In vivo detection of the epimer metabolites of sweroside via ultra-performance liquid chromatography time-of-flight mass spectrometry combined with DNPH derivatization.

The metabolites of sweroside were first investigated in vivo with ultra-performance liquid chromatography time-of-flight mass spectrometry (UPLC-TOF-MS) in combination with 2,4-dinitrophenylhydrazine derivatization. In addition, the mass detection sensitivity of the major metabolites, epinaucledal and naucledal, via UPLC-TOF-MS was significantly enhanced, and the epimer metabolites were distinctly discovered from plasma following gavage of sweroside in rats. The plasma concentration of epinaucledal and naucledal was quantified via UPLC-TOF-MS in negative mode using erythrocentaurin as the internal standard. The maximum mean plasma concentrations of naucledal and epinaucledal were 75.36 ± 20.10 and 43.52 ± 15.60 ng/ml within 2 h, respectively, following gavage of sweroside at 20 mg/kg. Moreover, the area under the concentration-time curve of naucledal was three times that of epinaucledal. The metabolic process of conversion of sweroside to epinaucledal and naucledal was deduced, and the pharmacological effects of epinaucledal and naucledal will clarify the clinical efficacy of sweroside.

Sweroside alleviates pressure overload-induced heart failure through targeting CaMK⠡δ to inhibit ROS-mediated NF-κB/NLRP3 in cardiomyocytes.

Ongoing inflammation in the heart is positively correlated with adverse remodeling, characterized by elevated levels of cytokines that stimulate activation of cardiac fibroblasts. It was found that CaMKIIδ response to Ang II or TAC triggers the accumulation of ROS in cardiomyocytes, which subsequently stimulates NF-κB/NLRP3 and leads to an increase in IL-6, IL-1ß, and IL-18. This is an important causative factor in the occurrence of adverse remodeling in heart failure. Sweroside is a biologically active natural iridoids extracted from Lonicerae Japonicae Flos. It shows potent anti-inflammatory and antioxidant activity in various cardiovascular diseases. In this study, we found that sweroside inhibited ROS-mediated NF-κB/NLRP3 in Ang II-treated cardiomyocytes by directly binding to CaMKIIδ. Knockdown of CaMKⅡδ abrogated the effect of sweroside regulation on NF-κB/NLRP3 in cardiomyocytes. AAV-CaMKⅡδ induced high expression of CaMKⅡδ in the myocardium of TAC/Ang II-mice, and the inhibitory effect of sweroside on TAC/Ang Ⅱ-induced elevation of NF-κB/NLRP3 was impeded. Sweroside showed significant inhibitory effects on CaMKIIδ/NF-κB/NLRP3 in cardiomyocytes from TAC/Ang Ⅱ-induced mice. This would be able to mitigate the adverse events of myocardial remodeling and contractile dysfunction at 8 weeks after the onset of the inflammatory response. Taken together, our findings have revealed the direct protein targets and molecular mechanisms by which sweroside improves heart failure, thereby supporting the further development of sweroside as a therapeutic agent for heart failure.

Sweroside attenuates podocyte injury and proteinuria in part by activating Akt/BAD signaling in mice.

In this study, we investigated the effects of sweroside on podocyte injury in diabetic nephropathy (DN) mice and elucidated its molecular mechanisms. We conducted in vivo experiments using a C57BL/6 mice model of DN to explore the effects of sweroside on proteinuria and podocyte injury in DN mice. In in vitro experiments, conditionally immortalized mouse podocytes were treated with high glucose and sweroside, and the protective effects of sweroside on podocyte injury were analyzed. In vitro, Akt/BAD pathways were detected using gene siRNA silencing assays and found to be involved in the protective roles of sweroside in high glucose-mediated podocyte injury. In vivo, sweroside significantly decreased albuminuria in DN mice (p < 0.01). periodic acid-Schiff staining showed that sweroside alleviated the glomerular volume and mesangium expansion in DN mice. Consistently, western blot and reverse transcription-polymerase chain reaction analyses showed that the profibrotic molecule expression in the glomeruli declined in sweroside-treated DN mice. Immunofluorescent results showed that sweroside preserved nephrin and podocin expression, and transmission electron microscopy showed that sweroside attenuated podocyte injury. In DN mice, sweroside decreased podocyte apoptosis, and increased nephrin, podocin expression and decreased desmin and HIF1α expression. These results confirmed that sweroside ameliorated albuminuria, glomerulomegaly, and glomerulosclerosis in these mice. Experiments in vitro revealed that sweroside improved HG-induced podocyte injury and apoptosis. Sweroside stimulated activation of the Akt/BAD pathway and upregulated Bcl-2-associated death promoter (BAD) and p-Akt. Overall, sweroside protected podocytes from injury and prevented the progression of DN, providing a novel strategy for the treatment of DN.

Sweroside plays a role in mitigating high glucose-induced damage in human renal tubular epithelial HK-2 cells by regulating the SIRT1/NF-κB signaling pathway.

Sweroside is a natural monoterpene derived from Swertia pseudochinensis Hara. Recently, studies have shown that sweroside exhibits a variety of biological activities, such as anti-inflammatory, antioxidant, and hypoglycemic effects. However, its role and mechanisms in high glucose (HG)-induced renal injury remain unclear. Herein, we established a renal injury model in vitro by inducing human renal tubular epithelial cell (HK-2 cells) injury by HG. Then, the effects of sweroside on HK-2 cell activity, inflammation, reactive oxygen species (ROS) production, and epithelial mesenchymal transition (EMT) were observed. As a result, sweroside treatment ameliorated the viability, inhibited the secretion of inflammatory cytokines (TNF-α, IL-1ß, and VCAM-1), reduced the generation of ROS, and inhibited EMT in HK-2 cells. Moreover, the protein expression of SIRT1 was increased and the acetylation of p65 NF-kB was decreased in HK-2 cells with sweroside treatment. More importantly, EX527, an inhibitor of SIRT1, that inactivated SIRT1, abolished the improvement effects of sweroside on HK-2 cells. Our findings suggested that sweroside may mitigate HG-caused injury in HK-2 cells by promoting SIRT1-mediated deacetylation of p65 NF-kB.

Sweroside alleviates hepatic steatosis in part by activating AMPK/mTOR-mediated autophagy in mice.

In this study, we investigated the effect of sweroside (SOS) on hepatic steatosis in mice and elucidated its molecular mechanisms. We conducted in vivo experiments using a C57BL/6 mice model of nonalcohol fatty liver disease (NAFLD) to explore the effect of SOS on hepatic steatosis in NAFLD mice. In in vitro experiments, primary mouse hepatocytes were treated with palmitic acid and SOS, and the protective effects of SOS on inflammation, lipogenesis, and fat deposition were analyzed. Autophagy-related protein levels and their related signaling pathways were evaluated in both in vivo and in vitro experiments. The results demonstrated that SOS decreased the high-fat-induced intrahepatic lipid content both in vivo and in vitro. The autophagy level in the liver was decreased in NAFLD mice but was reactivated following SOS intervention. SOS intervention was found to partially activate autophagy via the adenosine monophosphate-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) signaling pathway. Consequently, when the AMPK/mTOR pathway was suppressed or autophagy was inhibited, the beneficial effects of SOS intervention on hepatic steatosis were diminished. These results indicate that SOS intervention attenuates hepatic steatosis by promoting autophagy in the liver of NAFLD mice, in part by activating the AMPK/mTOR signaling pathway.

Sweroside: An iridoid glycoside of potential neuroprotective, antidiabetic, and antioxidant activities supported by molecular docking.

Oxidative stress can be a series burden on human health and may lead to many chronic diseases such as diabetes and neurological disorders. The use of natural products to scavenge the reactive oxygen species has attracted the attention of many researchers, to safely manage these conditions with fewer side effects, in available and cost-effective ways. The current study aimed at the isolation and structure elucidation of sweroside from Schenkia spicata (Gentianaceae) and the evaluation of its antioxidant, antidiabetic, neuroprotective, and enzyme inhibitory potential via in vitro and in silico studies. The antioxidant potential was evaluated by a variety of assays as ABTS, CUPRAC and FRAP, showing values of 0.34 ± 0.08, 21.14 ± 0.43, and 12.32 ± 0.20 mg TE/g, respectively, while demonstrating 0.75 ± 0.03 mmol TE/g for phosphomolybdenum (PBD) assay. Acetylcholinestrase (AChE), butyrylcholinesterase (BChE) and tyrosinase inhibitory activities were used to evaluate the neuroprotective effect, while the antidiabetic potential was evaluated by measuring α-amylase and glucosidase inhibitory activities. Results revealed that sweroside showed antioxidant and inhibitory effects on the enzymes tested with the exception of AChE. It demonstrated good tyrosinase inhibitory ability with 55.06 ± 1.85 mg Kojic acid equivalent /g. Regarding the antidiabetic ability, the compound displayed both amylase and glucosidase (0.10 ± 0.01 and 1.54 ± 0.01 mmol Acarbose equivalent/g, respectively) inhibitory activities. Molecular docking studies of sweroside on the active sites of the aforementioned enzymes in addition to NADPH oxidase were performed using Discovery Studio 4.1 software. Results revealed good binding affinities of sweroside to these enzymes mainly through hydrogen bonds and van der Waals interactions. Sweroside can be an important antioxidant and enzyme inhibitory supplement, yet further in vivo and clinical studies are required.

Inhibition of Advanced Glycation End-Products by Tamarindus indica and Mitragyna inermis Extracts and Effects on Human Hepatocyte and Fibroblast Viability.

Tamarindus indica and Mitragyna inermis are widely used by herbalists to cure diabetes mellitus. The aim of this study is to investigate the inhibitory potential of aqueous and various organic solvent fractions from both plants and some isolated compounds against advanced glycation end-products (AGEs). For this purpose, an in vitro BSA-fructose glycation model was used to evaluate the inhibition of AGE formation. Furthermore, the effects of the fractions on mouse fibroblast (NIH-3T3) and human hepatocyte (HepG2) survival were evaluated. The leaf, stem, and root fractions of both plants exhibited significant inhibition of AGEs formation. The IC50 values appeared to be less than 250 µg/mL; however, all fractions presented no adverse effects on NIH-3T3 up to 500 µg/mL. Otherwise, our phytochemical investigation afforded the isolation of a secoiridoid from the Mitragyna genus named secoiridoid glucoside sweroside (1), along with three known quinovic acid glycosides: quinovic acid-3ß-O-ß-d-glucopyranoside (2), quinovic acid-3-O-ß-d-6-deoxy-glucopyranoside, 28-O-ß-d-glucopyranosyl ester (3), and quinovic acid 3-O-α-l-rhamnopyranosyl-(4→1)-ß-d-glucopyranoside (4). In particular, 1-3 are compounds which have not previously been described in Mitragyna inermis roots. However, the isolated compounds did not exhibit AGE inhibitory activity. Further investigation on these potent antiglycation fractions may allow for the isolation of new antidiabetic drug candidates.

Sweroside Ameliorated Memory Deficits in Scopolamine-Induced Zebrafish (Danio rerio) Model: Involvement of Cholinergic System and Brain Oxidative Stress.

Sweroside is a secoiridoid glycoside and belongs to a large group of naturally occurring monoterpenes with glucose sugar attached to C-1 in the pyran ring. Sweroside can promote different biological activities such as antifungal, antibacterial, hepatoprotective, gastroprotective, sedative and antitumor, antioxidant, and neuroprotective activities. Zebrafish were given sweroside (12.79, 8.35, and 13.95 nM) by immersion once daily for 8 days, along with scopolamine (Sco, 100 µM) 30 min before the initiation of the behavioral testing to cause anxiety and memory loss. Employing the novel tank diving test (NTT), the Y-maze, and the novel object recognition test (NOR), anxiety-like reactions and memory-related behaviors were assessed. The following seven groups (n = 10 animals per group) were used: control, Sco (100 µM), sweroside treatment (2.79, 8.35, and 13.95 nM), galantamine (GAL, 2.71 µM as the positive control in Y-maze and NOR tests), and imipramine (IMP, 63.11 µM as the positive control in NTT test). Acetylcholinesterase activity (AChE) and the antioxidant condition of the brains were also evaluated. The structure of sweroside isolated from Schenkia spicata was identified. Treatment with sweroside significantly improved the Sco-induced decrease of the cholinergic system activity and brain oxidative stress. These results suggest that sweroside exerts a significant effect on anxiety and cognitive impairment, driven in part by the modulation of the cholinergic system activity and brain antioxidant action.

Sweroside Effectively Protects Against Ischemia/Reperfusion Injury by Regulating Excitation-contraction Coupling / 中国实验方剂学杂志

ObjectiveTo explore the effect of sweroside on the protection of cardiac systolic/diastolic function during ischemia/reperfusion （I/R） injury. MethodTwenty-four healthy male SD rats were randomly divided into control group， model group， 10 μmol·L-1 sweroside group and 1 μmol·L-1 digoxin group. The I/R injury was modeled by Langendorff and ligation of the left anterior descending coronary artery. The infarct size in each group was measured by 2，3，5-triphenyltetrazolium chloride （TTC） staining and hemodynamic parameters such as left ventricular diastolic pressure （LVDP）， left ventricular end-diastolic pressure （LVEDP）， left ventricular end-systolic pressure （LVESP）， maximum rate of rising of left ventricular pressure （+dp/dtmax） and maximum rate of decreasing of left ventricular pressure （-dp/dtmax） of rat isolated heart were detected by Powerlab. In addition， neonatal rat cardiomyocytes （NRCMs） were isolated and randomly divided into control group， model group， 1 μmol·L-1 sweroside group and 10 μmol·L-1 sweroside group. Hypoxia/reoxygenation （H/R） injury model was established. Cardiac systolic function and calcium transients were examined by multi-functional cell imaging analyzer and laser confocal microscope. Furthermore， real-time polymerase chain reaction（Real-time PCR） was used to verify the mRNA expression of excitation-contraction coupling genes such as L-type calcium channel （Cacnb2）， cytochrome c oxidase subunit 6A2 （Cox6a2）， troponin （Tnnc1， Tnni3， Tnnt2）， actin （Actc1）， and myosin （Myh6， Myl2， Myl4） according to the results of previous transcriptome sequencing and literature investigation. Differentially expressed genes were subjected to cluster analysis. ResultCompared with the conditions in the control group， increased cardiac infarction size （P<0.01） and LVEDP （P<0.01） and decreased LVDP （P<0.01） and LVESP （P<0.05） were observed in the model group， with +dp/dtmax of increasing trend while -dp/dtmax decreasing. Moreover， the cell viability， heart rate and contraction amplitude of NRCMs was reduced （P<0.01）， while the contraction duration， time to peak and relaxation time was elevated （P<0.01） in the model group. Interestingly， sweroside could reverse these indicators （P<0.05）. In addition， the expression of Cacnb2， Cox6a2， Tnnc1， Tnni3， Tnnt2， Actc1， and Myh6， Myl2， and Myl4 was down-regulated in the model group （P<0.05， P<0.01）， but sweroside could up-regulate the expression of the above genes （P<0.05）. ConclusionSweroside effectively regulated Ca2+ level in NRCMs， enhanced cardiac systolic function， and protected against H/R injury by regulating excitation-contraction coupling.

Promotion of osteogenesis by Sweroside via BMP2-involved signaling in postmenopausal osteoporosis.

Phlomis umbrosa has been traditionally used for bone diseases in traditional Korean Medicine. Sweroside (SOS), marker compounds of P. umbrosa, has been known to promote osteoblast differentiation. In this study, ameliorative effects of SOS on osteoporosis and potential target pathway were investigated. Ovariectomized mice were administered three doses of SOS three times a week for 4 weeks after inducing osteoporosis. Bone mineral content (BMC) and bone mineral density (BMD) were analyzed by dual energy X-ray absorptiometry. A human osteosarcoma cell line (SaOS-2) was differentiated to clarify the promoting effects of SOS on osteoblast differentiation and bone formation. Osteoblastic bone-forming markers were evaluated in lumbar vertebrae (LV) and mineralized SaOS-2 cells. SOS markedly elevated BMC and BMD levels and attenuated the bone marrow adipocytes in the femoral shaft. SOS increased the formation of bone matrix in SaOS-2 cells. Bone morphogenetic protein-2 (BMP2) and runt-related transcription factor 2 (CBFA1) in LV and SaOS-2 cells were up-regulated by SOS. SOS increased alkaline phosphatase (ALPL), osteopontin (SPP1), and bone sialoprotein-1 (BSPH1). In conclusion, SOS induced the formation of mineralized bone matrix by regulating BMP2/CBFA1-mediated molecules. Therefore, SOS could be a therapeutic compound of treatment for osteoporosis by producing the new bone matrix.

Sweroside Protects Against Myocardial Ischemia-Reperfusion Injury by Inhibiting Oxidative Stress and Pyroptosis Partially via Modulation of the Keap1/Nrf2 Axis.

Aims: Sweroside, a secoiridoid glucoside extracted from Swertia pseudochinensis Hara, is reported to possess antioxidant and anti-inflammatory activities. However, whether sweroside has a protective effect on myocardial ischemia-reperfusion (IR) injury is yet to be elucidated. The present study aimed to confirm the cardioprotective effect of sweroside and to identify its underlying mechanism. Methods and Results: H9c2 cells were pretreated with sweroside and then underwent hypoxia-reoxygenation. Cell Counting Kit-8, creatine kinase-myocardial band (CK-MB) and lactate dehydrogenase (LDH) assays were conducted to detect cell viability and myocardial injury, respectively. The Langendorff method was used to induce myocardial IR injury ex vivo. Triphenyltetrazolium chloride staining was performed to detect myocardial infarct size, while protein expression was analyzed using western blotting. Overall, the results indicated that sweroside pretreatment dose-dependently led to a significant enhancement in cell viability, a decrease in release of CK-MB and LDH, a reduction in infarct size, and an improvement in cardiac function. Additionally, sweroside pretreatment caused a marked suppression of oxidative stress, as evidenced by the fact that sweroside decreased the accumulation of reactive oxygen species and malondialdehyde, while enhancing the activities of superoxide dismutase and glutathione peroxidase. Moreover, sweroside was found to notably repress pyroptosis, as sweroside blocked pore formation in the cell membrane, inhibited caspase-1 and interleukin (IL)-1ß activity, and decreased the expression levels of NLR family pyrin domain containing 3 (NLRP3), apoptosis-associated speck-like protein containing a CARD, cleaved caspase-1, and IL-1ß. Mechanistically, it was found that sweroside inhibited Kelch-like ECH-associated protein 1 (Keap1) and induced nuclear factor E2-associated factor 2 (Nrf2) nuclear translocation. Furthermore, the inhibition of oxidative stress and pyroptosis by sweroside could be abrogated via the inhibition of Nrf2 expression, which suggested that the protective effect induced by sweroside was Nrf2-dependent. Conclusions: The present study demonstrated that sweroside pretreatment could protect against myocardial IR injury by inhibiting of oxidative stress and NLRP3 inflammasome-mediated pyroptosis partially via modulation of the Keap1/Nrf2 axis.

Anti-inflammatory Effects of Sweroside on LPS-Induced ALI in Mice Via Activating SIRT1.

Sweroside, as one of the main components of Swertia L. in Gentianaceae, has the effect of clearing heat and detoxifying. In previous studies, sweroside has been reported to have anti-inflammatory effect on LPS-induced inflammation by alleviating NF-κB signaling pathway. In this paper, we investigate the anti-inflammatory effects of sweroside by establishing LPS-induced acute lung injury (ALI) model in mice. Experimental results showed that sweroside could reduce the wet-to-dry ratio of the lung and inhibit MPO activity. In addition, it turned out that sweroside reduced pathological changes in lung tissue and the numbers of inflammatory cells. Moreover, sweroside significantly reduced the levels of inflammatory cytokines and down-regulated the NF-κB signaling pathway. And the results demonstrated that sweroside could increase the expression of SIRT1, and the protective effects of sweroside on LPS-induced ALI were reversed by SIRT1 inhibitor EX-527. In conclusion, sweroside can protect LPS-induced ALI mice through inhibiting inflammation.

[Effect of swertiamarin, gentiopicrin and sweroside on cell apoptosis and expression of Bcl-2 in rheumatoid arthritis fibroblast-like synoviocytes].

The aim of this paper was to discuss the effect of swertiamarin, gentiopicrin and sweroside on rheumatoid arthritis fibroblast-like synoviocytes(RA-FLSs) and B-cell lymphoma-2(Bcl-2) and their mechanisms. ZINC database and RCSB PDB database were retrieved for 3 D chemical structures of swertiamarin, gentiopicrin and sweroside and 3 D target protein structures. AutoDock Mgltools 1.5.6, AutoDockVina 1.1.2 and pyMOL 2.2.0 were applied for molecular docking to analyze the relationship between Bcl-2(1 GJH) target protein and important ingredients. The cell apoptosis of RA-FLSs was tested by Annexin V-FITC. The Bcl-2 protein expression of RA-FLSs treated with different ingredients was tested by Western blot. The Bcl-2 mRNA expression of RA-FLSs treated with different ingredients was tested by RT-PCR. Swertiamarin, gentiopicrin and sweroside were docked well with Bcl-2(1 GJH). The binding energy of swertiamarin was-6.9 kcal·mol~(-1), the binding energy of gentiopicrin was-6.7 kcal·mol~(-1) and the binding energy of sweroside was-6.4 kcal·mol~(-1). Compared with the blank group, the Bcl-2 protein expression of each group were reduced, while that of the gentiopicrin group was the highest(P<0.01). Compared with the blank group, the Bcl-2 mRNA expression of each groups were reduced. Gentiopicrin can reduce the Bcl-2 protein expression and the Bcl-2 mRNA expression, so as to promote the RA-FLSs apoptosis.

A new secoiridoid glycoside and other constituents from the roots and flowers of Fagraea fragrans Roxb. (Gentianaceae).

A new secoiridoid glycoside, 3'-O-p-trans-coumaroylsweroside (1), together with twenty-nine compounds (2-30), were isolated from the roots and flowers of Fagraea fragrans Roxb. (Gentianaceae). Their structures were identified by analysis of mass spectrometric and NMR spectroscopic data. Compounds 2, 4, 6, 11 and 13-15 showed weak to moderate cytotoxicity against cholangiocarcinoma cancer cells lines (KKU-213, KKU-055 and KKU-214). We report the first phytochemical investigation of the roots and flowers of F. fragrans, as only the essential oil of the latter has been investigated.

Plumeria rubra L.- A review on its ethnopharmacological, morphological, phytochemical, pharmacological and toxicological studies.

ETHNOPHARMACOLOGICAL RELEVANCE: Plumeria rubra L. (Apocynaceae) is a deciduous, commonly ornamental, tropical plant grown in home premises, parks, gardens, graveyards, because of its beautiful and attractive flowers of various colours and size. The different parts of the plant are used traditionally to treat various diseases and conditions like leprosy, inflammation, diabetic mellitus, ulcers, wounds, itching, acne, toothache, earache, tongue cleaning, pain, asthma, constipation and antifertility. AIM OF THE REVIEW: The main aim of this review is to provide an overview and critically analyze the reported ethnomedical uses, phytochemistry, pharmacological activities and toxicological studies of P. rubra and to identify the remaining gaps and thus supply a basis for further investigations. The review also focuses towards drawing attention of people and researchers about the wide spread pharmaceutical properties of the plant for its better utilization in the coming future. MATERIAL AND METHODS: All the relevant data and information on P. rubra was gathered using various databases such as PubMed, Springer, Taylor and Francis imprints, NCBI (National Center for Biotechnology Information), Science direct, Google scholar, Chemspider, SciFinder, research and review articles from peer-reviewed journals and unpublished data such as Phd thesis, etc. Some other 'grey literature' sources such as webpages, ethnobotanical books, chapters, wikipedia were also studied. RESULTS: More than 110 chemical constituents have been isolated from P. rubra including iridoids, terpenoids, flavonoids and flavonoid glycosides, alkaloids, glycosides, fatty acid esters, carbohydrates, animo acids, lignan, coumarin, volatile oils, etc. The important chemical constituents responsible for pharmacological activities of the plant are fulvoplumierin, plumieride, rubrinol, lupeol, oleanolic acid, stigmasterol, taraxasteryl acetate, plumieride-p-E-coumarate, rubranonoside, rubrajalellol, plumericin, isoplumericin, etc. The plant possess a wide range of pharmacological activities present namely antibacterial, antiviral, anti-inflammatory, antipyretic, antidiabetic, hepatoprotective, anticancer, anthelmintic, antifertility and many other activities. CONCLUSION: P. rubra is a valuable medicinal source and further study in this topic can validate the traditional and ethnobotanical use of the plant. However, many aspects of the plant have not been studied yet. The pharmacological activity of active chemical constituent isolated from the plant is proven only for a couple of activities hence, lack of bio-guided isolation strategies is observed. Further studies on bioavailability, pharmacokinetics, mechanism of action and structural activity relationship studies of isolated pure compounds will contribute more in understanding their pharmacological effects. Higher doses of plant extracts are administered to experimental animals, therefore their toxicity and side effects in humans are needed to be thoroughly studied, although no side effect or toxicity is seen or observed in experimental animals. Studies are also essential to investigate the long term in vivo toxicity and clinical efficacy of the plant.

Effect of swertiamarin, gentiopicrin and sweroside on cell apoptosis and expression of Bcl-2 in rheumatoid arthritis fibroblast-like synoviocytes / 中国中药杂志

The aim of this paper was to discuss the effect of swertiamarin, gentiopicrin and sweroside on rheumatoid arthritis fibroblast-like synoviocytes(RA-FLSs) and B-cell lymphoma-2(Bcl-2) and their mechanisms. ZINC database and RCSB PDB database were retrieved for 3 D chemical structures of swertiamarin, gentiopicrin and sweroside and 3 D target protein structures. AutoDock Mgltools 1.5.6, AutoDockVina 1.1.2 and pyMOL 2.2.0 were applied for molecular docking to analyze the relationship between Bcl-2(1 GJH) target protein and important ingredients. The cell apoptosis of RA-FLSs was tested by Annexin V-FITC. The Bcl-2 protein expression of RA-FLSs treated with different ingredients was tested by Western blot. The Bcl-2 mRNA expression of RA-FLSs treated with different ingredients was tested by RT-PCR. Swertiamarin, gentiopicrin and sweroside were docked well with Bcl-2(1 GJH). The binding energy of swertiamarin was-6.9 kcal·mol~(-1), the binding energy of gentiopicrin was-6.7 kcal·mol~(-1) and the binding energy of sweroside was-6.4 kcal·mol~(-1). Compared with the blank group, the Bcl-2 protein expression of each group were reduced, while that of the gentiopicrin group was the highest(P<0.01). Compared with the blank group, the Bcl-2 mRNA expression of each groups were reduced. Gentiopicrin can reduce the Bcl-2 protein expression and the Bcl-2 mRNA expression, so as to promote the RA-FLSs apoptosis.

Sweroside promotes osteoblastic differentiation and mineralization via interaction of membrane estrogen receptor-α and GPR30 mediated p38 signalling pathway on MC3T3-E1 cells.

BACKGROUND: Dipsaci Radix has been clinically used for thousands of years in China for strengthening muscles and bones. Sweroside is the major active iridoid glycoside isolated from Dipsaci Radix. It has been reported that sweroside can promote alkaline phosphatase (ALP) activity in both the human osteosarcoma cell line MG-63 and rat osteoblasts. However, the underlying mechanism involved in these osteoblastic processes is poorly understood. PURPOSE: This study aimed to characterize the bone protective effects of sweroside and to investigate the signaling pathway that is involved in its actions in MC3T3-E1 cells. METHODS: Cell proliferation, differentiation and mineralization were evaluated by the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay, ALP test and Alizarin Red S staining, respectively. The concentration of sweroside in intracellular and extracellular fluids was determined by ultra-performance liquid chromatography coupled to triple quadrupole xevo-mass spectrometry (UPLC/TQ-XS-MS). Proteins associated with the osteoblastic signaling pathway were analysed by western blot and immunofluorescence methods. RESULTS: Sweroside did not obviously affect the proliferation but significantly promoted the ALP activity and mineralization of MC3T3-E1 cells. The maximal absorption amount 0.465 ng/ml (1.3 × 10-9 M) of sweroside was extremely lower than the tested concentration of 358.340 ng/ml (10-6 M), indicating an extremely low absorption rate by MC3T3-E1 cells. Moreover, the ALP activity, the protein expression of ER-α and G protein-coupled receptor 30 (GPR30) induced by sweroside were markedly blocked by both the ER antagonist ICI 182780 and the GPR30 antagonist G15. In addition, sweroside also activated the phosphorylation of p38 kinase (p-p38), while the phosphorylation effects together with ALP and mineralization activities were completely blocked by a p38 antagonist, SB203580. Additionally, the phosphorylation of p38 induced by sweroside were markedly blocked by both the ER antagonist ICI 182780 and the GPR30 antagonist G15. CONCLUSIONS: The present study indicated that sweroside, as a potential agent in treatment of osteoporosis, might exert beneficial effects on MC3T3-E1 cells by interaction with the membrane estrogen receptor-α and GPR30 that then activates the p38 signaling pathway. This is the first study to report the specific mechanism of the effects of sweroside on osteoblastic differentiation and mineralization of MC3T3-E1 cells.

Sweroside ameliorated carbon tetrachloride (CCl4)-induced liver fibrosis through FXR-miR-29a signaling pathway.

To date, there are very few effective drugs for liver fibrosis treatment; therefore, it is urgent to develop novel therapeutic targets and approaches. In the present research, we sought to study the protective effect of sweroside contained in Lonicera japonica or blue honeysuckle berries in a mouse model of liver fibrosis and investigate the underlying mechanism. The mouse model of liver fibrosis in was induced by intraperitoneal injections of 10% CCl4 for 6 weeks (three times/week). At the beginning of the fourth week, sweroside was intragastrically administered once a day and at the end of the treatment, biochemical and histological studies were investigated. The expression of FXR, miR-29a and the downstream targets were analyzed as well. Moreover, the effect of sweroside on cell proliferation was observed in human hepatic stellate cells (HSCs) (LX-2), along with using the siRNA for FXR and miR-29a inhibitor to investigate the underpinning of the anti-fibrotic effect of sweroside. Sweroside successfully protected the liver fibrosis in CCl4-induced mouse model, accompanied by miR-29a induction. Furthermore, sweroside also induced miR-29a in HSCs, resulting in the inhibition of COL1 and TIMP1. Our data also showed that either silencing miR-29a or knockdown of FXR in LX-2 cell abolished the inhibition of COL1 and TIMP1 as well as the inhibition of cell proliferation by sweroside treatment. In conclusion, sweroside exerted its anti-fibrotic effect in vivo and in vitro by up-regulation of miR-29a and repression of COL1 and TIMP1, which was at least in part through FXR.

Terpenoids from aerial parts of Gendarussa vulgaris / 中草药

Objective: To isolate and identify the terpenoids from the aerial parts of Gendarussa vulgaris. Methods: The 95% EtOH extract of the aerial parts of G. vulgaris were isolated and purified by silica gel, Sephadex LH-20, reversed-phase ODS, macroporous adsorption resin AB-8 and semi-preparative high performance liquid chromatography. The compound structures were identified by physicochemical properties and spectroscopic data. Results: Ten terpenoids were identified as gvterpennoid A (1), 4,4,14α- trimethylpregn-8-en-3β,20α-diol (2), betulone (3), ergosterol endoperoxide (4), ursolic acid (5), oleanolic acid (6), 3β-hydroxyl- 11α,12α-epoxy olean-28,13β-lactone (7), sweroside (8), loganin (9), and dehydromorroniaglycone (10). Conclusion: Compound 1 is a new triterpene, named gvterpennoid A. Compound 2 is a new natural product, and its 13C- and 1H-NMR chemical shifts were first completely assigned on the basis of 1D and 2D NMR spectroscopic evidence. Compounds 3-5, 7-10 are isolated from Gendarussa genus for the first time.

Sweroside-mediated mTORC1 hyperactivation in bone marrow mesenchymal stem cells promotes osteogenic differentiation.

This paper aims to probe into the effect of sweroside (SOS) in osteoporosis (OP) and explains mechanisms of its molecular. Applying the ovariectomized (OVX) mouse model investigates the preventive effect of SOS against postmenopausal OP after 3 months of SOS treatment (120 mg/kg/day). Using hematoxylin and eosin (HE) staining and micro computed tomography (CT) observed the morphology of OP in each group. Immunohistochemical staining (IHC) was used to examine osteoblast markers. Experiments in vitro, bone marrow mesenchymal stem cells (BMSCs) from C57/BL6 mice were treated with SOS for 14 days. The staining of alizarin red and alkaline phosphatase activity were measured, and the presentation of osteoblast markers was detected by quantitative reverse transcription PCR. BMSCs were also treated with 1 µg/mL SOS with or without rapamycin, the expression of protein S6 (PS6), P-mTOR, runt-related transcription factor 2 (RUNX2), OSX, and osteocalcin (OCN) was detected by Western blotting. Experiments in vivo, HE results show that SOS can alleviate OP, CT results show that there are lower trabecular thickness, bone mineral density, and trabecular number in control OVX mice than those in the OVX + SOS group. IHC results showed that SOS can promote the expression of osteogenic markers and immunofluorescent results show that SOS can promote mTORC1 signal activation. Experiments in vitro revealed that SOS stimulated the activation of the mTORC1 signaling pathway and upregulated RUNX2, OSX, and OCN, rapamycin can reverse it. Our findings demonstrated that differentiated BMSCs into osteoblasts can be promoted by SOS via upregulating the expression of P-mTOR, PS6, RUNX2, OSX, and OCN. SOS effectively prevented OP by hyperactivation of the mTORC1/PS6 signaling pathway.