinases-α (IKK-α), IL-10, TNF-α were detected by immunoblot. SCE at concentrations ranging from 31.25 to 250 μg·mL-1 had no significant effect on cell activity. SCE treatment significantly inhibited NO release induced by LPS (P < 0.001, P < 0.01), increased the level of IL-10 (P < 0.05), and decreased the level of TNF-α (P < 0.001). In addition, SCE significantly reduced the expression of TNF-α, IBA-1, TLR4, and MyD88 (P < 0.01, P < 0.001) and elevated the expression of IL-10, Arg-1, NF-κB P65 and IKK-α (P < 0.001, P < 0.01, P < 0.05). SCE treatment could also promote the expression of miR-124 (P < 0.01). However, transfection with the miR-124 inhibitor increased TNF-α (P < 0.001), decreased the level of IL-10 (P < 0.05), increased the mRNA level and the protein expression of TNF-α and IBA-1 (P < 0.05, P < 0.01, P < 0.001), and decreased the mRNA level and protein expression of IL-10 and Arg-1 (P < 0.001, P < 0.01). In addition, the inhibition of TLR4 and MyD88 was attenuated. In conclusion, SCE appears to inhibit the activation of TLR4 signaling pathway by upregulating miR-124 so as to inhibit microglia M1 polarization and promote microglia M2 polarization.

Effects of raw and wine-processed Schisandra chinensis on neuro-endocrine-immune network in insomnia mice and its mechanism / 中国药房

OBJECTIVE To investigate the effect of raw and wine-processed Schisandra chinensis on neuro-immune-endocrine network in insomnia mice and its mechanism. METHODS Fifty mice were randomly divided into blank group， model group， diazepam group， raw S. chinensis group and wine-processed S. chinensis group， with 10 mice in each group. Except for blank group， the mice in the other groups were intraperitoneally injected with thyroxine solution to establish mice model of insomnia； at the end of each day’s modeling， the corresponding doses of diazepam，raw and wine-processed S. chinensis were given by gavage. The blank group and model group were given constant volume of normal saline. The general state of the mice was observed and recorded， and the total activity distance and upright times of the mice were detected； the EEG and EMG signals of mice were recorded， and the time ratio of sleep wake time （wake）， non-rapid eye movement （NREM） and rapid eye movement （REM） was analyzed； the contents of neurotransmitters [γ-aminobutyric acid （GABA）， 5-hydroxytryptamine （5-HT）， dopamine （DA）， norepinephrine （NE）， cortisol （CORT）] in brain suprachiasmatic nucleus （SCN） were detected； and the expressions of tumor necrosis factor α （TNF-α） and interleukin 1β （IL-1β） were detected； the mRNA expressions of clock gene Bmal1， circadian clock gene Clock and cycle gene Per2 were all detected. RESULTS Compared with the blank group， the mental state of the model group mice was relatively depressed， the amount of food and water increased， the body mass decreased， the hair was rough and shiny， and the circadian rhythm was irregular； the total activity distance and upright times decreased significantly； the time ratio of wake increased significantly， while the time ratios of REM and NREM decreased significantly； the content of 5- HT in brain SCN decreased significantly， while the content of NE， DA and CORT increased significantly； the fluorescence intensity of IL-1β and TNF-α was significantly increased； the relative expression level of Bmal1 and Clock mRNA was significantly increased， while the relative expression level of Per2 mRNA was significantly decreased （P＜0.05 or P＜0.01）. Compared with the model group， the general state of mice in diazepam group， raw S. chinensis group and wine-processed S. chinensis group was improved obviously， and most of the above index levels were significantly reversed （P＜0.05 or P＜0.01）. CONCLUSIONS Raw and wine-processed S. chinensis have a certain therapeutic effect on insomnia mice， the mechanism of which may be related to the regulation of neuro-endocrine-immune system related biological indicators in insomnia mice.

Investigation of pharmacodynamic material basis of Schisandra chinensis in the treatment of allergic asthma / 中国药房

OBJECTIVE To study the pharmacological basis of Schisandra chinensis in the treatment of allergic asthma. METHODS The common components of 10 batches of S. chinensis from different habitats were analyzed by UPLC-Q-TOF-MS/MS. Furthermore， the allergic asthma model was established by intraperitoneal injection of ovalbumin （OVA） and aluminum hydroxide for stimulation combined with atomization exitation； general behavioral observation and the contents of interferon γ （IFN-γ）， interleukin-4 （IL-4） and immunoglobulin E （IgE） in serum were taken as criteria for evaluating the therapeutic effect of S. chinensis from different habitats in the treatment of allergic asthma. Correlation coefficients between common peak area and efficacy evaluation index of each batch of medicinal material were analyzed through grey correlation degree and Pearson correlation analysis. RESULTS A total of 21 common components were identified in 10 batches of S. chinensis from different habitats. After administration of S. chinensis， symptoms such as shortness of breath， sneezing and curling of rats were alleviated. In addition， the content of IFN-γ was significantly increased while the contents of IL-4 and IgE in serum were distinctly decreased （P＜0.01）. Grey correlation analysis showed that 11 common components had high correlation coefficients with IFN-γ， IL-4 and IgE （rˉ＞0.8）. Pearson correlation analysis showed that 8 components were significantly positively correlated with the content of IFN-γ （P＜ 0.05）， and 9， 8 components were significantly negatively correlated with the content of IL-4 and IgE （P＜0.05）. Based on the results of grey correlation degree and Pearson correlation analysis， 7 components such as peak 3， 4， 6， 7， 9， 19 and 20， were highly related to S. chinensis in the treatment of allergic asthma. CONCLUSIONS Schisandrol A， schisandrin B， schisandrin C， gomisin M2， gomisin J， pregomisin and angeloylgomisin H are the potential pharmacodynamic substance basis of S. chinensis in the treatment of allergic asthma.

Research progress on chemical constituents of Schisandra chinensis and its effect on nonalcoholic fatty liver disease / 中国中药杂志

Schisandra chinensis, a traditional Chinese medicinal herb, is rich in chemical constituents, including lignans, triterpenes, polysaccharides, and volatile oils. Clinically, it is commonly used to treat cardiovascular, cerebrovascular, liver, gastrointestinal, and respiratory diseases. Modern pharmacological studies have shown that S. chinensis extract and monomers have multiple pharmacological activities in lowering liver fat, alleviating insulin resistance, and resisting oxidative stress, and have good application prospects in alleviating nonalcoholic fatty liver disease(NAFLD). Therefore, this study reviewed the research progress on chemical constituents of S. chinensis and its effect on NAFLD in recent years to provide references for the research on S. chinensis in the treatment of NAFLD.

Effect of Polysaccharides from Root， Stem， Leaf and Fruit of Schisandra chinensis on Exercise Endurance in D-galactose-induced Aging Mice / 中国实验方剂学杂志

ObjectiveTo observe the effect of polysaccharides from root， stem， leaf and fruit of Schisandra chinensis on exercise endurance in the aging mice induced by D-galactose. MethodMale ICR mice were randomly assigned into six groups： blank control group， model group， root polysaccharide group， stem polysaccharide group， leaf polysaccharide group and fruit polysaccharide group. The mice were administrated with distilled water or root， stem， leaf and fruit polysaccharide （total sugar content of 35 mg·kg-1） by gavage. Thirty minutes after the administration， the blank control group was subcutaneously injected with normal saline， and the other groups with D-galactose （300 mg·kg-1）， once daily for 6 weeks. The anti-fatigue effects were evaluated by rotarod test， forelimb grip strength test， and weight-loaded swimming test. The fatigue and oxidation indicators such as blood urea nitrogen （BUN）， serum lactic acid （LD）， lactic dehydrogenase （LDH）， creatine kinase （CK）， superoxide dismutase （SOD）， malondialdehyde （MDA）， glutathione peroxidase （GSH-Px）， and reactive oxygen species （ROS） were measured by chemical colorimetry. The protein levels of pro-apoptotic protein B cell lymphoma-2 （Bcl-2）-associated X protein （Bax）， anti-apoptotic Bcl-2 and cleaved cysteinyl aspartate-specific protease-3 （cleaved Caspase-3） in mouse skeletal muscle were detected by Western blot. ResultIn the rotarod test， the time on rod was shorter in the model group than in the blank control group （P<0.01） and the root， stem and fruit polysaccharide groups （P<0.01）. In the forelimb grip strength test， the forelimb grip strength in the model group was lower than that in the blank control group （P<0.01） and the root， stem， leaf and fruit polysaccharide groups （P<0.01）. In the weight-loaded swimming test， the weight-loaded swimming time in the model group was shorter than that in the blank control group （P<0.01） and the root， stem， leaf and fruit polysaccharide groups （P<0.01）. Compared with those in the blank control group， the BUN， LD， LDH and CK levels significantly increased in the model group （P<0.05， P<0.01）. The increases in BUN and LDH levels were decreased by the root， stem and fruit polysaccharides （P<0.05， P<0.01） and those in LD and CK by the root， stem， leaf and fruit polysaccharides （P<0.05， P<0.01）. Compared with the blank control group， the model group showed decreased SOD and GSH-Px activities （P<0.01） and increased MDA and ROS content （P<0.01）. Compared with the model group， the root， stem， and fruit polysaccharide increased the SOD activity （P<0.05， P<0.01） and decreased ROS content （P<0.01）. The root and stem polysaccharides decreased the MDA content （P<0.01） and increased the GSH-Px activity （P<0.05， P<0.01）. Compared with the blank control group， the model group showed up-regulated protein levels of Bax and cleaved Caspase-3 and down-regulated protein level of Bcl-2 （P<0.01）. Compared with the model group， the root， and stem polysaccharides down-regulated the protein levels of cleaved Caspase-3 （P<0.05） and up-regulated protein level of Bcl-2 （P<0.01）. ConclusionThe polysaccharides from the root， stem， leaf， and fruit of S. chinensis have anti-fatigue effect in D-galactose-induced aging mice. The polysaccharides may exert such effect by improving the antioxidant capacity and inhibiting the apoptosis of skeletal muscle cells.

Molecular cloning and characterization of three phenylalanine ammonia-lyase genes from Schisandra chinensis / 中国天然药物

Phenylalanine ammonia-lyase (PAL), which catalyzes the conversion from L-phenylalanine to trans-cinnamic acid, is a well-known key enzyme and a connecting step between primary and secondary metabolisms in the phenylpropanoid biosynthetic pathway of plants and microbes. Schisandra chinensis, a woody vine plant belonging to the family of Magnoliaceae, is a rich source of dibenzocyclooctadiene lignans exhibiting potent activity. However, the functional role of PAL in the biosynthesis of lignan is relatively limited, compared with those in lignin and flavonoids biosynthesis. Therefore, it is essential to clone and characterize the PAL genes from this valuable medicinal plant. In this study, molecular cloning and characterization of three PAL genes (ScPAL1-3) from S. chinensis was carried out. ScPALs were cloned using RACE PCR. The sequence analysis of the three ScPALs was carried out to give basic characteristics followed by docking analysis. In order to determine their catalytic activity, recombinant protein was obtained by heterologous expression in pCold-TF vector in Escherichia coli (BL21-DE3), followed by Ni-affinity purification. The catalytic product of the purified recombinant proteins was verified using RP-HPLC through comparing with standard compounds. The optimal temperature, pH value and effects of different metal ions were determined. Vmax, Kcat and Km values were determined under the optimal conditions. The expression of three ScPALs in different tissues was also determined. Our work provided essential information for the function of ScPALs.

Schisandra chinensis Oil Attenuates Aristolochic Acid I-Induced Nephrotoxicity in vivo and in vitro / 中国结合医学杂志

OBJECTIVE@#To investigate the protective effects of Schisandra chinensis oil (SCEO) against aristolochic acid I (AA I)-induced nephrotoxicity in vivo and in vitro and elucidate the underlying mechanism.@*METHODS@#C57BL/6 mice were randomly divided into 5 groups according to a random number table, including control group, AA I group, and AA I +SCEO (0.25, 0.5 and 1 g/kg) groups (n=5 per group). Pretreatment with SCEO was done for 2 days by oral administration, while the control and AA I groups were treated with sodium carboxymethyl cellulose. Mice of all groups except for the control group were injected intraperitoneally with AA I (5 mg/kg) from day 3 until day 7. Histopathological examination and apoptosis of kidney tissue were observed by hematoxylin and eosin and TdT-mediated dUTP nick-end labeling (TUNEL) staining, respectively. The levels of serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), and serum creatinine (SCr), as well as renal malondialdehyde (MDA), glutathione, r-glutamyl cysteingl+glycine (GSH), and superoxide dismutase (SOD) were analyzed using enzyme-linked immunosorbent assay (ELISA). Expressions of hepatic cytochrome P450 1A1 (CYP1A1), CYP1A2, and nad(p)hquinonedehydrogenase1 (NQO1) were analyzed using ELISA, quantitative real-time polymerase chain reaction (qPCR) and Western blot, respectively. In vitro, SCEO (40 µ g/mL) was added 12 h before treatment with AA I (40 µ mol/mL for 48 h) in human renal proximal tubule cell line (HK-2), then apoptosis and reactive oxygen species (ROS) were analyzed by flow cytometry.@*RESULTS@#SCEO 0.5 and 1 g/kg ameliorated histopathological changes and TUNEL+ staining in the kidney tissues of mice with AA I-induced nephrotoxicity, and reduced serum levels of ALT, AST, BUN and SCr (P<0.01 or P<0.05). SCEO 0.5 and 1 g/kg alleviated the ROS generation in kidney, containing MDA, GSH and SOD (P<0.01 or P<0.05). SCEO 1 g/kg increased the expressions of CYP1A1 and CYP1A2 and decreased NQO1 level in the liver tissues (P<0.01 or P<0.05). Besides, in vitro studies also demonstrated that SCEO 40 µ g/mL inhibited apoptosis and ROS generation (P<0.05 or P<0.01).@*CONCLUSIONS@#SCEO can alleviate AA I-induced kidney damage both in vivo and in vitro. The protective mechanism may be closely related to the regulation of metabolic enzymes, thereby inhibiting apoptosis and ROS production.

Effects of compatibility of Schisandra chinensis on Psoralea corylifolia -induced oxidative damage and endoplasmic reticulum stress in hepatocytes / 中国药房

OBJECTIVE To study the effects of the compatibility of Schisandra chinensis on Psoralea corylifolia -induced oxidative damage and endoplasmic reticulum stress in L 02 hepatocytes，and to provide reference for clinical use of the compatibility attenuation of P. corylifolia -S. chinensis . METHODS L02 cells were divided into blank control group ，P. corylifolia model group （1 200 μg/mL P. corylifolia ，calculated by crude drug ），P. corylifolia -S. chinensis low-dose，medium-dose and high-dose groups （1 200 μg/mL P. corylifolia combined with 600，1 200，2 400 μg/mL S. chinensis ，respectively，calculated by crude drug ）. After the cells in each group were cultured in culture medium or drug solution for 48 hours，the levels of aspartate aminotransferase（AST）and alanine aminotransferase （ALT）were detected ；the levels of glutathione （GSH），superoxide dismutase （SOD）and malondialdehyde （MDA）in cell culture medium were detected ；reactive oxygen species （ROS）level and mitochondrial membrane potential in cells were detected ；mRNA and protein expressions of glucose-regulated protein 78（GRP-78）and protein kinase R-like endoplasmic reticulum kinase （PERK）were detected. RESULTS Compared with blank control group ，the levels of AST，ALT，MDA and ROS ，mRNA and protein expressions of GRP- 78 and PERK were increased significantly in P. corylifolia model group （P＜0.05 or P＜0.01）；while GSH and SOD levels ，mitochondrial membrane potential were decreased significantly （P＜0.05 or P＜0.01）. Compared with P. corylifolia model group ，above indexes of P. corylifolia -S. chinensis low-dose， medium-dose and high-dose groups were all improved significantly （P＜0.05 or P＜0.01）. CONCLUSIONS The compatibility of P. corylifolia -S. chinensis can alleviate P. corylifolia -induced oxidative damage and endoplasmic reticulum stress of L 02 cells.

A new norsesquiterpenoid from Schisandra chinensis / 中国中药杂志

This study investigated the chemical components from the leaves and stems of Schisandra chinensis. Three norsesquiterpenoids were isolated from S. chinensis by various column chromatographies(silica gel, Sephadex LH-20, and MCI), reversed-phase medium-pressure preparative, and semi-preparative high-performance liquid chromatography(HPLC). Their structures were identified based on physicochemical properties, mass spectrometry(MS), nuclear magnetic resonance(NMR), ultraviolet(UV), and electro-nic circular dichroism(ECD) as(3R,4R,5R,6S,7E)-3,4,5,6-tetrahydroxy-7-megastigmen-9-one(1),(3S,5R,6R,7E)-3,5,6-trihydroxy-7-megastigmen-9-one(2), and(3S,4R,9R)-3,4,9-trihydroxymegastigman-5-ene(3). Compound 1 was a new compound, and its absolute configuration was determined by ECD. Compounds 2 and 3 were isolated from the Schisandra plant for the first time.

Pharmacological effects of Schisandra Chinensis Fructus polysaccharide on intestinal protection / 中国药理学与毒理学杂志

Schisandra Chinensis Fructus (SCF) is the fruit of Schisandra chinensis (Turcz.) Baill., a perennial vine. It was first recorded in Shen Nong's herbal classic and has a long application history. Studies have shown that SCF has anti-inflammatory, protective liver, antioxidant, antibacterial and other pharmacological effects. Ancient prescriptions are commonly used in the treatment of chronic diarrhea and other intestinal diseases and diabetes. Modern clinical phar?macology features of SCF polysaccharide (SCFP) in diabetes, liver diseases, enteritis and other aspects have achieved excellent results. Gut is an important digestive organ of human body, but intestinal diseases are varied, including Crohn's disease, ulcerative colitis, intestinal flora imbalance, etc.. It is a chronic and non-specific inflammatory disease. The disease is persisted for a long time and the incidence rate is expected to rise. Most of the symptoms are recurrent diarrhea, bloody stool and abdominal pain. It is considered by the World Health Organization as a refractory disease. At present, there is little possibility of complete cure, which is closely related to complex environmental factors, eating hab?its and heredity. In recent years, clinical studies have found that SCFP has a variety of pharmacological effects on intes?tinal protection.①Reduce inflammatory factors:intestinal mucositis is a common adverse reaction in patients with chemo?therapy. The development of mucositis is related to pro-inflammatory factors such as tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-1β, Interferon-γ(IFN-γ). SCFP can significantly reduce IL-6 TNF-α, IL-1β, and IL-8, as well as the accumulation of T cells in the process of resisting apoptosis, reduce the inflammatory reaction and protect the dam?age to villi and crypts, improve the symptoms of small intestinal mucositis caused by weight loss and diarrhea. ② Pro?mote immunoglobulin A secretion: intestinal mucosal immunity is the first line of defense of the body's immune system. Its main antibody is secretory immunoglobulin A, which can destroy and phagocytize microorganisms, bacteria and viruses. SCFP can improve intestinal immunity by increasing the number and activity of T lymphocytes, promoting the secre?tion of secretory immunoglobulin A, and affecting the activity of a variety of cytokines. ③ Regulation of intestinal flora:the flora in the intestine has the functions of auxiliary nutrient absorption, biological antagonism and immune regulation, and can form a natural barrier for the host's intestine. When the human intestinal flora is disordered, probiotics will be greatly reduced, harmful bacteria will proliferate and destroy the intestinal environment. Under these conditions, the intake of SCFP significantly increased the number of beneficial bacteria such as bifidobacteria and lactobacillus, and sig?nificantly decreased the number of conditional pathogens such as enterococcus and escherichia coli, indicating that SCFP can indeed regulate the intestinal disorder caused by lincomycin hydrochloride to a certain extent. This may be because beneficial bacteria in the intestine metabolize polysaccharides produce short chain fatty acids such as lactic acid and acetic acid, which reduces the pH value in the intestine and inhibits the growth of enterococcus and Escherichia coli. In conclusion, SCFP can treat and protect intestinal diseases to a certain extent, which provides a favorable basis for the treatment of intestinal diseases.

Establishment of HPLC Fingerprint of Wine-processed Schisandra chinensis and Analysis of Chemical Pattern Recognition / 中国药房

OBJECTIVE：To establish the fingerprint of wine-processed Schisandra chinensis ，and to conduct cluster analysis and principal component analysis. METHODS ：HPLC method was adopted. The determination was performed on Diamonsil C 18（2） column with mobile phased consisted of methanol-water （gradient elution ）at the flow rate of 1 mL/min. The detection wavelength was set at 250 nm，and the column temperature was 30 ℃；the injection volume was 10 μL. With schisandrol A as the reference peak，HPLC fingerprints of 15 batches of samples were drawn and their similarity were evaluated with Similarity Evaluation System of TCM Chromatographic Fingerprint （2012 edition）. The common peaks were determined. Cluster analysis and principal component analysis were performed by using SPSS 22.0 statistical software. RESULTS ：There were 20 common peaks in 15 batches of samples ，and the similarities were 0.983-0.999；a total of 8 common peaks were identified ，namely 5-hydroxymethyl furfural，schisandrol A ，schisandrol B ，schisantherin A ，schisantherin B ，deoxyschizandrin，γ-schizandrin，pseudo-γ-schizandrin. The results of cluster analysis showed that 15 batches of wine-processed S. chinensis could be clustered into 4 categories. Among them，S1-S4 and S 14 were clustered into one category ，S9-S11 were clustered into one category ，S5，S7-S8，S12-S13 were clustered into one category ，and S 6 and S 15 were clustered into one category. The results of principal component analysis showed that the cumulative variance contribution rate of first four principal component s was 85.381％；the classification results were basically consistent with the results of cluster analysis. Compared with S. chinensis ，5-hydroxymethyl furfural was newly found in S. chinensis after wine-processing ，with high content ；but there was no significant difference in the other chromatographic peaks. CONCLUSIONS：The established HPLC fingerprint is simple and easy to operate ，combined with cluster analysis and principal component analysis ，can be used for quality control of wine-processed S. chinensis decoction pieces.

Identification and characterization of DIR gene family in Schisandra chinensis / 中国中药杂志

Dirigent(DIR) proteins are involved in the biosynthesis of lignin, lignans, and gossypol in plants and respond to biotic and abiotic stresses. Based on the full-length transcriptome of Schisandra chinensis, bioinformatics methods were used to preliminarily identify the DIR gene family and analyze the physico-chemical properties, subcellular localization, conserved motifs, phylogeny, and expression patterns of the proteins. The results showed that a total of 34 DIR genes were screened and the encoded proteins were 156-387 aa. The physico-chemical properties of the proteins were different and the secondary structure was mainly random coil. Half of the DIR proteins were located in chloroplast, while the others in extracellular region, endoplasmic reticulum, cytoplasm, etc. Phylogenetic analysis of DIR proteins from S. chinensis and the other 8 species such as Arabidopsis thaliana, Oryza sativa, and Glycine max demonstrated that all DIR proteins were clustered into 5 subfamilies and that DIR proteins from S. chinensis were in 4 subfamilies. DIR-a subfamily has the unique structure of 8 β-sheets, as verified by multiple sequence alignment. Finally, through the analysis of the transcriptome of S. chinensis fruit at different development stages, the expression pattern of DIR was clarified. Combined with the accumulation of lignans in fruits at different stages, DIR might be related to the synthesis of lignans in S. chinensis. This study lays a theoretical basis for exploring the biological functions of DIR genes and elucidating the biosynthesis pathway of lignans in S. chinensis.

Effects of “Green Removing ”Processing Technology of the Fresh Fruit of Schisandra chinensis after Harvested on the Quality of Medicinal Material / 中国药房

OBJECTIVE：To stud y the effects of “green removing ”processing technology of fresh fruit of Schisandra chinensis after harvested on the quality of medicinal material ，and to provide new ideas for the scientific and rational processing of Chinese medicinal material. METHODS ：Fifteen fresh fruits of S. chinensis were used as samples ，with 3 samples in each group. The sample were processed preliminarily by 5 methods，such as drying at 50 ℃，drying in the sun ，drying at 50 ℃ after“green removing”processing with microwave ，drying at 50 ℃ after“green removing ”processing with blanching ，drying at 50 ℃ after “green removing ”processing with steaming. HPLC fingerprints of 15 batches of dried S. chinensis products were established and similarity evaluation was conducted according to Similarity Evaluation System of TCM Chromatographic Fingerprints （2012 edition）. Cluster analysis was used to evaluate the similarity of dried S. chinensis products with different processing methods. At the same time ，HPLC method was adopted to determine the content changes of seven lignans in dried products ，such as schisandrol A ， schisandrol B ，schisantherin A ，schisantherin B ，schizandrin A ，schisandrin B and schisandrin C. RESULTS ：A total of 7 common peaks were obtained in the fingerprints of 15 batches of dried S. chinensis products. Except that the similarity between the chromatograms of dried samples in the sun and the control fingerprint was relatively low ，the similarities of chromatograms of dried products by other processing methods were greater than 0.900. Cluster analysis showed that 6 samples dried at 50 ℃ after“green removing”processing with microwave and dried at 50 ℃ after“green removing ”processing with blanching were grouped into the first category ；3 samples dried at 50 ℃ after“green removing”processing with steaming were grouped into the second category ；6 samples dried at 50 ℃ and dried in sun were grouped into the third category. The content determination results showed that there was no significant difference in the total content of seven lignans in the samples dried at 50 ℃ and dried in the sun （P＞0.05）. The total contents of seven lignans in the samples dried at 50 ℃ after“green moving ” processing with microwave ，blanching and steaming were significantly higher than those dried at 50 ℃ and dried in sun （P＜0.01）. CONCLUSIONS：The quality of S. chinensis samples dried after “green moving ”processing with microwave and blanching is better than those directly dried in sun and dried in oven.

Establishment of Characteristic Pattern of Vinegar-processed Schisandra chinensis Formula Granules and Content Determination of 5 Components / 中国药房

OBJECTIVE：To establ ish characteristic pattern of vinegar-processed product of Schisandra chinensis formula granules from different habitats ，and to determine the contents of 5 components. METHODS ：HPLC method was adopted. The determination was performed on Agilent ZORBAX SB-C 18 column with mobile phase consisted of acetonitrile-water （gradient elution）at the flow rate of 1.0 mL/min. The column temperature was set at 30 ℃，and detection wavelength was set at 220 nm. The sample size was 10 µL. Using schisandrin as reference ，HPLC characteristic pattern of 19 batches of vinegar-processed S. chinensis formula granules was drawn. The similarity evaluation was performed with Similarity Evaluation System of TCM Chromatographic Fingerprints （2012 edition），and common peaks were confirmed. The contents of schisandrin ，schisandrol， angeloylgomisin H ，schizandrin and deoxyschizandrin were determined by same method. RESULTS ：There were 8 common peaks in 19 batches of vinegar-processed S. chinensis formula granules ，and the similarities were all above 0.996；five of them were identified as schisandrin ，schisandrol，angeloylgomisin H ，schizandrin and deoxyschizandrin ，respectively. The linear range of schisandrin，schisandrol，angeloylgomisin H ，schizandrin and deoxyschizandrin were 0.030-0.380，0.016-0.195，0.009-0.115， 0.006-0.078 and 0.011-0.138 μg（r＞0.999），respectively. RSDs of precision ，stability（24 h）and reproducibility tests were all lower than 2％. Average recoveries were 99.84％，99.54％，99.28％，100.03％，100.27％（RSD＜1.4％，n＝9）. Average contents of five components in 19 batches of samples were in the range of 0.15％-0.36％，0.02％-0.16％，0.02％-0.06％，0.02％-0.08％ and 0.08％-0.17％，respectively；among them ，total contents of five components in sample S 18 and S 19 from Hebei province were relatively high ，while those were relatively low in sample S 16 and S 17（RSD＝42％）；RSD of total content in other samples （S1-S15）was 12％，and was lower than that of Hebei province ；total content of five components were higher in sample from Jilin province. CONCLUSIONS ： Established characteristic pattern and method for the content determination are specific and reproducible，and can be used for the quality evaluation of vinegar-processed S. chinensis formula granules. The total content fluctuation of vinegar-processed product of S. chinensis formula granules from Liaoning ，Jilin and Heilongjiang provinces is lower than that in Hebei province ，and the quality of vinegar-processed Δ 基金项目：国家中药标准化项目（No.ZYB2H-Y-GD-13） ＊主管中药师 ，硕士 。研究方向 ：中药质量标准 。E-mail： S. chinensis formula granules from Jilin province is the best. lzz332@126.com

Effect of schisandra chinensis polysaccharide on serum inflammatory factors in T2DM rats and its mechanism / 吉林大学学报(医学版)

Objective: To observe the effect of Schisandra Chinensis polysaccharide (SCP) on the serum inflammatory factors in the rats with type 2 diabetes mellitus (T2DM) induced by high-fat diet and low dose of streptozotocin (STZ), and to explore its underlying mechanism in the treatment of T2DM. Methods: The male Wistar rats were given high-fat diet and introperieoneally injected with low dose of STZ (30 mg · kg-1) in one time to establish the rat T2DM models. The successful model rats were randomly divided into model group, low dose (25 mg · kg-1)of SCP group, middle dose (50 mg · kg-1) of SCP group and high dose (100 mg · kg-1) of SCP group; there were 10 rats in each group. Another 10 healthy rats were used as normal control group. Eight weeks after the intragastric administration of SCP, oral glucose tolerance test (OGTT) was performed in the rats of various groups. The fasting blood glucose (FBG) and insulin (INS) levels were detected by glucose oxidase method and radioimmunoassay method, respectively, and the insulin sensitivity index (ISI) was calculated. The levels of interleukin-6 (IL-6), C-reactive protein (CRP), interleukin-lβ (IL-lβ), tumor necrosis factor-α (TNF-α) and nuclear factor-κB (NF-κB) in serum of the rats were measured by enzyme linked immunosorbent assay (ELISA) method. HE staining was used to observe the pathomorphology of pancreas tissue of the rats. Results: Compared with normal control group, the serum FBG level of the rats in model group was significantly increased (P<0.01), and the area under the curve (AUC) of blood glucose of the rats was significantly increased, the serum INS level and the ISI were significantly decreased (P<0.05 or P<0.01); the levels of IL-6, CRP, IL-1β, TNF-α, and NF-κB in serum were all significantly increased (P<0.05 or P<0.01). Compared with model group, the serum FBG levels of the rats in different doses of SCP groups were markedly decreased (P<0.05), the AUC of blood glucose of the rats were significantly decreased, the INS and the ISI levels were significantly elevated (P<0.05); the levels of IL-6, CRP, IL-1β, TNF-α and NF-κB in serum were significantly decreased (P<0.05 or P<0.01). Compared with low dose of SCP group, the FBG levels of the rats in middle and high doses of SCP groups were significantly decreased (P<0.05), the serum INS level of the rats in high dose of SCP group was significantly increased (P<0.05). The HE staining results showed that compared with normal control group, the islets were atrophied, the number of cells in islets was decreased and the boundary was irregular in model group; compared with model group, the islet boundary in different doses of SCP groups became clear, the areas were increased, and the number cells was increased significantly. Conclusion: SCP can decrease the FBG level, increase the INS level and improve insulin resistance (IR) in the T2DM rats induced by high-fat diet combined with low dose of STZ, and its mechanism might be related to its inhibiting inflammation response.

Research progress of Schisandra chinensis and predictive analysis of Q-marker / 中草药

Schisandra chinensis fructus is the dried ripe fruit of S. chinensis from magnoliaceae, produced mainly in the three provinces in the northeast of China. This research systematically summarized the active constituent and pharmacological activity of S. chinensis fructus in recent years, and predicted the Q-marker of S. chinensis fructus in terms of component specificity, constituent validity, component measurability and component absorbed into blood based on the concept of the Q-marker of traditional Chinese medicine. The result suggested the component of lignans in diphenyl cyclooctene as the Q-marker of S. chinensis fructus to conduct the qualitative and quantitative analysis, which provides the scientific basis for establishing and improving the quality evaluation standard of S. chinensis fructus.

Cloning and expression analysis of PLR gene in Schisandra chinensis / 中草药

Objective: To obtain more information for further researches on mechanism of pinoresinol lariciresinol reductases (PLR) gene, which is the key enzyme gene involved with lignans synthesis in Schisandra chinensis, ScPLR gene and its promoter were cloned and analyzed, and the expression pattern of ScPLR gene at fruit development stages was also illustrated. Methods: On the basis of PLR gene sequence obtained by transcriptome sequencing, specific primers were designed, the open reading frame (ORF) of ScPLR gene was then cloned, and the bioinformation of ScPLR gene was analyzed through online software. Meanwhile, the promoter of ScPLR gene was amplified by TAIL-PCR method and analyzed. The expression patterns of ScPLR from fruits at different development stages were analyzed preliminarily. Results: The length of ScPLR gene ORF was 837 bp, which encoded 278 amino acids residues with molecular weight of 31419.85 and theoretical pI of 8.97; ScPLR consisted of a membrane structure, which was a hydrophobic stable protein without signal peptide, and mainly composed of α-helix and random curl; Subcellular localization prediction result showed that ScPLR protein is mainly located in cytoplasm; The results of phylogenetic analysis revealed that ScPLR is closest related to Ricinus communis PLR. The length of ScPLR gene promoter was 994 bp, which had regulatory elements including TATA-box, CAAT-box, also cis-regulatory elements related to light regulation, auxin response, anaerobic induction, defense and stress response, the presence of various cis-acting elements fully reflected the high efficiency and complexity of promoter regulation on gene expression at the transcriptional level. qRT-PCR results showed that ScPLR expression level displayed obvious up-regulation at fruit swelling stage, then down-regulation at fruit coloring period. Conclusion: The ScPLR gene and its promoter were cloned and analyzed, the trend of high expression level of ScPLR before fruit coloring period was consistent with the dynamic change of lignans accumulation in S. chinensis, which will lay foundation for the further research on function and expression regulation of ScPLR gene in lignans biosynthesis pathway.

A systematic strategy for screening therapeutic constituents of (Turcz) Baill infiltrated blood-brain barrier oriented in lesions using ethanol and water extracts: a novel perspective for exploring chemical material basis of herb medicines

, a widely used Chinese herbal medicine, was considered as central nervous system (CNS) drug for years. Both ethanol extracts (EES) and water extracts (WES) of it were applied clinically. Unfortunately, the difference of their efficacy and even effective material foundation of remains obscure. In this study, to explore the active constituents of , we compared pharmacodynamics and chemical profiles / of EES/WES for the first time using multiple chemical analysis, pharmacological and data processing approaches. It was proved that there was no significant difference in the anti-depressive effects between WES and EES. However, the contents of most components and in plasma were higher in EES than those in WES, which was unconvincing for their similar efficacy. Therefore, we further explored components of targeted onto brain and the results showed that 5 lignans were identified with definite absorptivity respectively both in EES and WES caused by the limitation of blood-brain barrier. Moreover, bioinformatic analysis predicted their anti-depressive action. Above all, the systematic strategy screened 5 brain-targeted effective substances of and it was suggested that exploring the components into nidi would promote the studies on herbs effective material basis.

Candidate genes involved in the biosynthesis of lignan in Schisandra chinensis fruit based on transcriptome and metabolomes analysis / 中国天然药物

Schisandra chinensis Turcz. (Baill.) is a plant species with fruits that have been well known in Far Eastern medicine for a long time. It has traditionally been used as a stimulating and fortifying agent in cases of physical exhaustion and to inhibit fatigue. The major bioactive compounds found in S. chinensis are lignans with a dibenzocyclooctadiene skeleton, but little is known about their biosynthesis in plants. S. chinensis is the ideal medicinal plant for studying the biosynthesis of lignans, especially the dibenzocyclooctadiene skeleton. Genomic information for this important herbal plant is unavailable. To better understand the lignan biosynthesis pathway, we generated transcriptome sequences from the fruit during ripening and performed de novo sequence assembly, yielding 136 843 unique transcripts with N50 of 1778 bp. Putative functions could be assigned to 41 824 transcripts (51.57%) based on BLAST searches against annotation databases including GO (Gene ontology) and KEGG (Kyoto Encyclopedia of Genes and Genomes). Furthermore, 22 candidate cytochrome P450 genes and 15 candidate dirigent proteins genes that were most likely involved in the lignan biosynthesis pathway were discovered based on transcriptome sequencing of S. chinensis. The genomic data obtained from S. chinensis, especially the identification of putative genes involved in the lignan biosynthesis pathway, will facilitate our understanding of lignan biosynthesis at the molecular level. The lignan metabolite profiles were analyzed by metabolomes, the accumulation patterns of 30 metabolites involved in the lignan pathway were studied. Co-expression network of lignan contents and transcriptional changes showed 355 strong correlations (correlation coefficient, R > 0.9) between 21 compounds and 153 transcripts. Furthermore, the comprehensive analysis and characterization of the genes involved in lignan pathways and the metabolite profiles of lignans are expected to provide better insight regarding the diversity of the chemical composition, synthetic characteristics, and regulatory mechanisms of this medical herb.

Cloning and expression analysis of 2-oxoglutarate-dependent dioxygenase from Schisandra chinensis / 药学学报

The 2-oxoglutarate-dependent dioxygenase (2-ODD) gene is regarded as the key enzyme gene involved with aryl naphthalene lignan-podophyllotoxin synthesis. To study the expression pattern and function of the Sc2-ODD gene, a full-length cDNA of the gene was cloned. Bioinformatic analysis, the expression pattern, and prokaryotic expression and purification were implemented. The open reading frame of Sc2-ODD gene was 1 077 bp and encoded 358 amino acids with a molecular weight of 40.16 kD. The Sc2-ODD protein contained the conserved 2OG-FeII-oxy sequence of the 2-ODD protein. The results of phylogenetic analysis revealed that Sc2-ODD is most closely related to Corchorus olitorius 2-ODD. qRT-PCR results showed that Sc2-ODD expression displayed obvious up-regulation at the fruit-swelling stage, then down-regulation in the fruit-coloring period. The Sc2-ODD gene was cloned into the bacterial expression vector pGS21T, the recombinant Sc2-ODD protein was expressed in Escherichia coli Rosetta (DE3) cells and the fusion protein was obtained and purified by GST fusion protein purification technology. This study will lay a foundation for further research on the function and expressional regulation of the Sc2-ODD gene in the aryl naphthalene lignans biosynthesis pathway, and also provides a scientific basis for improving the lignan content and the medicinal quality of Schisandra chinensis using plant genetic engineering.