Ophiopogonin D' inhibited tumour growth and metastasis of anaplastic thyroid cancer by modulating JUN/RGS4 signalling.

Anaplastic thyroid cancer (ATC), an aggressive malignancy with virtually 100% disease-specific mortality, has long posed a formidable challenge in oncology due to its resistance to conventional treatments and the severe side effects associated with current regimens such as doxorubicin chemotherapy. Consequently, there was urgent need to identify novel candidate compounds that could provide innovative therapeutic strategies for ATC. Ophiopogonin D' (OPD'), a triterpenoid saponin extracted, yet its roles in ATC has not been reported. Our data demonstrated that OPD' potently inhibited proliferation and metastasis of ATC cells, promoting cell cycle arrest and apoptosis. Remarkably, OPD' impeded growth and metastasis of ATC in vitro and in vivo, displaying an encouraging safety profile. Regulator of G-protein signalling 4 (RGS4) expression was significantly up-regulated in ATC compared to normal tissues, and this upregulation was suppressed by OPD' treatment. Mechanistically, we elucidated that the transcription factor JUN bound to the RGS4 promoter, driving its transactivation. However, OPD' interacted with JUN, attenuating its transcriptional activity and thereby disrupting RGS4 overexpression. In summary, our research revealed that OPD' bound with JUN, which in turn resulted in the suppression of transcriptional activation of RGS4, thereby eliciting cell cycle arrest and apoptosis in ATC cells. These findings could offer promise in the development of high-quality candidate compounds for treatment in ATC.

The Protective Effects of Ruscogenin Against Lipopolysaccharide-Induced Myocardial Injury in Septic Mice.

ABSTRACT: Sepsis-induced myocardial dysfunction commonly occurs in individuals with sepsis and is a severe complication with high morbidity and mortality rates. This study aimed to investigate the effects and potential mechanisms of the natural steroidal sapogenin ruscogenin (RUS) against lipopolysaccharide (LPS)-induced myocardial injury in septic mice. We found that RUS effectively alleviated myocardial pathological damage, normalized cardiac function, and increased survival in septic mice. RNA sequencing demonstrated that RUS administration significantly inhibited the activation of the NOD-like receptor signaling pathway in the myocardial tissues of septic mice. Subsequent experiments further confirmed that RUS suppressed myocardial inflammation and pyroptosis during sepsis. In addition, cultured HL-1 cardiomyocytes were challenged with LPS, and we observed that RUS could protect these cells against LPS-induced cytotoxicity by suppressing inflammation and pyroptosis. Notably, both the in vivo and in vitro findings indicated that RUS inhibited NOD-like receptor protein-3 (NLRP3) upregulation in cardiomyocytes stimulated with LPS. As expected, knockdown of NLRP3 blocked the LPS-induced activation of inflammation and pyroptosis in HL-1 cells. Furthermore, the cardioprotective effects of RUS on HL-1 cells under LPS stimulation were abolished by the novel NLRP3 agonist BMS-986299. Taken together, our results suggest that RUS can alleviate myocardial injury during sepsis, at least in part by suppressing NLRP3-mediated inflammation and pyroptosis, highlighting the potential of this molecule as a promising candidate for sepsis-induced myocardial dysfunction therapy.

Total synthesis and cytotoxicity evaluation of the spirostanol saponin gitonin.

The spirostanol saponin gitonin was efficiently synthesized in 12 steps (longest linear sequence) in 18.5% overall yield from the commercially available isopropyl ß-D-1-thiogalactopyranoside (IPTG) and tigogenin. A cascade two-step glycosylation and Schmidt's inverse procedure significantly facilitated the synthesis of gitonin and its derivatives. The cytotoxic activities of gitonin and its structural analogues were evaluated against A549, HepG2, and MCF-7, and most of them exhibited moderate to excellent inhibitory activity. Our study demonstrates that the removal of the ß-D-galactopyranosyl residue (attached at C-2 of the glucose unit) from gitonin would not decrease the inhibition activities; however, further cleavage of sugar units could seriously reduce the activities. A bioassay on these cancer cell lines also suggested that the presence of 2α-hydroxy on the aglycone weakened the cytotoxicity of the designed saponin.

Tributelosides A-D: Four Undescribed Spirostan Glycosides Isolated from the Branches and Leaves of Tribulus terrestris with Their NO Production Inhibitory Activity in LPS Activated RAW 264.7 Cells.

Four undescribed spirostan glycosides, (25S)-5α-spirostan- 12-one-2α,3ß-diol-3-O-ß-D-glucopyranosyl-(1→4)-ß-D-galactopyranoside (1), (25S)-5α-spirostan-12-one-2α,3ß-diol-3-O-ß-D-galatopyranosyl-(1→2)-ß-D-glucopyranosyl- (1→4)-ß-D-galactopyranoside (2), (25S)-5α-spirostan-12-one-2α,3ß-diol-3-O-ß-D-glucopyranosyl-(1→2)-[ß-D-glucopyranosyl-(1→3)]-ß-D-glucopyranosyl-(1→4)-ß-D-galactopyranoside (3), and hecogenin 3-O-ß-D-glucopyranosyl-(1→3)-[ß-D-xylopyranosyl-(1→2)]-ß-D-glucopyranosyl-(1→4)-[α-L-rhamnopyranosyl-(1→2)]-ß-D-galactopyranoside (4), together with eleven known compounds (5-15) were isolated from the branches and leaves of Tribulus terrestris. Their chemical structures were established through spectroscopic methods, including HR-ESI-MS, 1D-, and 2D-NMR spectra. Preliminary biological evaluation on NO production inhibitory activity in LPS activated RAW 264.7 cells showed that compounds 1-3, 5, and 6 had significant inhibitory effects with IC50 values ranging from 2.4 to 18.3 µM, compared to that of the positive control compound, dexamethazone (IC50 13.6 µM).

Polyhydroxylated Spirostanol Saponins from the Rhizomes of Paris dulongensis.

Three new polyhydroxylated spirostanol steroidal saponins, dulongenosides B-D (2-4), along with 14 known compounds, dulongenoside A (1), padelaoside B (5), parisyunnanoside G (6), polyphyllin D (7), ophiopogonin C' (8), formosanin C (9), dioscin (10), paris saponin VII (11), paris H (12), parisyunnanoside I (13), protodioscin (14), proprotogracillin (15), crustecdysone (16), and stigmasterol-3-O-ß-d-glucopyranoside (17), were isolated from the rhizomes of Paris dulongensis (Melanthiaceae). Their chemical structures were elucidated based on extensive analyses of NMR and MS data and acidic hydrolyses. The isolates were evaluated for their cytotoxicity to five human cancer cell lines (HL-60, SW480, MDA-MB-231, A549, and A549/Taxol) and the normal human bronchial epithelial cell line BEAS-2B by the MTS test. Compounds 7-12 and 14 showed cytotoxic activity, with IC50 values ranging from 0.20 to 4.35 µM. Proprotogracillin selectively inhibited A549 (IC50=0.58 µM) and A549/Taxol (IC50=0.74 µM) cells, with no significant cytotoxic activity against HL-60, SW480, MDA-MB-231, or BEAS-2B cells, with IC50 values greater than 40 µM.

Two New Isospirostanol-Type Saponins from the Bulbs of Lilium Brownii and Their Anti-Hepatocarcinogenic Activity.

Bulbs of Lilium brownii, commonly known as "Bai-he" in China, serve both edible and medicinal purposes in clinical practice. In this study, two new isospirostanol-type saponins were isolated from L. brownii, and their structures were identified by spectroscopic method, and absolute configurations were elucidated by comprehensive analysis of spectral data obtained from combined acid hydrolysis. Two compounds were finally identified as 3-O-[α-L-rhamnopyranosyl-(1→2)-ß-D-glucopyranoside]-(22R,25R)-5α-spirosolane-3ß-ol (1) and 3-O-{α-L-rhamnopyranosyl-(1→2)-[ß-D-glucopyranosyl-(1→4)]-ß-D-glucopyranoside}-(22R,25R)-5α-spirosolane-3ß-ol (2), respectively. Further, we found that compound 2 significantly suppressed the proliferation of SMMC-7721 and HepG2 cells with IC50 values of 26.3±1.08 µM and 30.9±1.59 µM, whereas compound 1 didn't inhibit both of the two hepatocellular carcinoma. Subsequently, compound 2 effectively decreased the levels of interleukin-1ß and tumor necrosis factor-α and the expression of Bcl-2, and increased the expression of Bax and Caspase-3 proteins. Which indicated that the anti-hepatocellular carcinoma effect of compound 2 involves reducing the level of inflammation and inducing apoptosis.

Investigation of Anti-Alzheimer's Mechanisms of Sarsasapogenin Derivatives by Network-Based Combining Structure-Based Methods.

Alzheimer's disease (AD), a neurodegenerative disease with no cure, affects millions of people worldwide and has become one of the biggest healthcare challenges. Some investigated compounds play anti-AD roles at the cellular or the animal level, but their molecular mechanisms remain unclear. In this study, we designed a strategy combining network-based and structure-based methods together to identify targets for anti-AD sarsasapogenin derivatives (AAs). First, we collected drug-target interactions (DTIs) data from public databases, constructed a global DTI network, and generated drug-substructure associations. After network construction, network-based models were built for DTI prediction. The best bSDTNBI-FCFP_4 model was further used to predict DTIs for AAs. Second, a structure-based molecular docking method was employed for rescreening the prediction results to obtain more credible target proteins. Finally, in vitro experiments were conducted for validation of the predicted targets, and Nrf2 showed significant evidence as the target of anti-AD compound AA13. Moreover, we analyzed the potential mechanisms of AA13 for the treatment of AD. Generally, our combined strategy could be applied to other novel drugs or compounds and become a useful tool in identification of new targets and elucidation of disease mechanisms. Our model was deployed on our NetInfer web server (http://lmmd.ecust.edu.cn/netinfer/).

Isolation and identification of hemostatic steroidal glycosides from Ypsilandra thibetica.

The phytoconstituents of the fraction with hemostatic activity of the 70% aqueous ethanol extract of Ypsilandra thibetica Franch. were investigated. As a result, fourteen previously unreported spirostanol saponins, ypsilandrosides Z1-Z14, and nine known analogues were isolated and characterized by MS, NMR, and chemical methods. Among them, ypsilandrosides Z1-Z4 (1-4) have a rare 12-O-ß-d-glucopyranosyl group, while ypsilandrosides Z5-Z8 (5-8) possess a rare double bond between C-4 and C-5, and a hydroxyl or carbonyl located at the C-6. All isolates were further tested for their hemostatic activity. The results suggested that five spirostanol tetraglycosides show favorable inducing platelet aggregation activities. Among them, ypsilandroside G (16) displayed significant inducing platelet aggregation activity with an EC50 value of 57.17 µM. Furthermore, the preliminary structure-activity relationship of these spirostanol glycosides' hemostatic activity was discussed.

Protective effects and mechanism of action of ruscogenin in a mouse model of ovalbumin-induced asthma.

OBJECTIVE: Ruscogenin is a natural product exhibiting anti-inflammatory, antioxidant, and anti-apoptotic effects; however, its effectiveness for asthma management has not yet been reported. The aim of this study was to explore the role of ruscogenin in airway inflammation and apoptosis in asthma. METHODS: In vivo, female 6- to 8-week-old CL57 mice were sensitized to ovalbumin and challenged intranasally for 7 days. One group was gavaged with ruscogenin before ovalbumin challenge. At the end of the challenge period, airway hyperresponsiveness and airway inflammation were evaluated. Enzyme-linked immunosorbent assay was used to estimate the oxidative stress levels. A terminal deoxynucleotidyl transferase dUDP nick-end labeling assay was used to determine the extent of apoptosis. Immunohistochemistry and western blotting were performed to examine VDAC1 expression. In vitro, human bronchial epithelial (HBE) cells were treated with H2O2, ruscogenin, or disulfonate salt, and flow cytometry was used to calculate the apoptosis ratio and detect mitochondrial calcium levels. RESULTS: In vivo, ruscogenin improved airway hyperresponsiveness and airway inflammation, while reducing oxidative stress, the apoptosis ratio and VDAC1 expression in asthmatic lungs. In vitro, ruscogenin attenuated apoptosis in HBE cells by decreasing the levels of VDAC1 expression and mitochondrial calcium. CONCLUSION: Ruscogenin reduced oxidative stress and apoptosis in the airway epithelium by inhibiting VDAC1 expression and mitochondrial handling of calcium.

Androgenic properties of the dietary supplement 5α-hydroxy-laxogenin.

Dietary supplements sold for anabolic benefits or performance enhancement often contain substances, which are non-approved and might lack quality controls. With regard to athletes, the inclusion of substances or methods in the prohibited list of the World Anti-Doping Agency is based on medical or scientific evidence. 5α-hydroxy-laxogenin is a synthetic spirostane-type steroid, which is contained in dietary supplements and advertised as anabolic agent. To date, evidence is missing on anabolic or androgenic activity of 5α-hydroxy-laxogenin. We investigated its androgenic potential in two in vitro bioassays. While no activity was observed in the yeast androgen screen, 5α-hydroxy-laxogenin was able to trans-activate the androgen receptor in human prostate cells in a dose-dependent manner. Interestingly, a biphasic response was observed with antagonistic properties at lower concentrations and agonistic effects at higher concentrations tested. The demonstrated androgenic properties of the higher concentrations demonstrate that further investigations should focus on the safety as well as on potential anabolic effects of 5α-hydroxy-laxogenin. This is of interest with regard to abuse for doping purposes.

Metabolomics Mechanism and Lignin Response to Laxogenin C, a Natural Regulator of Plants Growth.

Laxogenin C (LGC) is a natural spirostanol deriving from plant hormone which has shown growing regulation similar to those of brassinosteroids. In the present study, LGC showed a promoting effect on tomato seed germination and seedling growth in a dose-dependent manner. We applied LC-MS/MS to investigate metabolome variations in the tomato treated with LGC, which revealed 10 differential metabolites (DMs) related to KEGG metabolites, associated with low and high doses of LGC. Enrichment and pathway mapping based on the KEGG database indicated that LGC regulated expressions of 2-hydroxycinnamic acid and l-phenylalanine to interfere with phenylalanine metabolism and phenylpropanoids biosynthesis. The two pathways are closely related to plant growth and lignin formation. In our further phenotypic verification, LGC was confirmed to affect seedling lignification and related phenylpropanoids, trans-ferulic acid and l-phenylalanine levels. These findings provided a metabolomic aspect on the plant hormone derivates and revealed the affected metabolites. Elucidating their regulation mechanisms can contribute to the development of sustainable agriculture. Further studies on agrichemical development would provide eco-friendly and efficient regulators for plant growth control and quality improvement.

Chemistry, Biosynthesis and Pharmacology of Sarsasapogenin: A Potential Natural Steroid Molecule for New Drug Design, Development and Therapy.

Sarsasapogenin is a natural steroidal sapogenin molecule obtained mainly from Anemarrhena asphodeloides Bunge. Among the various phytosteroids present, sarsasapogenin has emerged as a promising molecule due to the fact of its diverse pharmacological activities. In this review, the chemistry, biosynthesis and pharmacological potentials of sarsasapogenin are summarised. Between 1996 and the present, the relevant literature regarding sarsasapogenin was obtained from scientific databases including PubMed, ScienceDirect, Scopus, and Google Scholar. Overall, sarsasapogenin is a potent molecule with anti-inflammatory, anticancer, antidiabetic, anti-osteoclastogenic and neuroprotective activities. It is also a potential molecule in the treatment for precocious puberty. This review also discusses the metabolism, pharmacokinetics and possible structural modifications as well as obstacles and opportunities for sarsasapogenin to become a drug molecule in the near future. More comprehensive preclinical studies, clinical trials, drug delivery, formulations of effective doses in pharmacokinetics studies, evaluation of adverse effects and potential synergistic effects with other drugs need to be thoroughly investigated to make sarsasapogenin a potential molecule for future drug development.

Isolation and Identification of Pennogenin Tetraglycoside from Cestrum nocturnum (Solanaceae) and Its Antifungal Activity against Fusarium kuroshium, Causal Agent of Fusarium Dieback.

Antifungal assay-guided fractionation of the methanolic crude extract of Cestrum nocturnum (Solanaceae), popular known as 'lady of the night', led the isolation and identification of the steroidal saponin named pennogenin tetraglycoside, which was identified for the first time in this plant species by spectroscopic means. The crude extract, fractions and pennogenin tetraglycoside exhibited mycelial growth inhibition of Fusarium solani and F. kuroshium. F. solani is a cosmopolitan fungal phytopathogen that affects several economically important crops. However, we highlight the antifungal activity displayed by pennogenin tetraglycoside against F. kuroshium, since it is the first plant natural product identified as active for this phytopathogen. This fungus along with its insect symbiont known as Kuroshio shot hole borer (Euwallacea kuroshio) are the causal agents of the plant disease Fusarium dieback that affects more than 300 plant species including avocado (Persea americana) among others of ecological relevance. Scanning electron microscopy showed morphological alterations of the fungal hyphae after exposure with the active fractions and 12 phenolic compounds were also identified by mass spectrometry dereplication as part of potential active molecules present in C. nocturnum leaves.

[Protective effect of ophiopogonin D against isoproterenol-induced cardiomyocyte injury and targets].

This study aims to unveil the effect of ophiopogonin D(OPD) on isoproterenol(ISO)-induced apoptosis of rat cardiomyocytes and the possible targets, which is expected to provide clues for further research on the myocardial protection of ophiopogonins. Cell count kit-8(CCK-8) assay was used to detect viability of cells treated with OPD and ISO, Western blot to examine the effect of OPD and ISO on the expression of endoplasmic reticulum stress-related Bip, Bax, Perk, ATF4, caspase-12, and CHOP, flow cytometry to determine cell apoptosis rate, and Hoechst 33258 and Tunel staining to observe cell apoptosis and morphological changes. In addition, the probe for calcium ion-specific detection was employed to investigate calcium ion release from the endoplasmic reticulum, and OPD-bond epoxy-activated agarose solid-phase microspheres were prepared and used as affinity matrix to capture OPD-binding target proteins in H9 c2 cell lysate. For the target proteins of OPD identified by high-resolution mass spectrometry, the related signal pathways were enriched and the potential targets of OPD against cardiomyocyte injury were discussed. The experimental result showed that 10 µmol·L~(-1) ISO can significantly induce the expression of endoplasmic reticulum stress-related proteins and promote cell apoptosis. Different concentration of OPD can prevent the damage of myocardial cells caused by ISO. According to mass spectrometry results, 19 proteins, including Fam129 a and Pdia6, were involved in multiple signaling pathways such as the unfolded protein reaction bound by the ERN1 sensor, tricarboxylic acid cycle, and Nrf2 signal transduction pathway. The above results indicate that OPD protects cardiomyocytes by regulating multiple signaling pathways of target proteins and affecting cell cycle progression.

Gracillin shows potent efficacy against colorectal cancer through inhibiting the STAT3 pathway.

Colorectal cancer (CRC) accounts for about 10% of all annually diagnosed cancers and cancer-related deaths worldwide. STAT3 plays a vital role in the occurrence and development of tumours. Gracillin has shown a significant antitumour activity in tumours, but its mechanism remains unknown. The human CRC cell lines HCT116, RKO, and SW480 and immunodeficient mice were used as models to study the effects of gracillin on cell proliferation, migration and apoptosis. These were evaluated by cell viability, colony formation, wound-healing migration and cell apoptosis assays. Luciferase reporter assay, and immunostaining and western blot analyses were used to explore the specific mechanism through which gracillin exerts its effects. Gracillin significantly reduces viability and migration and stimulates apoptosis in human CRC cells. It also significantly inhibits tumour growth with no apparent physiological toxicity in animal model experiments. Moreover, gracillin is found to inhibit STAT3 phosphorylation and STAT3 target gene products. In addition, gracillin inhibits IL6-induced nuclear translocation of P-STAT3. Gracillin shows potent efficacy against CRC by inhibiting the STAT3 pathway. It should be further explored as a unique STAT3 inhibitor for the treatment of CRC.

Apigenin restores endothelial function by ameliorating oxidative stress, reverses aortic stiffening, and mitigates vascular inflammation with aging.

We assessed the efficacy of oral supplementation with the flavanoid apigenin on arterial function during aging and identified critical mechanisms of action. Young (6 mo) and old (27 mo) C57BL/6N mice (model of arterial aging) consumed drinking water containing vehicle (0.2% carboxymethylcellulose; 10 young and 7 old) or apigenin (0.5 mg/mL in vehicle; 10 young and 9 old) for 6 wk. In vehicle-treated animals, isolated carotid artery endothelium-dependent dilation (EDD), bioassay of endothelial function, was impaired in old versus young (70% ± 9% vs. 92% ± 1%, P < 0.0001) due to reduced nitric oxide (NO) bioavailability. Old mice had greater arterial reactive oxygen species (ROS) production and oxidative stress (higher nitrotyrosine) associated with greater nicotinamide adenine dinucleotide phosphate oxidase (oxidant enzyme) and lower superoxide dismutase 1 and 2 (antioxidant enzymes); ex vivo administration of Tempol (antioxidant) restored EDD to young levels, indicating ROS-mediated suppression of EDD. Old animals also had greater aortic stiffness as indicated by higher aortic pulse wave velocity (PWV, 434 ± 9 vs. 346 ± 5 cm/s, P < 0.0001) due to greater intrinsic aortic wall stiffness associated with lower elastin levels and higher collagen, advanced glycation end products (AGEs), and proinflammatory cytokine abundance. In old mice, apigenin restored EDD (96% ± 2%) by increasing NO bioavailability, normalized arterial ROS, oxidative stress, and antioxidant expression, and abolished ROS inhibition of EDD. Moreover, apigenin prevented foam cell formation in vitro (initiating step in atherosclerosis) and mitigated age-associated aortic stiffening (PWV 373 ± 5 cm/s) by normalizing aortic intrinsic wall stiffness, collagen, elastin, AGEs, and inflammation. Thus, apigenin is a promising therapeutic for arterial aging.NEW & NOTEWORTHY Our study provides novel evidence that oral apigenin supplementation can reverse two clinically important indicators of arterial dysfunction with age, namely, vascular endothelial dysfunction and large elastic artery stiffening, and prevents foam cell formation in an established cell culture model of early atherosclerosis. Importantly, our results provide extensive insight into the biological mechanisms of apigenin action, including increased nitric oxide bioavailability, normalization of age-related increases in arterial ROS production and oxidative stress, reversal of age-associated aortic intrinsic mechanical wall stiffening and adverse remodeling of the extracellular matrix, and suppression of vascular inflammation. Given that apigenin is commercially available as a dietary supplement in humans, these preclinical findings provide the experimental basis for future translational studies assessing the potential of apigenin to treat arterial dysfunction and reduce cardiovascular disease risk with aging.

Sarsasapogenin improves adipose tissue inflammation and ameliorates insulin resistance in high-fat diet-fed C57BL/6J mice.

Insulin resistance is a major cause of type 2 diabetes and metabolic syndrome. Macrophage infiltration into obese adipose tissue promotes inflammatory responses that contribute to the pathogenesis of insulin resistance. Suppression of adipose tissue inflammatory responses is postulated to increase insulin sensitivity in obese patients and animals. Sarsasapogenin (ZGY) is one of the metabolites of timosaponin AIII in the gut, which has been shown to exert anti-inflammatory action. In this study, we investigated the effects of ZGY treatment on obesity-induced insulin resistance in mice. We showed that pretreatment with ZGY (80 mg·kg-1·d-1, ig, for 18 days) significantly inhibited acute adipose tissue inflammatory responses in LPS-treated mice. In high-fat diet (HFD)-fed obese mice, oral administration of ZGY (80 mg·kg-1·d-1, for 6 weeks) ameliorated insulin resistance and alleviated inflammation in adipose tissues by reducing the infiltration of macrophages. Furthermore, we demonstrated that ZGY not only directly inhibited inflammatory responses in macrophages and adipocytes, but also interrupts the crosstalk between macrophages and adipocytes in vitro, improving adipocyte insulin resistance. The insulin-sensitizing and anti-inflammatory effects of ZGY may result from inactivation of the IKK /NF-κB and JNK inflammatory signaling pathways in adipocytes. Collectively, our findings suggest that ZGY ameliorates insulin resistance and alleviates the adipose inflammatory state in HFD mice, suggesting that ZGY may be a potential agent for the treatment of insulin resistance and obesity-related metabolic diseases.

Ruscogenin attenuates particulate matter-induced acute lung injury in mice via protecting pulmonary endothelial barrier and inhibiting TLR4 signaling pathway.

The inhalation of particulate matter (PM) is closely related to respiratory damage, including acute lung injury (ALI), characterized by inflammatory fluid edema and disturbed alveolar-capillary permeability. Ruscogenin (RUS), the main active ingredient in the traditional Chinese medicine Ophiopogonis japonicus, has been found to exhibit anti-inflammatory activity and rescue LPS-induced ALI. In this study, we investigated whether and how RUS exerted therapeutic effects on PM-induced ALI. RUS (0.1, 0.3, 1 mg·kg-1·d-1) was orally administered to mice prior to or after intratracheal instillation of PM suspension (50 mg/kg). We showed that RUS administration either prior to or after PM challenge significantly attenuated PM-induced pathological injury, lung edema, vascular leakage and VE-cadherin expression in lung tissue. RUS administration significantly decreased the levels of cytokines IL-6 and IL-1ß, as well as the levels of NO and MPO in both bronchoalveolar lavage fluid (BALF) and serum. RUS administration dose-dependently suppressed the phosphorylation of NF-κB p65 and the expression of TLR4 and MyD88 in lung tissue. Furthermore, TLR4 knockout partly diminished PM-induced lung injury, and abolished the protective effects of RUS in PM-instilled mice. In conclusion, RUS effectively alleviates PM-induced ALI probably by inhibition of vascular leakage and TLR4/MyD88 signaling. TLR4 might be crucial for PM to initiate pulmonary lesion and for RUS to exert efficacy against PM-induced lung injury.

In vitro study on the effect of ophiopogonin D on the activity of cytochrome P450 enzymes.

Ophiopogonin D is a commonly used herb in cardiology and pediatrics for its variuos pharmacological effects. It is necessary to investigate the effect of ophiopogonin D on the activity of cytochrome P450 enzymes (CYP450s) to provide more guidance for the clinical application of ophiopogonin D. Eight isoforms of CYP450s, including CYP1A2, 2A6, 2C8, 2C9, 2C19, 2D6, 2E1, and 3A4 were incubated with 100 µM ophiopogonin D in pooled human liver microsomes. The inhibition model and corresponding parameters were also investigated. Ophiopogonin D exerted a significant inhibitory effect on the activity of CYP3A4, 2C9, and 2E1 in a dose-dependent manner with the IC50 values of 8.08, 12.92, and 22.72 µM, respectively (p < 0.05). The inhibition of CYP3A4 by ophiopogonin D was performed non-competitively and time-dependently with the Ki value of 4.08 µM and the KI/Kinact value of 5.02/0.050 min-1·µM-1. Whereas, ophiopogonin D acts as a competitive inhibitor of CYP2E1 and 2C9 with the Ki value of 6.69 and 11.07 µM, respectively. The inhibitory effect of ophiopogonin D on the activity of CYP3A4, 2C9, and 2E1 indicated the potential drug-drug interaction between ophiopogonin D and drugs metabolized by these CYP450s, which needs further in vivo investigation and validation.

Insight into the cardiovascular mechanisms of blood pressure lowering effect of gitogenin: a steroidal saponin.

Background/objectives: Steroidal saponins are widely distributed in medicinal plants with potential applications in cardiovascular disorders. Gitogenin, a saponin, has not been explored as antihypertensive; this investigation was aimed to explore its blood pressure lowering potential and underlying mechanisms.Methodology: The effect of gitogenin was evaluated on blood pressure in vivo, using normotensive rat model and the underlying cardiovascular mechanism(s) in vitro, in isolated rat aorta and in atria preparations using PowerLab data acquisition system (ADInstrument, Australia).Results: Intravenous injection of gitogenin decreased mean arterial pressure (MAP) in anesthetized rats. Atropine (1 mg/kg) and L-NAME (100 mg/kg) pretreatment significantly (*p < .05) attenuated effect on MAP to gitogenin. In isolated intact aortic rings, gitogenin induced endothelium-dependent vasodilatation (maximum 65%), which was ablated (maximum 22%) with L-NAME (100 mg/kg) and atropine (1 µM) pretreatment or endothelium removal. Gitogenin was found more potent against angiotensin II precontractions without effect on high K+ and low K+ precontractions. In isolated rat right atria, gitogenin suppressed rate and force of contractions. Atropine (1 µM) pretreatment partially inhibited effect of gitogenin on force and eliminated its effect on rate. Combined atropine (10 µM) and atenolol (0.5 µM) pretreatment was without effect on force of contractions but eliminated effect of gitogenin on rate with 25% increase.Conclusion: These findings indicate that antihypertensive effect of gitogenin is the outcome of vascular and cardiac effects; agonistic effect on vascular M3 and cardiac M2 receptors; and being more selective for M2. Increase in the rate of atrial contraction might be of clinical importance.