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.

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 Saponin AG8 from Ardisia gigantifolia stapf. Induces Triple Negative Breast Cancer Cells Apoptosis through Oxidative Stress Pathway.

Triple-negative breast cancers (TNBCs) are associated with poor patient survival because of the absence of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) expressions. Our previous studies have shown that the triterpenoid saponin AG8 from Ardisia gigantifolia stapf. inhibits the proliferation of MDA-MB-231 cells. In this study, the effects of AG8 were further analyzed in different TNBC cell types: MDA-MB-231, BT-549, and MDA-MB-157 cells. AG8 inhibited the viability of MDA-MB-231, BT-549, and MDA-MB-157 cells in a dose-dependent manner and showed stronger cytotoxicity to African American (AA) and mesenchymal (M) subtypes than Caucasian (CA) and mesenchymal stem-like (MSL) subtypes, respectively. AG8 impaired the uptake of MitoTracker Red CMXRos by the mitochondria of TNBC cells in a dose-dependent manner, and this was recovered by N-acetyl-l-cysteine (NAC). AG8 affected GSH, SOD, and MDA levels of TNBC cells, but different TNBC subtypes had different sensitivities to AG8 and NAC. In addition, we found that AG8 increased the Bax/Bcl-2 ratio and the levels of cytoplasmic cytochrome c and significantly decreased phosphorylation of ERK and AKT in BT549 and MDA-MB-157 cells. AG8 elicited its anticancer effects through ROS generation, ERK and AKT activation, and by triggering mitochondrial apoptotic pathways in TNBC cells. AG8 had selective cytotoxic effects against the AA and M TNBC subtypes and markedly induced MDA-MB-157 (AA subtype) cell apoptosis through pathways that were not associated with ROS, which was different from the other two subtypes. The underlying mechanisms should be further investigated.

Antiangiogenic effects of AG36, a triterpenoid saponin from Ardisia gigantifolia stapf.

AG36 is a triterpenoid saponin from Ardisia gigantifolia stapf. Our recent studies proved that AG36 displayed prominent cytotoxicity against breast cancer cells both in vitro and in vivo. However, whether AG36 has antiangiogenic properties is unknown. Therefore, in the present study, we evaluated the antiangiogenic effect of AG36 and the underlying mechanism. The results indicated that AG36 could significantly inhibit the proliferation, migration and invasion of human umbilical vein endothelial cells (HUVEC). Further antiangiogenic molecular mechanism investigation showed that AG36 significantly suppressed phosphorylated FAK and AKT, and downregulated the expressions of vascular endothelial growth factor (VEGF) and vascular endothelial growth factor receptor 2 (VEGFR2) in HUVECs. PI3K inhibitor (LY294002) and FAK inhibitor (PF562271) pretreatment could markedly enhance AG36-induced inhibition of HUVEC proliferation and p-FAK suppression, respectively. In addition, AG36 inhibited the tumor growth in xenograft model and expressions of p-VEGFR2 and p-Akt in vivo. Molecular docking simulation indicated that AG36 formed hydrogen bonds and hydrophobic interactions within the ATP binding pocket of VEGFR2 kinase domain. The present study firstly revealed the high antiangiogenic potency and related underlying molecular of AG36, demonstrating that AG36 maybe a potential antiangiogenic cancer therapy agent or lead candidate.

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.

AG36 Inhibits Human Breast Cancer Cells Proliferation by Promotion of Apoptosis In vitro and In vivo.

AG36 is the biotransformation product of triterpenoid saponin from Ardisia gigantifolia stapf. In this study, the antitumor activity and underlying molecular mechanisms of AG36 against human breast MCF-7, MDA-MB-231, and SK-BR-3 cancer cells were investigated. AG36 inhibited the viability of MCF-7, MDA-MB-231, and SK-BR-3 cells in a dose and time-dependent manner, with an IC50 of approximately 0.73, 18.1, and 23.4 µM at 48 h, respectively. AG36 obviously induced apoptosis and G2/M arrest of all the three breast cancer cells. Moreover, AG36 decreased the protein expression of cycle regulatory proteins cyclin B1 or cyclin D1. In MCF-7 and MDA-MB-231 cells, AG36 strongly increased the cleaved caspase-3 and -8 protein expressions, while in SK-BR-3 cells, AG36 only increased the protein expression of cleaved caspase-3. In all the three breast cancer cells, the ratio of Bax/Bcl-2 and cytosolic cytochrome c content increased significantly compared with control group. The death receptor-related proteins Fas/FasL, TNFR1, and DR5 were detected by Western blot, it showed that different breast cancer cells activated the death receptor-mediated extrinsic caspase-8 pathway through different receptors. In addition, the caspase-8 inhibitor z-IETD-fmk could significantly block AG36-triggered MCF-7 cells apoptosis. The in vivo studies showed that AG36 significantly inhibited the growth of MCF-7 xenograft tumors in BALB/c nude mice comparing with control. In conclusion, AG36 inhibited MCF-7, MDA-MB-231, and SK-BR-3 cells proliferation by the intrinsic mitochondrial and the extrinsic death receptor pathways and AG36 might be a potential breast cancer therapeutic agent.

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.

UPLC-Q-TOF-MS(E) analysis of the constituents of Ding-Zhi-Xiao-Wan, a traditional Chinese antidepressant, in normal and depressive rats.

Ding-Zhi-Xiao-Wan (DZXW) is a traditional Chinese medicine widely used for treating depression. To clarify the bioactive constituents of DZXW, a new rapid ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS(E)) method was established in this study, with the whole extract of the formula separated into multiple components to facilitate the analytical process. In total, 97 compounds were detected and 88 were identified in DZXW. Based on their exact masses, fragmentation characteristics, and retention times, 85 of the 88 compounds were confirmed either conclusively or tentatively, and three potentially novel compounds were identified. In addition, following a three-day oral administration of DZXW, 60 and 28 compounds were observed in the plasma of normal and depressive rats, respectively. Finally, by combining our data with pharmacological information, 10 compounds were predicted as the likely bioactive constituents of DZXW as an antidepressant agent. Our approach provided a rapid method for characterising the chemical constituents of DZXW, and we were the first to screen for bioactive indexes in the plasma of depressive rats. Furthermore, our results provide useful chemical information that could be employed for further study of the pharmacodynamic material basis of DZXW's antidepressant effects.

One new cycloartane triterpene glycoside from Beesia calthaefolia.

One new cycloartane triterpene glycoside (1) was isolated from the whole plant of Beesia calthaefolia. Its structure was elucidated on the basis of extensive spectroscopic data analysis. Its inhibitory effect was measured by the classical pathway of the complement system, and compared with those of known related cycloartane glycosides 2 and 3, previously isolated by us from the same plant. Compounds 1 and 2 exhibited inhibitory activity of complement system with IC50 of 395.3 and 214 µM, respectively. The results suggested that OH at C-12, C-18 and C-15 along with the polarity could affect the inhibitory activity.

[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.

Two new triterpenoid saponins obtained by microbial hydrolysis with Alternaria alternata AS 3.6872.

Compound 1, a triterpenoid saponin from Ardisia gigantifolia Stapf showing potential anti-tumour activity, was hydrolysed into two deglycosyl derivatives (2 and 3) by Alternaria alternata AS 3.6872. Both these derivatives are new compounds. Their structures were elucidated on the basis of 1D, 2D NMR, HR-ESI-MS and optical rotation spectral data. Compounds 1-3 were evaluated for their cytotoxicity against human hepatocellular carcinoma and normal liver cells by Cell Counting Kit 8 colorimetric assay.

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).

Effect of Tenuifoliside A isolated from Polygala tenuifolia on the ERK and PI3K pathways in C6 glioma cells.

Tenuifoliside A (TFSA) is a bioactive oligosaccharide ester component of Polygala tenuifolia Wild, a traditional Chinese medicine which was used to manage mental disorders effectively. The neuroprotective and anti-apoptotic effects of TFSA have been demonstrated in our previous studies. The present work was designed to study the molecular mechanism of TFSA on promoting the viability of rat glioma cells C6. We exposed C6 cells to TFSA (or combined with ERK, PI3K and TrkB inhibitors) to examine the effects of TFSA on the cell viability and the expression and phosphorylation of key proteins in the ERK and PI3K signaling pathway. TFSA increased levels of phospho-ERK and phospho-Akt, enhanced release of BDNF, which were blocked by ERK and PI3K inhibitors, respectively (U0126 and LY294002). Moreover, the TFSA caused the enhanced phosphorylation of cyclic AMP response element binding protein (CREB) at Ser133 site, the effect was revoked by U0126, LY294002 and K252a. Furthermore, when C6 cells were pretreated with K252a, a TrkB antagonist, known to significantly inhibit the activity of brain-derived neurotrophic factor (BDNF), blocked the levels of phospho-ERK, phospho-Akt and phosphor-CREB. Taking these results together, we suggested the neuroprotection of TFSA might be mediated through BDNF/TrkB-ERK/PI3K-CREB signaling pathway in C6 glioma cells.

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.

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.

Cycloartane-type triterpene glycosides from Beesia calthaefolia and their anticomplement activity.

Two new 9,19-cycloartane triterpenes (1, 2), together with three known compounds (3-5), were isolated from the whole plant of Beesia calthaefolia. Their structures were determined by the application of spectroscopic analyses and chemical methods. Compound 1 showed moderate anticomplement activity similar to that of the positive control (rosmarinic acid), while compounds 3 and 4 showed weak activity. The results suggest that 6'-O-(4″-hydroxy-3″-methoxy-benzoyl)-ß-D-glucosyl of 9,19-cycloartane-type triterpenoids could be a structure that is essential for anticomplement activity.

Effects of (20S*,24R*)-epoxy-9,19-cyclolanstane-3ß,12ß,16ß,25-pentaol-3-O-ß-D-xylopyranoside extracted from rhizoma Beesia on immunoregulation and anti-inflammation.

(20S*,24R*)-epoxy-9,19-cyclolanstane-3ß,12ß,16ß,25-pentaol-3-O-ß-D-xylopyranoside (BC1) is a kind of natural bioactive substance extracted from Beesia calthaefolia (Maxim.)Ulbr. This study was designed to evaluate the effects of BC1 on the proliferation of lymphocytes, phagocytosis of peritoneal macrophage, and cytokine secretion, such as tumor necrosis factor (TNF)-α and interleukin (IL)-1ß, and the foot pad thickness index, which is beneficial for understanding the mechanism of BC1 on immunoregulation and anti-inflammation and also will benefit our further research. The proliferation of splenic lymphocyte induced by mitogen (concanavalin A or lipopolysaccharide (LPS)) was detected using the cell counting kit assay. The neutral red phagocytic test of macrophages was determined by colorimetric method. The gene and protein expressions of TNF-α and IL-1ß were measured by real time RT-PCR and ELISA in serum, spleen, and lymphocytes, respectively. In vitro, our present study has shown that BC1 (31.25-250 µg/ml) could inhibit the proliferation of splenic lymphocyte and phagocytosis of macrophages, and inhibit the increased production of TNF-α and IL-1ß in protein and gene levels. In mice, LPS could increase the gene and protein expressions of TNF-α and IL-1ß, respectively, but BC1 (12.5-50 µg/kg) could recover the increased gene and protein expressions of TNF-α and IL-1ß induced by LPS in the spleen and serum of mice. Treatment of arthritic rats with BC1 (1.5 mg/kg body weight) resulted in a significant reduction in foot pad thickness index and serum TNF-α level comparable to the indomethacin-treated arthritic rats, proving its anti-inflammatory effect. Thus, the function of immunoregulation of BC1 may be accomplished through modulating the gene and protein expressions of TNF-α and IL-1ß.

Cycloartane triterpenes from Beesia calthaefolia (Maxim.).

Three new cycloartane triterpenoids (1-3) and two known compounds (4, 5) were isolated from the whole plant of Beesia calthaefolia. Their structures were elucidated by 1D and 2D NMR, HRESIMS and optical rotation spectral data. All isolates were investigated for their inhibitory effects on the classical pathway of the complement system. Among them, compound 4 showed stronger inhibitory activity (IC50 136.7 µM) than positive control (Rosmarinic acid, IC50 181.8 µM) while compounds 2 and 3 were moderately active with IC50 value of 206 µM and 200.9 µM. Chemical compound studied in this article: Rosmarinic acid (PubChem CID: 5281792).