Triterpenoid Saponins from Ardisia gigantifolia and Mechanism on Inhibiting Proliferation of MDA-MB-231 Cells.

Seventeen 13,28-epoxy triterpenoid saponins obtained from Ardisia gigantifolia STAPF. were evaluated their anti-proliferative activities on MCF-7 cells. The structure-activity relationship analysis indicated that CH3 group at C-30, four saccharide units with L-rhamnose at R6 in the sugar units are crucial for the cytotoxic activity on MCF-7. Compounds 1, 2, 6, 7, 12, and 14 were selected to identify the anti-proliferative activity on the other three breast cancer cell lines (T47D, MDA-MB-231 and SK-BR-3). Compounds 2, 6, and 7 with good activity on MCF-7 also showed activity on T47D, MDA-MB-231, and SK-BR-3. Compounds 12 and 14 without cytotoxic activity on MCF-7 almost showed no activities on the other three cell lines. For the triple-negative breast cancer MDA-MB-231, Saponins 7 and 14 showed selective cytotoxic activity, 7 showed much more activity than 14, suggesting the six saccharide units in sugar units and CH3 on C-30 were the key moieties for the anti-proliferative activities. Further molecular mechanism of saponin 7 was studied on inhibiting cell proliferation of MDA-MB-231 cells. Saponin 7 could enhance apoptosis, arrest cell cycles, decrease mitochondrial membrane potentials (MMPs), and considered the involvement of reactive oxygen species (ROS) may explain this conundrum.

Proteomic Analysis of the Antidepressant Effects of Shen-Zhi-Ling in Depressed Patients: Identification of Proteins Associated with Platelet Activation and Lipid Metabolism.

Shen-Zhi-Ling (SZL) is a Chinese medicine formulated from a Kai-Xin-San decoction that is commonly used to treat depression caused by dual deficiencies in the heart and spleen. However, the underlying mechanisms remain unclear. We investigated biological changes in depression patients (DPs) exhibiting antidepressant responses to SZL treatment using proteomic techniques. We performed label-free quantitative proteomic analysis and liquid chromatography-tandem mass spectrometry to discover and examine altered proteins involved in depression and antidepressant treatment. Serum samples were collected from DPs, DPs who underwent 8 weeks of SZL treatment and healthy controls (HCs). The proteins that differed among the three groups were further validated by Western blot analysis. By performing multivariate analyses, we identified 12 potential serum biomarkers that were differentially expressed among the HC, DP, and SZL groups. We then confirmed the significant changes in alpha-1-antitrypsin, von Willebrand factors, apolipoprotein C-III, and alpha-2-macroglobulin among the three groups by performing Western blot analysis, which supported the proteomic results. Profiling the proteomic changes in DPs treated with SZL could improve our understanding of the pathways involved in SZL responses, such as alterations in platelet activation, inflammatory regulation, and lipid metabolism. Future studies involving larger patient cohorts are necessary to draw more definitive conclusions.

Cycloartane triterpenes from Beesia calthaefolia and their anticomplement structure-activity relationship study.

Fifteen cycloartane triterpenes were isolated from Beesia calthaefolia and among them one was new cycloartane triterpenoid. The structure of new compound was determined by the application of spectroscopic analyses and chemical methods. The fifteen compounds were evaluated for their anticomplement activity by classic pathway. The structure-activity relationship analysis indicated that the configurations of 12-OH is preferable to be α than ß, and 18-OH can decrease while 15-OH can increase the anticomplement activity, but saponin with both 15-OH and 18-OH lost most of its activity. The glycosyl moiety of most isolated cycloartane triterpenes is xylosyl. When xylosyl was substituted by glucosyl or galactosyl, their anticomplement activities were decreased or increased, respectively. Further structure-activity relationship (SAR) studies must be carried out to achieve general conclusions regarding the effect of further functionalizations on the anticomplement saponins.

AG4, a compound isolated from Radix Ardisiae Gigantifoliae, induces apoptosis in human nasopharyngeal cancer CNE cells through intrinsic and extrinsic apoptosis pathways.

3ß-O-{α-L-Pyran rhamnose-(1→3)-[ß-D-xylopyranose-(1→2)]-ß-D-glucopyranose-(1→4)-[ß-D-lucopyranose-(1→2)]-α-L-pyran arabinose}-cyclamiretin A (AG4) is a saponin component obtained from the Giantleaf Ardisia Rhizome (Rhizoma Ardisiae Gigantifoliae). The present study aimed to investigate the antitumor potential of AG4 and its possible mechanisms in human nasopharyngeal carcinoma cells (CNE). We exposed tumor cells to AG4 to investigate which cell line was the most sensitive to AG4. Cell viability was assessed using the MTT reduction assay, and the effects of AG4 on apoptosis, reactive oxygen species (ROS) content, mitochondrial membrane potential (MMP), and cell cycle were detected using a flow cytometer; the glutathione, superoxide dismutase and malondialdehyde activities were measured using colorimetric methods. The relative expressions of Bax, Bad, Bid, Bcl-2, and Fas mRNA were calculated using the (Equation is included in full-text article.)comparative method by real-time PCR studies and protein was detected by western blotting. AG4 markedly inhibited the growth of CNE cells by decreasing cell proliferation, inducing apoptosis, and blocking the cell cycle in the S phase. The release of caspase-3, caspase-8, and caspase-9 was stimulated by AG4 in CNE, and the decreased proliferation induced by AG4 was blocked by the inhibitor of pan caspase (Z-VAD-FMK). Moreover, the MMP was decreased in AG4-treated cells, and AG4-induced cell apoptosis was accompanied by a rapid and lasting increase in ROS, which was abolished by N-acetyl-L-cysteine (NAC); glutathione, superoxide dismutase, and malondialdehyde were regulated by AG4. AG4 inhibited Bcl-2 mRNA and protein expression and stimulated Bax, Bad, Bid, Fas mRNA, and protein expression in CNE cultures, suggesting an effect at the transcriptional and protein level. In addition, both the FasL inhibitor (AF-016) and the Bcl-2 family inhibitor (GX15-070) could prevent the cell apoptosis induced by AG4. The findings suggested that AG4-induced apoptosis in CNE cells involved a death receptor pathway and a Bcl-2 family-mediated mitochondrial signaling pathway by decreasing the MMPs in an ROS-dependent manner and regulating genes and proteins relative to apoptosis; also, regulation of cell cycles may also play a role in the antitumor mechanism of AG4.

Biotransformation on the triterpenoid saponin of Ardisia gigantifolia by Aspergillus avenaceus AS 3.4454.

Compound 1, a triterpenoid saponin from Ardisia gigantifolia Stapf. showing potential anti-tumor activity, was transformed into three derivatives (2-4) by Aspergillus avenaceus 3.4454. Among them, compounds 2 and 3 are new compounds. Their structures were elucidated on the basis of 1D NMR, 2D NMR, HR-ESI-MS, and optical rotation data. Compounds 1-3 were evaluated for their cytotoxicity against human hepatocellular carcinoma and normal liver cells by cell counting kit 8 colorimetric assay. Compound 3 displayed better cytotoxicity against Bel-7402 and HepG2 cell lines and much weaker cytotoxicity against normal liver L02 cell than that of positive control (epirubicin hydrochloride).

[Identification of the metabolites of Dingzhi Xiaowan extract in depressive rat plasma, urine, feces and bile after intragastric administration].

Dingzhi Xiaowan is a widely used traditional Chinese medicine in treating depression, which is a similar formula of Kaixinsan. In this research, a rapid ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS(E)) method was established to analyze the metabolites of Dingzhi Xiaowan in depressive model rat plasma, bile, urine and feces. After we established Chronic unpredictable mild stress (CUMS) model rats and orally administrated Dingzhi Xiaowan, rat plasma, bile, urine and feces samples were collected and prepared. Using Waters Cortects UPLC C18 column (2.1 mm x 50 mm, 1.6 µm), acetonitrile-0.1% formic acid mobile phase gradient, these samples were analyzed and 33 metabolites of nine bioactive compounds were detected and tentatively identified by Metabolynx. Among the 33 metabolites, three metabolites were identified from plasma sample, three came from bile sample, and 27 metabolites were identified from urine and feces samples. This approach provided a rapid method for characterizing the metabolites of Dingzhi Xiaowan and gave the truly active structures and the action mechanism of their antidepressant effects.

Antidepressant and neuroprotective effect of the Chinese herb kaixinsan against lentiviral shRNA knockdown brain-derived neurotrophic factor-induced injury in vitro and in vivo.

Depression has been associated with reduced expression of brain-derived neurotrophic factor (BDNF) in the hippocampus. Previous studies have demonstrated that the herbal medicine formula, 'kaixinsan' (KXS), could ameliorate the severity of depression and increase cAMP response element-binding protein expression. There is direct evidence suggesting that the reduction of the BDNF protein in specific brain sites can provoke depressive-like behaviour or affect neurogenesis in vivo. However, the biological mechanisms between the antidepressant and neuroprotective effect of KXS and the alterations in BDNF levels in in vivo and in vitro models remain unclear. Using BDNF knockdown mediated by lentiviral vectors (LV-shBDNF-3) transfected with primary hippocampal neurons and following injection into the dentate gyrus of the hippocampus, it was demonstrated that a reduction in BDNF expression affects cell viability and animal behaviours associated with depression. During treatment with KXS after the lentiviral shRNA silencing of BDNF in cell and animal, cell viability, body weight, the sucrose preference test (SPT), the open field test (OFT) the Morris Water Maze (MWM) task and BDNF expression were measured. KXS attenuated LV-shBDNF-3-induced cell death in primary hippocampal neurons and also improved the sucrose intake in SPT, ambulatory response in OFT and learning ability in MWM against LV-shBDNF-3-induced depressive-like syndromes. Moreover, immunoblot analysis confirmed that KXS could reverse LV-shBDNF-induced BDNF reduction either in vitro or in vivo. These findings provide substantial evidence for supporting a neurotrophic hypothesis of depression and specify BDNF targets for potential antidepressant interventions. Moreover, the antagonism between LV-shRNA BDNF knockdown and KXS may depend on multiple compounds with synergistic mechanisms that modulate the different signal transduction networks directly or indirectly, increasing BDNF expression and exerting its neuroprotective and antidepressant-like effects.

Antitumor activity of triterpenoid saponin-rich Adisia gigantifolia extract on human breast adenocarcinoma cells in vitro and in vivo.

The aim of this study was to explore whether the ethanolic extract of Ardisia gigantifolia rhizomes (AGB-5), a traditional herbal medicine from China, could affect the proliferation of human breast adenocarcinoma (MCF-7) cells in vitro and to explore the antitumor effects of AGB-5 in BALB/c mice engrafted with MCF-7 cells. The results showed that AGB-5 markedly inhibited the proliferation of MCF-7 cells with an IC50 value of 11.89±1.12 µg/mL, increased the S phase and decreased the G2/M phase without influence on G1 phase. MCF-7 cells treated with AGB-5 presented a dose-dependent increase of apoptosis compared with the control group. AGB-5 also significantly increased the activity of caspase-3 and -9 in a dose-dependent manner in MCF-7 cells. Furthermore, in an in vivo model, AGB-5 reduced tumor volume, brought back the red blood cell (RBC) and white blood cell (WBC) count near to normal value, enhanced superoxide dismutase and catalase level of MCF-7 bearing mice. This is the first study to verify the antitumor activity of A. gigantifolia in vivo. The results suggest that AGB-5 may have potential beneficial effects against human breast adenocarcinoma.

Methanolysis of triterpenoid saponin from Ardisia gigantifolia stapf. and structure-activity relationship study against cancer cells.

Thirteen 13,28-epoxy triterpenoid saponins were isolated from Ardisia gigantifolia stapf. and one potential anti-tumor saponin was methanolysised by H2SO4 to afford four new compounds. The seventeen compounds were evaluated for their anti-proliferative activity on A549, HCT-8 and Bel-7402 cells. The structure-activity relationship analysis indicated that the incorporation of =O group at C-16, L-rhamnose at R(5) and acetyl group at OH-6 of the D-glucose lead to a significant increase of the cytotoxic activity on A549 and HCT-8 but significant reduction of the cytotoxic activity on Bel-7402 cells. The synthesized saponins losing 13,28-epoxy and CHO at C-30, losed their cytotoxicities on A549 and HCT-8 cells, suggesting that the two moieties play an essential role for activity. 3ß-O-α-L-rhamnopyranosyl-(1→3)-[ß-D-xylopyranosyl-(1→2)]-ß-D-glucopyranosyl-(1→4)-[ß-D-glucopyranosyl-(1→2)]-α-l-arabinopyranoside-16α-hydroxy-13,28-epoxy-oleanane (2) showed better inhibitory activity to Bel-7402 (IC50 0.86 µM) than that of 5-FU (IC50 8.30 µM), which indicate that five saccharide and methyl moiety at C-30 are important for anti-proliferative activity. The activities of saponins 15>14, 17>16, suggested that the configuration of 28,30-epoxy is preferable to be 30(R) rather than 30(S) on Bel-7402 cells. Further molecular mechanism studies of saponins 1 and 2 were carried out on the cell cycle distribution of Bel-7402 cells.

A new triterpenoid saponin from Ardisia gigantifolia.

A new triterpenoid saponin, named 3-O-ß-d-glucopyranosyl-(1 â†’ 3)-ß-d-xylopyranosyl-(1 â†’ 2)-[α-l-rhamnopyranosyl-(1 â†’ 3)]-ß-d-glucopyranosyl-(1 â†’ 4)-[ß-d-glucopyranosyl-(1 â†’ 2)]-α-l-arabinopyranosyl-3ß,16α,28,30-tetrahydroxy-olean-12-ene (1), along with four known triterpenoids (2-5), was isolated from the rhizomes of Ardisia gigantifolia. Their structures were elucidated by spectroscopic methods. Compounds 1-4 showed cytotoxic activity against Hela, EJ, BCG, and HepG-2 cell lines. The percentage of early apoptotic cells after treatment with 1 was significantly increased compared with control cells (p < 0.05).

Chemical compositions of Souliea vaginata (Maxim) Franch rhizome and their potential therapeutic effects on collagen-induced arthritis in rats.

ETHNOPHARMACOLOGICAL REVEVANCE: Rheumatoid arthritis (RA) is a global prevalent chronic autoimmune inflammatory disease and acceptable safety drugs are lack for its treatment. The rhizomes of Souliea vaginata (Maxim) Franch (SV) possess anti-inflammatory functions and are used as substitution of Coptis chinensis Franch. SV is also traditional Chinese medicine and Tibetan medicine for the treatment of conjunctivitis, enteritis and rheumatic. For searching complementary and alternative anti-RA drugs, it is necessary to characterize the potential anti-arthritic activity of SV and underlying mechanism involved. AIM OF THE STUDY: The aim of the study was to test the chemical compositions, evaluate the anti-arthritic effects and underlying mechanisms of SV. MATERIALS AND METHODS: The chemical compositions of SV were analyzed using liquid chromatography-ion trap-time of flight tandem mass spectrometry (LCMS-IT-TOF). From day 11 to day 31, SV (0.5, 1.0 and 1.5 g/kg body weight) and Tripterygium glycosidorum (TG, 10 mg/kg body weight) were administered orally to the CIA model rats once a day. Thickness of paw and body weights were measured once every two days from day 1 to day 31. Histopathological changes were measured using hematoxylin-eosin (HE) staining. Effects of SV on the levels of IL-2, TNF-α, IFN-γ, IL-4 and IL-10 in serum of CIA rats were measured by enzyme-linked immunosorbent assay (ELISA) kits. CD3+, CD4+, CD8+ and CD4+CD25+ T cells populations were measured using flow cytometric analysis. To evaluate the possible hepatotoxicity and nephrotoxicity, the serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea (UREA) and creatinine (CREA) in CIA rats were also tested using blood auto analyzer. RESULTS: 34 compounds were identified from SV based on LCMS-IT-TOF, and triterpenoids are major anti-arthritic compositions. SV significantly relieved CIA rats' paw swelling without obvious influence on the body weight growth. SV decreased the serum levels of IL-2, TNF-α and IFN-γ in CIA rat, and increased the serum levels of IL-4 and IL-10. SV significantly increased and decreased the percentages of CD4+ and CD8+, with no significant effects on CD3+ in lymphocytes of CIA model rats. Moreover, SV simultaneously decreased thymus and spleen indexes and no hepatotoxicity and nephrotoxicity was observed after short-term treatment. CONCLUSION: These findings suggest that SV possesses preventive and therapeutic effect on RA by modulating the inflammatory cytokines, T-lymphocyte, thymus and spleen indexes and shows no hepatotoxicity and nephrotoxicity.

Synthesis and inhibitory effect of piperine derivates on monoamine oxidase.

A series of piperine derivates (1-19) have been designed, synthesized and evaluated in vitro for their monoamine oxidase (MAO) A and B inhibitory activity and selectivity. It is worth noting that most of the small amine moieties substituted on the piperidine ring proved to be potent and selective inhibitors of MAO-B rather than of MAO-A. 5-(3,4-methylenedioxyphenyl)-2E,4E-pentadienoic acid n-propyl amide (3) showed the greatest MAO-B inhibitory activity (IC(50)(MAO-B)=0.045 µM) and good selectivity (IC(50)(MAO-A)=3.66 µM). The conjugated double bond and carbonyl group of piperine are proved to be an essential feature for piperine and related alkylamides to exhibit MAO-inhibitory activity. Binding mode of the titled compounds was predicted using FlexX algorithm. The design and optimization of novel small molecule monoamine oxidase inhibitors will be guided by the results of this report.

Cytotoxic triterpenoid saponins from Ardisia gigantifolia.

Three new triterpenoid saponins, 1- 3, together with two known saponins, 4 and 5, were isolated from the rhizome of Ardisia gigantifolia. Their structures were elucidated by 1D and 2D NMR spectroscopic studies. Saponins 1, 2, 4, and 5 exhibited significant cytotoxicity against four human cancer cell lines, namely, Hela human cervical carcinoma cells, EJ human bladder tumor cells, HepG-2 human hepatoma cells, and BCG human gastric carcinoma cells with IC (50) values in the range of 1.9-4.8 µM.

Antidepressant Mechanism of Kaixinsan and Its Active Compounds Based on Upregulation of Antioxidant Thioredoxin.

Objectives: Kaixinsan (KXS), a traditional Chinese medicine formula, has been demonstrated to be effective in the treatment of depression. The present study applied a network pharmacology approach to dig out the new targets and mechanism of action of KXS and the active compounds in the treatment of depression. Methods: A network pharmacology approach based on public databases including ADME (absorption, distribution, metabolism, and excretion) evaluation, targets prediction, construction of networks, and molecule docking was used and validated the predicted new antioxidant targets and mechanisms in vitro. Based on an in vitro experiment, we verified the AKT1/Nrf2 pathway related to the thioredoxin (Trx) antioxidant mechanism. Results: The present study sorted 31 pharmacologically active components (kaempferol, ginsenoside rh2, ginsenoside rh4, stigmasterol, etc.) through the ADME algorithm from KXS. 136 potential molecular targets (AKT1, TNF, IL-1b, JUN, ESR1, NOS3, etc.) were predicted, of which there were 69 targets clearly related to depression. By compound-depression targets (C-DTs) network constructed, and protein-protein interaction networks (PPI) and KEGG pathway enrichment analyzed, we identified active compounds mediating depression-related targets to exert synergism on the predictive AKT1/Nrf2 pathway related to thioredoxin (Trx) antioxidant mechanism and other inflammation-related signaling pathways as well as neurotransmitter related signaling pathways. In the H2O2 induced SH-SY5Y cell damage model, this showed kaempferol and ginsenoside rh2 could enhance the activity of the Trx system by upregulation of AKT1 to activate Nrf2 in vitro. Conclusions: Taken together, by comprehensive systems pharmacology approach analysis, we found that KXS and its active compounds might exhibit antioxidant effects by stimulating the AKT1/Nrf2 pathway in the treatment of depression, which might shed new light on innovative therapeutic tactics for the new aspects for depression in traditional Chinese medicine in future studies.

[Study on chemical constituents of rhizome of Ardisia gigantifolia].

OBJECTIVE: To study the chemical constituents of the dried rhizome of Ardisia gigantifolia. METHOD: The 60% ethanol extract was extracted with EtOAc, and then separated and purified by column chromatography using silica gel and preparative HPLC. Their structures were identified on the basis of spectral analysis and physico-chemical properties. RESULT: Nine compounds were isolated and identified as 11-O-galloylbergenin (1), 11-O-syringylbergenin (2), 11-O-protocatechuoylbergenin (3), 4-O-galloylbergenin (4), 11 -O-vanilloylbergenin (5), (-) -epicatechin-3-gallate (6), stigmasterol-3-O-beta-D-glucopyranoside (7), (-) -4'-hydroxy-3-methoxyphenyl-beta-D-[6-O-(4"-hydroxy-3", 5"-dimethoxybenzoyl)] -glucopyranoside (8), and beta-sitosterol (9). CONCLUSION: Compounds 3, 4 and 7 were isolsted from the genus Ardisia for the first time, while compounds 1, 2, 5 and 6 were isolated from this plant for the first time.

Immunosuppressive activity of a cycloartane triterpene glycoside from Beesia calthaefolia by inhibiting T cell proliferation.

BC-1 is a cycloartane triterpene glycoside isolated from the whole plant of Beesia calthaefolia. Our recent studies proved that BC-1 inhibited proliferation of splenic lymphocyte and phagocytosis of macrophages, and inhibited the increased production of TNF-α and IL-1ß. However, it lacks of study about the immunomodulatory effect of BC-1 on purified T lymphocytes. Therefore, in the present study, we evaluated the suppressive potentials of BC-1 on immune responses in vitro. BC-1 markedly suppressed anti-CD3/CD28 mAbs (mAbs) induced murine T lymphocytes proliferation, the expression levels of CD69 and CD25 of CD3+ T cells. BC-1 could strongly decrease ratio of CD4+/CD8+, decrease the Th1/Th2 cytokines production (IL-2, IFN-γ, IL-4, and IL-10) of CD4+ T-cells. In addition, we studied signal transduction pathways about T-cell activation on puried murine CD4+ T lymphocytes by western-blot assay. The data revealed that BC-1 could inhibit the activation of JNK, ERK and PI3K/AKT signal transduction pathways. These results indicated that BC-1 possesses potential downregulating effect on the immune system and might be developed as an immunosuppressive agent in treatment of CD4+ T cell-mediated inflammatory and undesired immune responses.

Triterpenoid saponins from Ardisia gigantifolia.

Four new triterpenoid saponins (1-4) were isolated from the rhizome of Ardisia gigantifolia STAPF. The structures of new saponins were established as 3beta-o-alpha-L-rhamnopyranosyl-(1-->3)-[beta-D-xylopyranosyl-(1-->2)]-beta-D-glucopyranosyl-(1-->4)-[beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl-16alpha-hydroxy-13,28-epoxy-oleanane (1), 3beta-o-alpha-L-rhamnopyranosyl-(1-->3)-[beta-D-xylopyranosyl-(1-->2)]-beta-D-glucopyranosyl-(1-->4)-[beta-D-glucopyrano-syl-(1-->2)]-alpha-L-arabinopyranosyl-16alpha-hydroxy-13,28-epoxy-30-acetoxy-oleanane (2), 3beta-o-alpha-L-rhamnopyranosyl-(1-->3)-[beta-D-glucopyranosyl-(1-->4)-beta-D-xylopyranosyl-(1-->2)]-beta-D-glucopyranosyl-(1-->4)-[beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl-16alpha-hydroxy-13,28-epoxy-oleanane (3) and 3beta-o-alpha-L-rhamnopyranosyl-(1-->3)-[beta-D-xylopyranosyl-(1-->2)]-beta-D-glucopyranosyl-(1-->4)-[beta-D-6-O-acetylglucopranosyl-(1-->2)]-alpha-L-arabinopyranosyl-16alpha-hydroxy-13,28-epoxy-oleanane (4) were isolated from Ardisia gigantifolia STAPF. Their structures were elucidated by means of (1)H- and (13)C-NMR spectroscopic studies, including 2D-NMR technique. The cytotoxic activities of saponins 1-4 are reported against three human cancer cell lines, namely, Hela human cervical carcinoma cells, EJ human bladder tumor cells, and BCG-823 human gastric carcinoma cells.

Antioxidant activity of oligosaccharide ester extracted from Polygala tenuifolia roots in senescence-accelerated mice.

The constituents of the ethanol extract from the root of Polygala tenuifolia Willd. (Polygalaceae) were investigated for antioxidant activity in senescence-accelerated mice. Consequently, two relevant samples were obtained, a fraction separated by macroporous resin (YZ-OE), and a major pure crystal of 3,6'-disinapoyl sucrose (DISS). Based on HPLC-ESI-MS analysis, the most constituents in the YZ-OE fraction from the extract of P. tenuifolia were oligosaccharide esters. The antioxidant activities of these two samples were evaluated using the accelerated senescence-prone, short-lived mice (SAMP) in vivo. The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX) were increased significantly in SAMP mice fed oligosaccharide esters (YZ-OE 50 mg/kg) and its constituents (DISS 50 mg/kg). However, the content of malondialdehyde (MDA) was increased in the blood and liver of SAMP mice. But when given YZ-OE, it could be decreased, by 44.3% and 47.5%, respectively, compared with the SAMP model. Results from the analyses indicated that the oligosaccharide esters (YZ-OE) from roots of P. tenuifolia had a high in vivo antioxidant activity.

[Study of immunogenicity of chlorogenic acid-BSA with different number of CGA].

OBJECTIVE: To investigate the immunogenicity of chlorogenic acid-BSA(CGA-BSA) and the foundation for the allergization of CGA-BSA. METHOD: The CGA-BSA with different number of CGA was synthesized to allergize the BALB/c mice and SD rats to get antiserum. The level of IgE and histamine was analyzed by ELISA, and the passive cutaneous anaphylaxis (PCA) test was carried to analyze the antibody titer. RESULT: When the coupling rate of CGA-BSA is 20, the highest IgE and histamine level in BALB/c mice and the antibody titer in PCA are the highest. CONCLUSION: It has better immunogenicity when the coupling rate of CGA-BSA is between 10 to 20, while the CGA-BSA 20 with the best immunogenicity.

Establishment of a human MYH6 compound heterozygous knockout hESC line to model cardiomyopathy and congenital heart defects by CRISPR/Cas9 system.

MYH6 encodes the alpha heavy chain subunit of cardiac myosin. Mutations in MYH6 cause cardiomyopathy and congenital heart defects. However, due to embryonic lethality in MYH6 knockout mice, the precise roles of MYH6 in cardiomyopathy, congenital heart defects and development process remain largely unknown. In this study, we generated a human MYH6 compound heterozygous knockout hESC line using CRISPR/Cas9 technology. The establishment cell line WAe009-A-46 carried a compound heterozygous 2 bp deletion/7 bp deletion in MYH6, expressed pluripotency markers, showed a normal karyotype and exhibited capability to differentiate into the three germ layers in vitro. MYH6 protein was not detectable in WAe009-A-46 line. This cell line provides a useful tool for studying the role of MYH6 in cardiomyopathy and congenital heart defects.