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Lung cancer, one of the most often diagnosed malignancies, is the top cause of death in both men and women globally. In both developed and emerging countries, high incidences of cancer are becoming a huge health burden. Natural resources, including plants, have always been a possible source of lead compounds in the identification of optimal medications for cancer treatment, with natural resources accounting for around half of all anticancer drugs. Ruscogenin, a natural saponin, is a major component of Radix Ophiopogon japonicus with a well-established anticancer activity. In this study, the anticancer potential of ruscogenin against a B(a)P-challenged lung cancer model in mice was assessed. The mice were categorized into four groups: group I was as the control group, group II mice were challenged with B(a)P, group III rodents were treated with ruscogenin prior to challenge with B(a)P, and group IV rodents were treated with ruscogenin after B(a)P administration. Tumor incidence was calculated, and the following parameters were analyzed: body weight, lung weight, immunoglobulin (Ig) levels (IgG, IgA, and IgM), key marker enzymes, and proinflammatory cytokines in both treated and control mice. Lung tissues were analyzed via histopathological analysis. According to our results, all the markers that favor the growth of cancer were increased in the lung cancer group. After administration of ruscogenin, all the markers returned to their original levels, revealing the anticancer potential of ruscogenin.

Atopic dermatitis (AD) is a pruritic, chronic, relapsing inflammatory skin disease. The research aims to study the effects of Sarsasapogenin and its combination with Fluticasone in 2, 4-Dinitrofluorobenzene (DNFB) induced atopic dermatitis in BALB/c mice.Thirty male Balb/c mice were divided into 5 groups: (i) Normal control (NC), (ii) Disease control (DNFB), (iii) Sarsasapogenin (SG) (50 µg/mice), (iv) Fluticasone (FC) (50 µg/mice), (v) Sarsasapogenin + Fluticasone (SG + FC) combination (25 µg/mice). Dermatitis was induced by repeated application of DNFB in Balb/c mice. On topical application of SG, FC, and SG + FC combination on the ear and skin lesions, body weight, ear weight, ear thickness, erythema score, spleen weight, cytokines, immunoglobulin E (IgE) levels, nitric oxide (NO) level, hematological parameters, and oxidative stress markers were evaluated. Histological analysis of the ear tissue was also done.The results stated that SG and SG + FC treatment to mice considerably decrease the ear weight, ear thickness, spleen weight, serum IgE, cytokines, NO levels, and restoration of antioxidant stress markers with elevation in the hematological parameters. The observations were further confirmed by histopathological analysis of ear tissue.These data specify that SG has been demonstrated as a probable therapy for the treatment of allergic skin diseases in combination with FC by decreasing its dose from 50 to 25 µg/mice to avoid the chronic side effects of FC. Hence, it can be concluded that SG and SG + FC combination significantly improved the AD-like symptoms in the DNFB sensitized mice through mitigating the production of proinflammatory mediators and restoration of oxidative stress markers.

Ulcerative colitis (UC) is a chronic inflammatory bowel condition considered by oxido-nitrosative stress and the release of pro-inflammatory cytokines that affects the mucosal lining of the colon. Sarsasapogenin (SG), as an active component, has been found in many plants, and it exhibits potential protective effects, such as anti-inflammatory, antioxidant, anti-psoriasis, anti-arthritis, anti-asthma, anti-depressant and anti-cancer. However, the effects of SG on UC remain unknown.The purpose of this study was to investigate the effects of SG on 2, 4, 6-trinitrobenzene sulfonic acid (TNBS)-induced UC in rats.Thirty Wistar rats were randomized into five groups: (i) Normal control, (ii) Disease control (TNBS), (iii) Sarsasapogenin (SG) (50 µg/rat), (iv) Fluticasone (FC) (50 µg/rat), (v) Sarsasapogenin + Fluticasone (SG + FC) (25 µg/rat). UC was induced in rats by trans-rectal instillation of TNBS (10 mg/kg). SG, FC and SG + FC were administered for 11 days and on the 8th day colitis was induced. Several molecular, biochemical and histological alterations were evaluated in the colon tissue. All treatment group results were compared to the TNBS group results.The study results revealed that treatment of rats with SG and SG + FC combination significantly decreased the colon weight/length ratio, macroscopic inflammation score, lesions score, diarrhea score and adhesion score. Combination treatment in rats significantly reduced the production of biochemical parameters, proinflammatory cytokines, haematological parameters, serum IgE levels and restored the oxidative stress markers. SG and SG + FC treatment also considerably restored the histopathological changes induced by TNBS.Thus, SG and SG + FC combination could alter the disease progression and could be a hopeful therapeutic target for the management of UC by reducing its dose in combination with FC to elude the long term adverse effects of FC.

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-β-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 EC

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.Ruscogenin reduced oxidative stress and apoptosis in the airway epithelium by inhibiting VDAC1 expression and mitochondrial handling of calcium.

Sarsasapogenin (Sar), a natural steroidal compound, shows neuroprotection, cognition-enhancement, antiinflammation, antithrombosis effects, and so on. However, whether Sar has ameliorative effects on diabetes-associated cognitive impairment remains unknown. In this study, we found that Sar ameliorated diabetes-associated memory impairment in streptozotocin-induced diabetic rats, evidenced by increased numbers of crossing platform and percentage of time spent in the target quadrant in Morris water maze tests, and suppressed the nucleotide-binding domain and leucine-rich repeat containing protein 1 (NLRP1) inflammasome in hippocampus and cerebral cortex. Furthermore, Sar inhibited advanced glycation end-products and its receptor (AGEs/RAGE) axis and suppressed up-regulation of thrombin receptor protease-activated receptor 1 (PAR-1) in cerebral cortex. On the other hand, Sar mitigated high glucose-induced neuronal damages, NLRP1 inflammasome activation, and PAR-1 up-regulation in high glucose-cultured SH-SY5Y cells, but did not affect thrombin activity. Moreover, the effects of Sar were similar to those of a selective PAR-1 antagonist vorapaxar. Further studies indicated that activation of the NLRP1 inflammasome and NF-κB mediated the effect of PAR-1 up-regulation in high glucose condition by using PAR-1 knockdown assay. In summary, this study demonstrated that Sar prevented memory impairment caused by diabetes, which was achieved through suppressing neuroinflammation from activated NLRP1 inflammasome and NF-κB regulated by cerebral PAR-1. HIGHLIGHTS: Sarsasapogenin ameliorated memory impairment caused by diabetes in rats. Sarsasapogenin mitigated neuronal damages and neuroinflammation by down-regulating cerebral PAR-1. The NLRP1 inflammasome and NF-κB signaling mediated the pro-inflammatory effects of PAR-1. Sarsasapogenin was a pleiotropic neuroprotective agent and memory enhancer in diabetic rodents.

ObjectivesSepsis-associated acute lung injury (ALI) occurs with the highest morbidity and carries the highest mortality rates among the pathogenies of ALI. Ruscogenin (RUS) has been found to exhibit anti-inflammation property and rescue lipopolysaccharide-induced ALI, but little is known about its role in sepsis-triggered ALI. The aim of this study was to investigate the potential role of RUS in sepsis-induced ALI and the probable mechanism.MethodsMice model of cecal ligation and puncture (CLP) was replicated, and three doses of RUS (0.01, 0.03 and 0.1 mg/kg) were administrated 1 h before CLP surgeries.Key findingsRUS significantly extended the survival time and attenuated the lung pathological injury, oedema and vascular leakage in sepsis-induced ALI mice. RUS efficiently decreased the level of MPO in lung tissue and the WBC, NEU counts in BALF. In addition, RUS rescued the expression of VE-cadherin and p120-catenin and suppressed the TLR4/Src signalling in lung tissue.ConclusionsRUS attenuated sepsis-induced ALI via protecting pulmonary endothelial barrier and regulating TLR4/Src/p120-catenin/VE-cadherin signalling pathway.

Jianqiong Yang,1,* Ling Cao,2,* Yamei Li,2,* Hai Liu,2,3,* Minhong Zhang,1 Huamou Ma,1 Biao Wang,1 Xiaoliang Yuan,4 Qian Liu5,6 1Department of Clinical Medicine Research Center, The First Affiliated Hospital of Gannan Medical University, Ganzhou, Jiangxi, People’s Republic of China; 2College of Pharmacy, Gannan Medical University, Ganzhou, Jiangxi, People’s Republic of China; 3National Engineering Research Center for Modernization of Traditional Chinese Medicine - Hakka Medical Resources Branch, Gannan Medical University, Ganzhou, Jiangxi, People’s Republic of China; 4Department of Respiratory Medicine, The First Affiliated Hospital of Gannan Medical University, Ganzhou, Jiangxi, People’s Republic of China; 5Key Laboratory of Prevention and Treatment of Cardiovascular and Cerebrovascular Diseases of Ministry of Education, Gannan Medical University, Ganzhou, Jiangxi, People’s Republic of China; 6Department of Pediatric Surgery, The First Affiliated Hospital of Gannan Medical University, Ganzhou, Jiangxi, People’s Republic of China*These authors contributed equally to this workCorrespondence: Qian LiuGannan Medical University, #1 Yixueyuan Road, Ganzhou, Jiangxi, People’s Republic of ChinaTel +86 797 8169666Fax +86 797 8169600Email liuqiangmu2017@126.comBackground: Reineckia carnea is commonly used to treat cough, pneumonia and other diseases in China. In our previous study, it was found that the ethanol extracts of Reineckia carnea have a strong inhibitory effect on the proliferation of human lung cancer A549 cells. Here, we isolated gracillin from ethanol extracts for the first time.Purpose: Clarify the antiproliferation effect of gracillin on A549 cells and further explore its mechanisms via the mitochondrial pathway.Methods: Gracillin was isolated and purified by silica gel, D-101 macroporous resin and preparative RP-HPLC, then identified by NMR and HR-MS. The inhibitory effects of gracillin on the proliferation of A549 cells were detected by the MTS method. Its mechanisms were further explored by flow cytometry and Western blot.Results: A steroid saponin, gracillin, was isolated and identified from Reineckia carnea for the first time. In a concentration-dependent and time-dependent manner, gracillin significantly inhibited the proliferation of A549 cells with an IC50 value at 2.54 μmol/L and induced morphological changes. The results of flow cytometry analysis showed that the apoptosis rate of A549 cells was significantly increased (p < 0.05), and the cells proportion was obviously arrested in S phase. The concentration of intracellular calcium was raised (p < 0.01), and the mitochondrial membrane potential was greatly decreased (p < 0.01). In addition, the expression levels of Bax, caspase-3, cleaved caspase-3, and cytochrome C were dramatically up-regulated while Bcl-2 was down-regulated (p < 0.05) in A549 cells.Conclusion: This study confirmed that gracillin has a significant antiproliferative effect on A549 cells. Gracillin could induce the apoptosis of A549 cells through the mitochondrial pathway, which might be associated with regulation of the concentration of intracellular calcium, the mitochondrial membrane potential and the expression levels of Bax, Bcl-2, caspase-3, cleaved caspase-3, and cytochrome C.Keywords: Reineckia carnea, gracillin, antiproliferative activity, A549 cells, apoptosis

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.

To investigate the pharmacodynamic effects of ruscogenin on acute kidney injury and the Rev-erb

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.

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-1β, 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.

Jiaxuan Peng,1,* Kangxian Zhao,2,* Jiling Zhu,3,* Yanben Wang,4,5 Peng Sun,2 Qichang Yang,2 Tan Zhang,4 Weiqi Han,4 Wenjun Hu,4 Wanlei Yang,4 Jianwei Ruan,6 Yu Qian1,2,4 1Guangxi Key Laboratory of Regenerative Medicine, Guangxi Medical University, Guangxi 530021, People’s Republic of China; 2The Second Affiliated Hospital and Yuying Children’s Hospital of Wenzhou Medical University, Wenzhou, Zhejiang 325000, People’s Republic of China; 3Department of Clinical Medicine, Medical College of Shaoxing University, Shaoxing, Zhejiang 312000, People’s Republic of China; 4Department of Orthopaedics, Shaoxing People’s Hospital, Zhejiang University School of Medicine, Shaoxing 312000, Zhejiang, People’s Republic of China; 5Department of Orthopaedics, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, People’s Republic of China; 6Department of Orthopaedics, Taizhou Municipal Hospital, Taizhou 318000, Zhejiang, Republic of China*These authors contributed equally to this workCorrespondence: Yu QianDepartment of Orthopaedics, Shaoxing People’s Hospital, Zhejiang University School of Medicine, Shaoxing 312000, Zhejiang, People’s Republic of ChinaEmail doctor120@hotmail.comIntroduction: Osteoclasts are giant polynuclear cells; their main function is bone resorption. An increased number of osteoclasts and enhanced bone resorption exert significant effects on osteoclast-related bone-lytic diseases, including osteoporosis. Given the limitations of current therapies for osteolytic diseases, it is urgently required to develop safer and more effective alternatives. Sarsasapogenin, a major sapogenin from Anemarrhena asphodeloides Bunge, possesses potent antitumor effects and inhibits NF-κB and MAPK signaling. However, the manner in which it affects osteoclasts is unclear.Methods: We investigated the effects of anti-osteoclastogenic and anti-resorptive of sarsasapogenin on bone marrow-derived osteoclasts.Results: Sarsasapogenin inhibited multiple RANKL-induced signaling cascades, thereby inhibiting the induction of key osteoclast transcription factor NFATc1. The in vivo and in vitro results were consistent: sarsasapogenin treatment protected against bone loss in a mouse osteolysis model induced by lipopolysaccharide.Conclusion: Our research confirms that sarsasapogenin can be used as a new treatment for osteoclast-related osteolytic diseases.Keywords: sarsasapogenin, osteoclast, osteoclastogenesis, NF-κB, MAPK, NFATc1, therapeutics

The dried root of Asparagus cochinchinensis (RAC) has been used as an important traditional Chinese medicine for a long time in China. Steroidal saponins (SSs) are considered to be the main active ingredients of this herb. However, the isolation and structural determination of SSs from RAC are time-consuming and laborious. For this reason, the development of new methods for the separation and characterization of SSs is highly desirable. In this study, a new high-performance liquid chromatography coupled to electrospray ionization and quadrupole time-of-flight mass spectrometry (HPLC-ESI-QTOF-MS/MS) method with precursor ions and the corresponding fragment ions was developed for the identification of SSs in RAC. Finally, 30 SSs have been detected and identified, including 17 potential new compounds. This is the first systematic study of SSs in RAC by HPLC-ESI-QTOF-MS/MS method.

Sepsis induces critical myocardial dysfunction, resulting in an increased mortality. Gracillin (GRA) is a natural steroidal saponin, showing strong capacities of anti-inflammation, but its pharmacological effects on lipopolysaccharide (LPS)-induced acute cardiac injury still remain unclear. In this study, we attempted to explore if GRA was effective to attenuate cardiac injury in LPS-challenged mice and the underlying mechanisms. First, we found that GRA treatments markedly up-regulated the expression of miR-29a in cardiomyocytes. LPS-induced cytotoxicity in cardiomyocytes was significantly alleviated by GRA treatment, as evidenced by the improved cell viability and reduced lactate dehydrogenase (LDH) release. In addition, LPS-triggered apoptotic cell death was clearly ameliorated in cardiomyocytes co-treated with GRA. Notably, LPS-exposed cells showed significantly reduced expression of miR-29a, while being rescued by GRA treatment. In vivo, LPS apparently impaired cardiac function in mice, which was, however, alleviated by GRA administration. In addition, GRA markedly attenuated apoptosis in hearts of LPS-challenged mice by decreasing the expression of cleaved Caspase-3. LPS-triggered inflammatory response in cardiac tissues was also suppressed by GRA through blocking nuclear factor κB (NF-κB) signaling pathway. We also found that miR-29a expression was highly reduced in hearts of LPS-treated mice but was rescued by GRA pretreatment. Besides, miR-29a mimic alleviated LPS-induced apoptosis and inflammation in cardiomyocytes; however, LPS-caused effects were further accelerated by miR-29a. Of note, the protective effects of GRA on LPS-injured cardiac tissues were significantly abrogated by miR-29a suppression. In conclusion, our findings demonstrated that GRA exerted an effective role against LPS-induced acute cardiac injury through impeding apoptosis and inflammation regulated by miR-29a.

Objective: Asthma is a very common airway inflammatory disease for which the existing drug therapy options are insufficient. In this study, we explored the mechanisms underlying the anti-in...

karagulle, onur olgac/0000-0002-1575-5751 WOS:000509485100002 PubMed ID: 31345657 Background/Objective: The present study investigated the potent therapeutic effects of Ruscogenin, main steroid sapogenin of traditional Chinese plant called 'Ophiopogon japonicas', on chronic ulcer model established with acetic acid in rats. Methods: 24 rats were attenuated to the sham (2 ml/kg/day isotonic solution), control (untreated ulcer) and treatment (3 ml/kg/day ruscogenin) groups. After treatment for 2 weeks, gastric tissues were collected and prepared for light microscopic (H&E), immunohistochemical (Collagen I, III and IV) and biochemical analysis [Epidermal growth factor (EGF), Prostaglandin E2 (PGE2), Tumor Necrosis Factor alpha (TNF-alpha), Interleukin 6 and 8 (IL-6 and IL-8), Lipid Peroxidase (LPO), Myeloperoxidase (MPO), Glutathione (GSH) and Glutathione Peroxidase (GSH-Px)] and transmission electron microscopy (TEM). Results: Macroscopic scoring showed that the ulceration area of ruscogenin-treated group decreased compared with control group. Immunohistochemical analysis revealed ruscogenin ameliorated and restored the levels of Collagen I and IV to the levels of sham group. Tissue levels of EGF and PGE2 enhanced significantly in untreated ulcer group while were higher in treated ulcer group than the control group. TNF-alpha, IL-6, IL-8, LPO, MPO levels increased significantly in control group whereas decreased in treated rats after ruscogenin treatment. However, levels of GSH and GSH-Px increased significantly in treatment group. TEM showed chief cells and parietal cells of ulcer group having degenerated organelles while ruscogenin group had normal ultrastructure of cells. Conclusion: There are potent anti-inflammatory and anti-oxidant effects of ruscogenin on gastric ulcer and may be successfully used as a safe and therapeutic agent in treatment of peptic ulcer. (C) 2019 Asian Surgical Association and Taiwan Robotic Surgery Association. Publishing services by Elsevier B.V. Research and Development Expenses Account under Circulating Capital Budget of University of Health Science Bagcilar Training and Research Hospital, Istanbul, Turkey [2016/12] The present project was supported by Research and Development Expenses Account under Circulating Capital Budget of University of Health Science Bagcilar Training and Research Hospital, Istanbul, Turkey (Project No: 2016/12).

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.

Ophiopogonin‑B (OP‑B) is a bioactive component from the root of

Antifungal assay-guided fractionation of the methanolic crude extract of

Danhong Pan,1 Jinqiang Zhu,2 Liexiang Cao,2 Beilei Zhu,3 Lili Lin1 1Emergency Care Unit, The First People’s Hospital of Wenling, Wenling, Zhejiang, 317500, People’s Republic of China; 2Emergency Department, The First People’s Hospital of Wenling, Wenling, Zhejiang, 317500, People’s Republic of China; 3Department of Neurology, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, 325000, People’s Republic of ChinaCorrespondence: Lili Lin, Emergency Care Unit, The First People’s Hospital of Wenling, No. 333, Chuan’an South Road, Chengxi Street, Wenling, Zhejiang, 317500, People’s Republic of China, Tel +86– 13616760809, Email lllin_24126_zh@hotmail.comIntroduction: Endothelial dysfunction (ED) is associated with the progression of sepsis. Ruscogenin (RUS) has shown considerable efficacy in treating ED and sepsis. In the current study, the effects of RUS on sepsis-induced ED were assessed, and the mechanism was explored by focusing on the interactions of RUS with miRs.Methods: Sepsis was induced in mice and in human umbilical vein endothelial cells (HUVECs) using LPS method. Expression profile of miRs responding to sepsis was determined. Symptoms associated with sepsis and ED were examined after treatment with RUS. Changes in mouse survival, arterial structure, systemic inflammation, cell viability, apoptosis, and the miR-146a-5p/NRP2/SSH1 axis were analyzed.Results: Based on the microarray results, miR-146a-5p was selected as the therapeutic target. RUS improved survival rates and arterial structure, suppressed proinflammatory cytokines, down-regulated miR-146a-5p, and up-regulated NPR2 and SSH1 in septic mice. In HUVECs, RUS increased cell viability, suppressed apoptosis, inhibited inflammation, downregulated miR-146a-5p, and increased NRP2 and SSH1 levels. The re-induction of miR-146a-5p-5p impaired the protective effects of RUS on HUVECs.Discussion: Effects of RUS on sepsis-induced impairments in endothelium relied on the suppression of miR-146a-5p.Keywords: endothelial dysfunction, ruscogenin, miR-146a-5p, NRP2, sepsis

Interaction of intestinal barrier dysfunction and intestinal inflammation promotes the progression of Crohn’s disease (CD). A more recent study has suggested that ruscogenins (RUS) can exert anti-inflammatory effects through activation of the Nrf2/NQO1 pathway. The current study is aimed at determining the functionalization of RUS on CD-like colitis. Wild-type (WT) mice induced with trinitrobenzene sulfonic acid (TNBS) exhibit a significant inflammation in their colon and are hence widely used for CD models. In the current study, the mice were treated with the Nrf-2 antagonist (ML385) or ruscogenin (RUS) whereas normal WT mice were kept as the negative control. Comparative analysis was then performed on the inflammation and barrier function of the colons. In vitro analysis of mouse colonic organoid systems revealed the influence of RUS on LPS-induced apoptosis, cytokine, and chemokine expressions in the intestinal epithelium. It was found that RUS ameliorates murine colitis through activation of the Nrf2/NQO1 pathway which was presented as a decrease in inflammation score and downregulated levels of cytokine and chemokine synthesis, as well as increased intestinal permeability. Further, it was noted that RUS alleviated LPS-induced apoptosis in the intestinal epithelium cells through upregulation of the Nrf2/NQO1 signaling pathway in the mouse colonic organoids. In addition, ruscogenin (RUS) attenuated the levels of Bax and C-caspase-3 through activation of the Nrf2/HO1 signaling pathway both in vivo and in vitro. Therefore, it was evident that RUS can be applied as a potential alternative therapeutic agent in CD based on its protective effects on the barrier function and anti-inflammatory activity.

Saponins, a diverse group of natural compounds, offer an interesting pool of derivatives with biomedical application. In this study, three structurally related spirostanol saponins were isolated and identified from the leek flowers of Allium porrum L. (garden leek). Two of them were identical with the already known leek plant constituents: aginoside (1) and 6-deoxyaginoside (2). The third one was identified as new component of A. porrum, however, it was found identical with yayoisaponin A (3) obtained earlier from a mutant of elephant garlic Allium ampeloprasun L. It is a derivative of the aginoside (1) with additional glucose in its glycosidic chain, identified by MS and NMR analysis as (2α, 3β, 6β, 25R)-2,6-dihydroxyspirostan-3-yl β-D-glucopyranosyl-(1 → 3)-β-D-glucopranosyl-(1 → 2)-[β-D-xylopyranosyl-(1 → 3)]-β-D-glucopyranosyl]-(1 → 4)-β-D-galactopyranoside, previously reported also under the name alliporin. The leek native saponins were tested together with other known and structurally related saponins (tomatonin and digitonin) and with their related aglycones (agigenin and diosgenin) for in vitro cytotoxicity and for effects on NO production in mouse peritoneal cells. The highest inhibitory effects were exhibited by 6-deoxyaginoside. The obtained toxicity data, however, closely correlated with the suppression of NO production. Therefore, an unambiguous linking of obtained bioactivities of saponins with their expected immunobiological properties remained uncertain.

Background/objectives: Steroidal saponins are widely distributed in medicinal plants with potential applications in cardiovascular disorders. Gitogenin, a saponin, has not been explored as antihype...

To explore the effect of ophiopogonin D on main fatty acid metabolic enzymes in human cardiomyocyte AC-16,so as to provide reference for cardiovascular protection mechanism and safe clinical application of Ophiopogon japonicus.CCK-8 (cell counting kit-8) was used to detect the effect of different concentrations of ophiopogonin D on the viability of cardiomyocytes.Meanwhile,the effect of different concentrations of ophiopogonin D on the morphology and quantity of cardiomyocytes was observed under microscope.The effect of ophiopogonin D on the mRNA expression of CYP2J2,CYP4F3,CYP4A11,CYP4A22 and CYP4F2 in cardiomyocytes was detected by RT-PCR.Western blot was used to detect the protein expression of CYP4F3 in different concentrations of ophiopogonin D.Compared with the control group,low-concentration ophiopogonin D had no effect on the viability of cardiomyocytes.However,ophiopogonin D with a concentration of higher than 20μmol·L~(-1)could promote the viability.Under the microscope,ophiopogonin D with a concentration of below 100μmol·L~(-1)had no significant effect on the morphology and number of cardiomyocytes.RT-PCR results showed that compared with the control group,5μmol·L~(-1)ophiopogonin D could slightly up-regulate mRNA expressions of CYP2J2 and CYP4F3,while high-concentration ophiopogonin D (10 and 20μmol·L~(-1)) could significantly induce mRNA expressions of CYP2J2and CYP4F3 in a dose-dependent manner (Plt;0.05).The same concentration of ophiopogonin D had a little effect on the mRNA expressions of CYP4A11,CYP4A22 and CYP4F2.Western blot results showed that 20μmol·L~(-1)ophiopogonin D could significantly induce the protein expression of CYP4F3 in a dose-dependent manner (Plt;0.05).Based on the above results,ophiopogonin D (less than100μmol·L~(-1)) has no effect on the viability of AC-16 cardiomyocytes.Ophiopogonin D (less than 100μmol·L~(-1)) can selectively induce the expressions of CYP2J2 and CYP4F3,regulate the metabolic pathway of fatty acid signaling molecules,and thus protecting the cardiovascular system.

Gut microbiota has been proven to play an important role in many metabolic diseases and cardiovascular disease, particularly atherosclerosis. Ophiopogonin D (OPD), one of the effective compounds in Ophiopogon japonicus, is considered beneficial to metabolic syndrome and cardiovascular diseases. In this study, we have illuminated the effect of OPD in ApoE knockout (ApoE[Formula: see text] mice on the development of atherosclerosis and gut microbiota. To investigate the potential ability of OPD to alleviate atherosclerosis, 24 eight-week-old male ApoE[Formula: see text] mice (C57BL/6 background) were fed a high-fat diet (HFD) for 12 weeks, and 8 male C57BL/6 mice were fed a normal diet, serving as the control group. ApoE[Formula: see text] mice were randomly divided into the model group, OPD group, and simvastatin group ([Formula: see text]= 8). After treatment for 12 consecutive weeks, the results showed that OPD treatment significantly decreased the plaque formation and levels of serum lipid compared with those in the model group. In addition, OPD improved oral glucose tolerance and insulin resistance as well as reducing hepatocyte steatosis. Further analysis revealed that OPD might attenuate atherosclerosis through inhibiting mTOR phosphorylation and the consequent lipid metabolism signaling pathways mediated by SREBP1 and SCD1 in vivo and in vitro. Furthermore, OPD treatment led to significant structural changes in gut microbiota and fecal metabolites in HFD-fed mice and reduced the relative abundance of Erysipelotrichaceae genera associated with cholesterol metabolism. Collectively, these findings illustrate that OPD could significantly protect against atherosclerosis, which might be associated with the moderation of lipid metabolism and alterations in gut microbiota composition and fecal metabolites.

Ethnopharmacological relevance Maidong (Liliaceae) is used as a yin-nourishing medication for the treatment of cardiovascular disease, inflammation, and assistant cancer chemotherapy in the clinic. Ophiopogonin B (OP-B), a major saponin extracted from Maidong, is reported to have potential antitumor activities against various human cancers. However, the effects of OP-B on human ovarian cancer (OC) and the potential mechanisms of action are yet elusive. Aim of the study: In this study, we aimed to explore the potential molecular mechanisms of OP-B in the treatment of OC using network pharmacology. In vivo and in vitro experiments were conducted to further verify the therapeutic effects of OP-B on OC. Materials and methods To investigate the functions of OP-B against OC holistically, the related targets of OP-B and OC were each predicted based on four public databases. Subsequently, the identified PPI network was constructed to detect the hub potential targets. In addition, GO and KEGG enrichment analysis were applied by Metascape database. Furthermore, we simultaneously investigated the anticancer effects of OP-B on SKOV3 and A2780 human ovarian cancer cells using a cell viability assay, transwell assay, and an image-based cytometric assay. The quantitative real-time PCR and western-blot assay were used to validate the RNA and protein levels of target genes in OP-B treated OC cells. At last, SKOV3-bearing BALB/c nude mice were applied to observe the effectiveness and toxicity of OP-B. Results Through network pharmacological analysis, OP-B was found to play a critical role in OC via multiple targets and pathways, especially the STAT3 signaling pathways. In addition, in vitro experiments found OP-B suppressed SKOV3 and A2780 cells proliferation in a time and concentration dependent manner, and markedly impaired cancer cell migration. Flow cytometry analysis revealed that OP-B significantly increased early and late apoptosis, induced G2/M phase cell cycle arrest in SKOV3 cells and G0/G1 phase cell cycle arrest in A2780 cells. Moreover, OP-B administration down-regulated the expression of p-STAT3 protein, whereas the RNA expression and total protein levels of STAT3 were not altered. Finally, in vivo experiments confirmed the therapeutic effects of OP-B on OC in nude mice with low toxicity in heart, liver, lung, and kidney. Conclusion OP-B could efficiently suppress OC cellular proliferation, migration and induce apoptosis, cell cycle arrest mainly via the regulation of STAT3 signaling pathway. This study provides a promising potential application for an alternative to chemotherapy in ovarian cancer.

Radix Ophiopogonis is often an integral part of many traditional Chinese formulas, such as Shenmai injection used to treat cardio-cerebrovascular diseases. This study aimed to investigate t...

Shenmai injection (SMI) is a patented traditional Chinese medicine that is extracted from Panax ginseng and Ophiopogon japonicus and is commonly used to treat cardiovascular diseases and tumors. The O. japonicus extract Ophiopogonin D' (OPD') is highly cardiotoxic. Mitochondria are central to OPD'-induced cardiotoxicity, although the precise mechanisms remain unclear. Excessive mitophagy activation and mitochondrial dysfunction lead to apoptosis, and the PTEN-induced kinase 1(PINK1)/Parkin pathway is critical in regulating mitophagy and mitochondrial function. We investigated the role of the PINK1/Parkin pathway in OPD'-induced mitochondrial damage and cardiotoxicity in AC16 cells. Concentrations of 2 μM OPD' and above inhibited cardiomyocyte viability and increased lactate dehydrogenase (LDH) release in a concentration- and time-dependent manner. OPD' was toxic to cells and mitochondria and increased the rate of apoptosis, triggering pyknosis, decreasing mitochondrial membrane potential (MMP), and decreasing the protein expression of the biogenesis regulator peroxisome proliferator-activated receptor γ coactivator-1 alpha (PGC-1α). The increased ratio of microtubule-associated proteins 1 A/1B light chain 3B (LC3-II/LC3-I) in mitochondria indicated that OPD' induced mitophagy. OPD' significantly induced oxidative stress and apoptosis, including increased reactive oxygen species (ROS) generation and decreased nuclear factor erythroid-2 related factor 2 (Nrf2), heme oxygenase-1(HO-1), and B-cell lymphoma 2 (Bcl-2) protein expression. OPD' activated the PINK1/Parkin pathway and promoted PINK1/Parkin translocation to mitochondria. Inhibiting mitophagy attenuated OPD'-induced PINK1/Parkin pathway activation and preserved mitochondrial biogenesis, consequently mitigating OPD'-induced mitochondrial dysfunction and apoptosis. These findings suggest that OPD'-induced cardiomyocyte mitophagy and mitochondrial damage are at least partially mediated by dysregulation of the PINK1/Parkin pathway.

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.

Background A crosstalk exists between diabetes and Alzheimer's disease (AD), and diabetic encephalopathy displays AD-like disorders. Sarsasapogenin (Sar) has strong anti-inflammatory efficacy, showing neuroprotection and memory-enhancement effects. Purpose This study aims to verify the ameliorative effects of Sar on diabetic encephalopathy in vivo and in vitro, and to clarify the mechanisms from attenuation of AD-like pathology. Methods Streptozotocin-induced type 1 diabetic rats and high glucose-cultured SH-SY5Y cells were used in this study. After Sar treatment (20 and 60 mg/kg) for consecutive 9 weeks, Morris water maze and novel object recognition tasks were performed. Hematoxylin-eosin staining was used for examining loss of neurons in CA1 area and ki67 expression for reflecting neurogenesis in DG area of hippocampus. Aβ production pathway and tau phosphorylation kinase cascade were examined in these two models. Results Sar improved learning and memory ability, loss of neurons and reduction of neurogenesis in the hippocampus of diabetic rats. Moreover, Sar suppressed Aβ overproduction due to up-regulation of BACE1 in protein and mRNA and tau hyperphosphorylation from inactivation of AKT/GSK-3β cascade in the hippocampus and cerebral cortex of diabetic rats and high glucose-cultured SH-SY5Y cells, and PPARγ antagonism abolished the effects of Sar on key molecules in the two pathways. Additionally, it was found that high glucose-stimulated Aβ overproduction was prior to tau hyperphosphorylation in neurons. Conclusion Sar alleviated diabetic encephalopathy, which was obtained through inhibitions of Aβ overproduction and tau hyperphosphorylation mediated by the activation of PPARγ signaling. Hence, Sar is a good candidate compound for AD-like disorders.

Podocyte injury following abnormal podocyte autophagy plays an indispensable role in diabetic nephropathy (DN), therefore, restoration of podocyte autophagy is considered as a feasible strategy for the treatment of DN. Here, we investigated the preventive effects of sarsasapogenin (Sar), the main active ingredient in Anemarrhena asphodeloides Bunge, on the podocyte injury in diabetic rats, and tried to illustrate the mechanisms underlying the effects in high glucose (HG, 40 mM)-treated podocytes (MPs). Diabetes model was established in rats with single streptozocin (60 mg· kg−1) intraperitoneal administration. The rats were then treated with Sar (20, 60 mg· kg−1· d−1, i.g.) or a positive control drug insulin (INS) (40 U· kg−1· d−1, i.h.) for 10 weeks. Our results showed that both Sar and insulin precluded the decreases of autophagy-related proteins (ATG5, Beclin1 and LC3B) and podocyte marker proteins (podocin, nephrin and synaptopodin) in the diabetic kidney. Furthermore, network pharmacology was utilized to assess GSK3β as the potential target involved in the action of Sar on DN and were substantiated by significant changes of GSK3β signaling in the diabetic kidney. The underlying protection mechanisms of Sar were explored in HG-treated MPs. Sar (20, 40 μM) or insulin (50 mU/L) significantly increased the expression of autophagy- related proteins and podocyte marker proteins in HG-treated MPs. Furthermore, Sar or insulin treatment efficiently regulated phosphorylation at activation and inhibition sites of GSK3β. To sum up, this study certifies that Sar meliorates experimental DN through targeting GSK3β signaling pathway and restoring podocyte autophagy.

Cholinesterase (ChE) inhibition is an important treatment strategy for Alzheimer’s disease (AD) as acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are involved in the pathology of AD. In the current work, ChE inhibitory potential of twenty-four natural products from different chemical classes (i.e., diosgenin, hecogenin, rockogenin, smilagenin, tigogenin, astrasieversianins II and X, astragalosides I, IV, and VI, cyclocanthosides E and G, macrophyllosaponins A-D, kokusaginin, lamiide, forsythoside B, verbascoside, alyssonoside, ipolamide, methyl rosmarinate, and luteolin-7-O-glucuronide) was examined using ELISA microtiter assay. Among them, only smilagenin and kokusaginine displayed inhibitory action against AChE (IC50 = 43.29 ± 1.38 and 70.24 ± 2.87 µg/mL, respectively). BChE was inhibited by only methyl rosmarinate and kokusaginine (IC50 = 41.46 ± 2.83 and 61.40 ± 3.67 µg/mL, respectively). IC50 values for galantamine as the reference drug were 1.33 ± 0.11 µg/mL for AChE and 52.31 ± 3.04 µg/mL for BChE. Molecular docking experiments showed that the orientation of smilagenin and kokusaginine was mainly driven by the interactions with the peripheral anionic site (PAS) comprising residues of hAChE, while kokusaginine and methyl rosmarinate were able to access deeper into the active gorge in hBChE. Our data indicate that similagenin, kokusaginine, and methyl rosmarinate could be hit compounds for designing novel anti-Alzheimer agents.

The sustained activation of the nuclear factor κB (NF-κB) signaling pathway has been observed in human inflammatory bowel disease (IBD). Ophiopogonin D (OP-D) is a small molecular compound isolated from Ophiopogon japonicus, a widely used herbal remedy. In this study, dextran sodium sulfate was used to make a mouse model of experimental colitis and verify the effect of OP-D on the mouse model of experimental colitis. Small molecule-protein molecular docking approaches were also used to discover the mechanisms underlying the OP-D-induced regulation of colitis. In colitis, the OP-D can inhibit the apoptosis of intestinal mucosa cells, restore the intestinal barrier, and alleviate inflammation. The molecular docking simulations showed that OP-D had a high affinity with the REL-homology domain of NF-κB-p65 that affected its translocation to the nucleus. In a cell study, the effects of OP-D on inflammation and barrier dysfunction were significantly decreased by a small interfering RNA targeting NF-κB-p65. Further, the LPS-induced increase in NF-κB-p65 in the nucleus was also significantly inhibited by OP-D. OP-D alleviated experimental colitis by inhibiting NF-κB. New insights into the pathogenesis and treatment options of colitis are provided through this study.

Nasopharyngeal carcinoma (NPC) is a common malignant tumor in South China and is characterized by a high death rate. Ophiopogonin B (OP‑B) is a bioactive component of Radix Ophiopogon japonicus, which is frequently used in traditional Chinese medicine to treat cancer. The present study aimed to examine the anti‑cancer properties of OP‑B on NPC cells. Cell viability and cell proliferation were measured using MTT and EdU assays. Flow cytometry was used to measure cell apoptosis, reactive oxygen species and mitochondrial membrane potential. Western blotting was used to investigate the expression of apoptosis and Hippo signaling pathway proteins. OP‑B inhibited the proliferation of NPC cells by inducing apoptosis and disturbing the mitochondrial integrity. OP‑B enhanced ROS accumulation. In addition, OP‑B promoted the expression of mammalian STE20‑like kinase 1, large tumor suppressor 1 and phosphorylated yes‑associated protein (YAP) and suppressed the expression of YAP and transcriptional enhanced associate domain in NPC cells. OP‑B increased the expression of forkhead box transcription factor O1 in the nuclear fraction. In conclusion, OP‑B has therapeutic potential and feasibility in the development of novel YAP inhibitors for NPC.

Lung cancer is a leading cause of tumor-associated death worldwide. Autophagy plays a key role in regulating lung cancer progression, and is a promising option for lung cancer treatment. Saponins are a group of naturally occurring plant glycosides, characterized by their strong foam-forming properties in aqueous solution, and exert various biological properties, such as anti-inflammation and anti-cancer. In the present study, we for the first time explored the effects of gitogenin (GIT), an important saponin derived from Tribulus longipetalus, on lung cancer progression both in vitro and in vivo. We found that GIT markedly reduced the proliferation and induced apoptosis in lung cancer cells through increasing the cleavage of Caspase-3 and poly (ADP-ribose) polymerases (PARPs). In addition, GIT-incubated lung cancer cells exhibited clear accumulation of autophagosome, which was essential for GIT-suppressed lung cancer. Mechanistically, GIT-induced autophagy initiation was mainly through activating AMP-activated protein kinase (AMPK) and blocking protein kinase B (AKT) signaling pathways, respectively. Moreover, the autophagic flux was disrupted in GIT-treated lung cancer cells, contributing to the accumulation of impaired autophagolysosomes. Importantly, we found that suppressing autophagy initiation could abolish GIT-induced cell death; however, autophagosomes accumulation sensitized lung cancer cells to cell death upon GIT treatment. More in vitro experiments showed that GIT led to reactive oxygen species (ROS) production in lung cancer cells, which was also involved in the modulation of apoptosis. The in vivo findings confirmed the effects of GIT against lung cancer progression with undetectable toxicity to organs. In conclusion, we provided new insights into the treatment of lung cancer, and GIT might be an effective strategy for future clinical application.

Ophiopogonin D (OPD), a compound from the Chinese herb Radix Ophiopogonis, reportedly induces increased levels of cytochrome P450 2J3 (CYP2J3)/epoxyeicosatrienoic acids (EETs) and Ca2+ in rat cardiomyocytes. Little is known regarding the specific mechanism between CYP2J3 and Ca2+ homeostasis. Here, we investigated whether CYP2J3 is involved in the protective action of OPD on the myocardium by activating the Ca2+ homeostasis-related protein complex (SERCA2a and PLB) in H9c2 rat cardiomyoblast cells. The interaction between SERCA2a and PLB was measured using fluorescence resonance energy transfer. OPD attenuated heart failure and catalyzed the active transport of Ca2+ into the sarcoplasmic reticulum by inducing the phosphorylation of PLB and promoting the SERCA2a activity. These beneficial effects of OPD on heart failure were abolished after knockdown of CYP2J3 in a model of heart failure. Together, our results identify CYP2J3 as a critical intracellular target for OPD and unravel a mechanism of CYP2J3-dependent regulation of intracellular Ca2+.

Ophiopogonin D (OPD) and Ophiopogonin D′ (OPD′) are two bioactive ingredients in Ophiopogon japonicus. Previously published studies have often focused on the therapeutic effects related to OPD's antioxidant capacity but underestimated the cytotoxicity-related side effects of OPD′, which may result in unpredictable risks. In this study, we reported another side effect of OPD′, hemolysis, and what was unexpected was that this side effect also appeared with OPD. Although hemolysis effects for saponins are familiar to researchers, the hemolytic behavior of OPD or OPD′ and the interactions between these two isomers are unique. Therefore, we investigated the effects of OPD and OPD′ alone or in combination on the hemolytic behavior in vitro and in vivo and adopted chemical compatibility and proteomics methods to explain the potential mechanism. Meanwhile, to explain the drug-drug interactions (DDIs), molecular modeling was applied to explore the possible common targets. In this study, we reported that OPD′ caused hemolysis both in vitro and in vivo, while OPD only caused hemolysis in vivo. We clarified the differences and DDIs in the hemolytic behavior of the two isomers. An analysis of the underlying mechanism governing this phenomenon showed that hemolysis caused by OPD or OPD′ was related to the destruction of the redox balance of erythrocytes. In vivo, in addition to the redox imbalance, the proteomics data demonstrated that lipid metabolic disorders and mitochondrial energy metabolism are extensively involved by hemolysis. We provided a comprehensive description of the hemolysis of two isomers in Ophiopogon japonicus, and risk warnings related to hemolysis were presented. Our research also provided a positive reference for the development and further research of such bioactive components.

Previously, various steroidal glycosides were reported from plants of Cestrum species. However, phytochemical investigation has not been conducted on Cestrum newellii. A systematic phytochemical investigation of the leaves of C. newellii resulted in the isolation of eight novel steroidal glycosides (1–8), which were classified into three spirostanol glycosides (1–3), two furostanol glycosides (4 and 5), two pseudofurostanol glycosides (6 and 7), and one cholestane glycoside (8). In addition, three known cholestane glycosides (9–11) were isolated and identified. The structures of the new compounds were determined based on spectroscopic data and chemical transformations. Compounds 1 and 2 are spirostanol glycosides having hydroxy groups at C-2, C-3, C-12, and C-24 of the aglycone moiety. Although C. newellii is known to be a poisonous plant, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay exhibited that none of the isolated compounds were cytotoxic to HL-60 human promyelocytic leukemia cells.

Many Chinese herbs are well known for their neuroprotective and anti-oxidant properties. Extracts of Salvia miltiorrhiza and Anemarrhenae asphodeloides, tanshinone IIA (tanIIA), salvianolic acid B (Sal B) and sarsasapogenin (ML-1), were selected to study their dissociation potential towards Aβ42 peptide fibrils and neuroprotective effect on cells. Moreover, derivatives of sarsasapogenin (ML-2, ML-3 and ML-4) have been prepared by the addition of modified carbamate moiety. TanIIA and Sal B have shown to possess a strong ability to dissociate Aβ42 fibrils. The dissociation potential of ML-1 increased upon the introduction of carbamate moiety with N-heterocycles. In silico data revealed that derivatives ML-4 and Sal B interact with Aβ42 regions responsible for fibril stabilization through hydrogen bonds. Contrary, tanIIA binds close to a central hydrophobic region, which may lead to destabilization of fibrils. Sarsasapogenin derivative ML-2 decreased nitride oxide production, and derivative ML-4 enhanced the growth of neurites. The reported data highlight the possibility of using active compounds to design novel treatment agents for Alzheimer's disease.

Yucca schidigera Roezl (Mojave), a kind of ornamental plant belonging to the Yucca genus (Agavaceae), whose extract exhibits important roles in food, beverage, cosmetic and feed additives owing to its rich spirostanol saponins. To provide a comprehensive chemical profiling of the spirostanol saponins in it, this study was performed by using a multi-phase liquid chromatography method combining a reversed phase chromatography T3 column with a normal phase chromatography silica column for the separation and an ESI-Q-Exactive-Orbitrap MS in positive ion mode as the detector. By comparing the retention time and ion fragments with standards, thirty-one spirostanol saponins were identified. In addition, according to the summary of the chromatographic retention behaviors and the MS/MS cleavage patterns and biosynthetic pathway, another seventy-nine spirostanol saponins were speculatively identified, forty ones of which were potentially new ones. Moreover, ten novel spirostanol saponins (three pairs of (25R/S)-spirostanol saponin isomer mixtures) were targeted for isolation to verify the speculation. Then, the comprehensive chemical profiling of spirostanol saponins from Y. schidigera was reported here firstly.

An efficient method of thiol group introduction to the structure of common natural products and synthetic active compounds with recognized biological efficacy such genistein (1), 5,11-dimethyl-5H-indolo[2,3-b]quinolin (2), capecitabine (3), diosgenin (4), tigogenin (5), flumethasone (6), fluticasone propionate (7), ursolic acid methyl ester (8), and &beta, sitosterol (9) was developed. In most cases, the desired compounds were obtained easily via two-step processes involving esterification reaction employing S-trityl protected thioacetic acid and the corresponding hydoxy-derivative, followed by removal of the trityl-protecting group to obtain the final compounds. The results of our preliminary experiments forced us to change the strategy in the case of genistein (1), and the derivatization of diosgenin (4), tigogenin (5), and capecitabine (3) resulted in obtaining different compounds from those designed. Nevertheless, in all above cases we were able to obtain thiol-containing derivatives of selected biological active compounds. Moreover, a modelling study for the two-step thiolation of genistein and some of its derivatives was accomplished using the density functional theory (B3LP). A hypothesis on a possible reason for the unsuccessful deprotection of the thiolated genistein is also presented based on the semiempirical (PM7) calculations. The developed methodology gives access to new sulphur derivatives, which might find a potential therapeutic benefit.

Ethnopharmacological relevance Shengmai San (SMS) has been commonly used as a traditional Chinese medicine for the treatment of cardiovascular disorders, of which drug interactions need to be assessed for the safety concern. There is little evidence for the alterations of hepatic and intestinal drug-metabolizing enzymes after repeated SMS treatments to assess drug interactions. Aim of the study The studies aim to illustrate the effects of repeated treatments with SMS on cytochrome P450s (CYPs), reduced nicotinamide adenine dinucleotide (phosphate)-quinone oxidoreductase (NQO), uridine diphosphate-glucuronosyltransferase (UGT), and glutathione S-transferase (GST) using in vivo rat model. Materials and methods The SMS was prepared using Schisandrae Fructus, Ginseng Radix, and Ophiopogonis Radix (OR) (1:2:2). Chromatographic analyses of decoctions were performed using ultra-performance liquid chromatography (UPLC) and LC-mass spectrometry. Sprague-Dawley rats were orally treated with the SMS and its component herbal decoctions for 2 or 3 weeks. Hepatic and intestinal enzyme activities were determined. CYP3A expression and the kinetics of intestinal nifedipine oxidation (NFO, a CYP3A marker reaction) were determined. Results Schisandrol A, schisandrin B, ginsenoside Rb1 and ophiopogonin D were identified in SMS. SMS selectively suppressed intestinal, but not hepatic, NFO activity in a dose- and time-dependent manner. Hepatic and intestinal UGT, NQO and GST activities were not affected. A 3-week SMS treatment decreased the maximal velocity of intestinal NFO by 50%, while the CYP3A protein level remained unchanged. Among SMS component herbs, the decoction of OR decreased intestinal NFO activity. Conclusions These findings demonstrate that 3-week treatment with SMS and OR suppress intestinal, but not hepatic CYP3A function. It suggested that the potential interactions of SMS with CYP 3A drug substrates should be noticed, especially the drugs whose bioavailability depends heavily on intestinal CYP3A.

OBJECTIVES CD31hi EMCNhi vessels (CD31, also known as PECAM1 [platelet and endothelial cell adhesion molecule 1]; EMCN, endomucin), which are strongly positive for CD31 and endomucin, couple angiogenesis and osteogenesis. However, the role of CD31hi EMCNhi vessels in bone regeneration remains unknown. In the present study, we investigated the role of CD31hi EMCNhi vessels in the process of bone regeneration. MATERIALS AND METHODS We used endothelial-specific Kruppel like factor 3 (Klf3) knockout mice and ophiopogonin D treatment to interfere with CD31hi EMCNhi vessel formation. We constructed a bone regeneration model by surgical ablation of the trabecular bone. Immunofluorescence and micro-computed tomography (CT) were used to detect CD31hi EMCNhi vessels and bone formation. RESULTS CD31hi EMCNhi vessels participate in the process of bone regeneration, such that endothelial-specific Klf3 knockout mice showed increased CD31hi EMCNhi vessels and osteoprogenitors in the bone regeneration area, and further accelerated bone formation. We also demonstrated that the natural compound, ophiopogonin D, acts as a KLF3 inhibitor to promote vessels formation both in vitro and in vivo. Administration of ophiopogonin D increased the abundance of CD31hi Emcnhi vessels and accelerated bone healing. CONCLUSIONS Our findings confirmed the important role of CD31hi Emcnhi vessels in bone regeneration and provided a new target to treat bone fracture or promote bone regeneration.