Marsdeniae tenacissimae extract(MTE) suppresses cell proliferation by attenuating VEGF/VEGFR2 interactions and promotes apoptosis through regulating PKC pathway in human umbilical vein endothelial cells

Marsdeniae tenacissimae extract(MTE), commonly known as Xiao-Ai-Ping in China, is a traditional Chinese herb medicine capable of inhibiting proliferation and metastasis and boosting apoptosis in various cancer cells. However, little is known about the contribution of MTE towards tumor angiogenesis and the underlying mechanism. The present study aimed to evaluate the effects of MTE on the proliferation and apoptosis of human umbilical vein endothelial cells(HUVECs) and the molecular ism. 3-(4,5-dimethylthiazol-2-yl)-5(3-carboxymethoxyphenyl)-2-(4-sulfopheny)-2H-tetrazolium, inner salt(MTS) and PI-stained flow cytometry assays revealed that MTE dose-dependently reduced the proliferation of HUVECs by arresting cell cycle at S phase(P < 0.05). Annexin V-FITC/PI-stained flow cytometry confirmed that MTE(160 μL·m L-1) enhanced the apoptosis of HUVECs significantly(P < 0.001). Real-time quantitative RT-PCR and Western blot analyses showed an increase in Bax expression and a sharply decline in Bcl-2 expression; caspase-3 was activated simultaneously in a dose-dependent manner(P < 0.05). Further study observed the dose-dependent down-regulation of vascular endothelial growth factor(VEGF) receptor-2(VEGFR-2), P2Y6 receptor(P2Y6R), and chemokine(C-C motif) ligand 2(CCL-2), along with the activation of PKC δ and up-regulation of p53 in a dose-dependent manner in MTE-treated selected cells(P < 0.05). Collectively, the results from the present study suggested that MTE suppressed the proliferation by attenuating CCL-2-mediated VEGF/VEGFR2 interactions and promoted the apoptosis through PKCδ-induced p53-dependent mitochondrial pathway in HUVECs, supporting that MTE may be developed as a potent anti-cancer medicine.

Cytotoxicity of anti-tumor herbal Marsdeniae tenacissimae extract on erythrocytes

Marsdeniae tenacissimae extract （MTE） has been used as an adjuvant medicine for cancer therapy for a long time. Although massive studies demonstrated its considerable anti-cancer effect, there is no research on its influence on erythrocytes, which are firstly interacted with MTE in the circulation. To investigate the influence of MTE on erythrocytes, we used a flow cytometer to detect the MTE-treated alternations of morphology, calcium concentration, and reactive oxygen species （ROS） level in erythrocytes. We used hemolysis under different osmotic solutions to evaluate the fragility of erythrocytes. Data showed that MTE treatment dose-dependently increased the ratio of erythrocyte fragmentation （P〈0.001） and shrinking, and elevated the forward scatter （FSC） value （P〈0.001） and calcium accumulation （P〈0.001）. MTE induced ROS production of erythrocytes under the high glucose condition （P〈0.01） and consequently caused a rise in fragility （P〈0.05）. These results suggest that MTE induces cytotoxicity and aging in erythrocytes in a dose-dependent manner, and presents the possibility of impairment on cancer patients＇ circulating erythrocytes when MTE is used as an anti-cancer adjuvant medicine.

Clinical value of Tongguanteng(Radix seu Herba Marsdeniae Tenacissimae) extract combined with chemotherapy in the treatment of advanced non-small cell lung cancer： a Meta-analysis

OBJECTIVE: To investigate the clinical efficacy and safety of Tongguanteng(Radix seu Herba Marsdeniae Tenacissimae) extract combined with chemotherapy in the treatment of advanced non-small celllung cancer(NCSLC) compared with chemotherapy alone.METHODS: Databases including Chinese National Knowledge Infrastructure, China Biology Medicine Disc, Wanfang, and MEDLINE were searched until April 1, 2014. Two assessors independently reviewed each trial. The primary outcome was the effective rate(ER) of Tongguanteng(Radix seu Herba Marsdeniae Tenacissimae) extract combined with chemotherapy. The secondary outcomes included quality of life improvement rate(QOLIR) and adverse reactions. Statistical calculations were performed by using Cochrane Collaboration Review Manager 5.2.RESULTS: A total of 888 patients from 15 studies,13 randomized controlled trials(RCT) and two controlled clinical trials, were included. Compared with chemotherapy alone, Tongguanteng(Radix seu Herba Marsdeniae Tenacissimae) extract plus chemotherapy significantly improved ER [Risk ratio(RR) =1.32, 95% CI,(1.14, 1.54)](based on 15 studies) and QOLIR [RR = 2.04, 95% CI,(1.69, 2.47)](based on 13studies). Compared with chemotherapy alone,Tongguanteng(Radix seu Herba Marsdeniae Tenacissimae) extract plus chemotherapy significantly inhibited chemotherapy-induced white blood cell decline [RR = 0.79, 95% CI,(0.70, 0.90)(based on 10 studies), chemotherapy-induced platelet decline[RR = 0.77, 95% CI,(0.60, 0.98)](based on 8 studies),and significantly alleviated nausea and vomiting(NV) [RR = 0.83, 95% CI,(0.71, 0.97)](based on 7studies). There was no significant difference in hemoglobin decline between the two therapies [RR =0.88, 95% CI,(0.70, 1.09)](based on 6 studies).CONCLUSION: This Meta-analysis suggests that Tongguanteng(Radix seu Herba Marsdeniae Tenacissimae) extract combined with chemotherapy may be more efficacious in the treatment of advanced NSCLC than chemotherapy alone. This effect includes enhancing ER and QOLIR, and weakening chemotherapy toxicity. However, large-scale RCTs are required to further investigate the short- and long-term effects of Tongguanteng(Radix seu Herba Marsdeniae Tenacissimae) extract.

Two New C_(21) Steroids from Marsdenia tenacissima

Two new C21 steroids were isolated from the CHCl3 extract of the stem of Marsdenia tenacissima. On the basis of spectroscopic analysis and chemical methods, their structures were elucidated as 17β-tenacigenin B (2) and 3-O-6-deoxy-3-O-methyl-β-D-allopyranosyl-(1→4)-β-D- oleandropyranosyl-tenacigenin C (3). The structure of the known aglycon tenacigenin C was revised as 5α, 9α, 17β-pregnane-3β, 8β, 11α, 12β, 14β-pentanol-20-one. Compound 3 is the first reported glycoside of tenacigenin C.

Polyoxypregnanes from the stems of Marsdenia tenacissima

From the stems ofMarsdenia tenacissima two new polyoxypregnanes were isolated, their structures were elucidated by 1D, 2D-NMR as 11α,12β-di-O-tigloyl-tenacigenin B （1） and tenacigenoside E （2）.

A new C_(21) steroid glycoside from Marsdenia tenacissima

A new C21 steroid glycoside,11α-O-tigloyl-12-β-O-acetyl tenacigenin B 3-O-β-D-glucopyranosyl-（1→4）-β-D-glucopyr- anosyl-（1→4）-3-O-methyl-6-deoxy-β-D-allopyranosyl-（1→4）-β-D-oleandropyranoside（1）,named tenacissoside N was isolated from the stems of Marsdenia tenacissima.The structure of the glycoside was identified by HR-ESI-MS and NMR spectra.

Research progress on anti-tumor properties of Marsdenia tenacissima

Tongguanteng （Marsdenia tenacissima）, which is mainly distributed in the Yunnan and Guizhou provinces of China, wasfirst recorded in Diannanbencao by Lan Mao of the Ming dynasty of China. According to recent pharmacological studies,the chemical composition of Tongguanteng （Marsdenia tenacissima） is complex and contains C21 steroidal saponins,polysaccharides, alkaloids, and other molecules, which show anti-cancer effects on various tumor cell lines. It inhibitstumor cell proliferation and growth mainly by increasing the expression of apoptosis- and cell cycle-related proteins topromote apoptosis and arrest tumor cells in the G2/M or S phase. Downregulation of the expression of vascularendothelial growth factor-2/A and matrix metalloprotease-2/9 suppresses the formation of the tumor microvasculature,leading to tumor malnutrition, increased expression of interleukin-2, glutathione peroxidase, catalase, and superoxidedismutase, and decreased interleukin-10 and malondialdehyde expression, thereby enhancing immunity andantioxidation in the body. Additionally, inhibition of epidermal growth factor receptor, hepatocyte growth factor receptor,and tyrosine-protein kinase receptor activation enhances the anti-tumor efficacy of epidermal growth factorreceptor-tyrosine kinase inhibitors as well as inhibits P-glycoprotein and cytochrome P450 to increase the concentrationof anti-tumor drugs in tumor cells.

傣药“雅解”:宾蒿、傣百解及竹叶兰的生物活性及其解毒机制

傣药“雅解”是傣族传统医学中最具特色的药物和治疗方法。其中,宾蒿(Clerodendrum chinense var.simplex)、傣百解(Marsdenia tenacissima)和竹叶兰(Arundina graminifolia)作为3种常见的“雅解”被广泛使用。本研究使用石油醚、乙酸乙酯、正丁醇或水依次萃取3种“雅解”的乙醇提取物获得各极性部位,并制备3种“雅解”的水煎剂。经测定3种“雅解”的总黄酮质量分数为22.41~586.39 mg/g,高于总多酚质量分数(2.76~28.66 mg/g)。3种“雅解”的提取物均可清除DPPH、·OH和ABTS自由基,其中对ABTS自由基的清除率高于其他自由基(IC50>380μg/mL)。它们对大肠杆菌、铜绿假单胞菌和金黄色葡萄球菌的抑制作用较弱。此外,3种“雅解”具有一定的抗炎作用,在60μg/mL的质量浓度下即可显著下调脂多糖刺激巨噬细胞产生的NO、肿瘤坏死因子-α、白细胞介素-1β和白细胞介素-6。综上所述,本研究通过现代科学手段初步揭示了3种“雅解”的解毒作用机理,为“雅解”的进一步优化与研究奠定了基础。

通关藤的适宜采收期研究

测定不同生长期通关藤(Marsdenia tenacissima)的茎粗和茎长,采用香草醛-高氯酸法显色,在462 nm处测定通关藤中总皂苷的含量,研究通关藤的最适宜采收期。结果表明,通关藤总皂苷含量在两年生时达到最高,以后缓慢下降;产量在三年生时增长最快,之后增长缓慢。通关藤的最佳采收期要兼顾其产量和有效成分总皂苷的含量,两者都在相对较高的时候才是最佳采收期,确定其最佳采收期为两年半生至三年生的藤茎。

Marsdenia tenacissima injection induces the apoptosis of prostate cancer by regulating the AKT/GSK3β/STAT3 signaling axis

Marsdenia tenacissima injection,a standard Marsdenia tenacissima extract(MTE),has been approved as an adjuvant therapeutic agent for various cancers.Our previous study showed that MTE inhibited the proliferation and metastasis of prostate cancer(PCa)cells.However,the underlying mechanisms and active ingredients of MTE against PCa were not completely understood.This study revealed that MTE induced significant decreases in cell viability and clonal growth in PCa cells.In addition,MTE induced the apoptosis of DU145 cells by reducing the mitochondrial membrane potential and increasing the expression of Cleaved Caspase 3/7,Cyt c,and Bax.In vivo,DU145 xenografted NOD-SCID mice treated with MTE showed significantly decreased tumor size.TUNEL staining and Western blot confirmed the pro-apoptotic effects of MTE.Network pharmacology analysis collected 196 ingredients of MTE linked to 655 potential targets,and 709 PCa-associated targets were retrieved,from which 149 overlapped targets were screened out.Pathway enrichment analysis showed that the HIF-1,PI3K-AKT,and ErbB signaling pathways were closely related to tumor apoptosis.Western blot results confirmed that MTE increased the expression of p-AKT^(Ser473) and p-GSK3β^(Ser9),and decreased the expression of p-STAT3^(Tyr705) in vitro and in vivo.A total of 13 compounds in MTE were identified by HPLC-CAD-QTOF-MS/MS and UPLCQTOF-MS/MS.Molecular docking analysis indicated that six compounds may interact with AKT,GSK3β,and STAT3.In conclusion,MTE induces the endogenous mitochondrial apoptosis of PCa by regulating the AKT/GSK3β/STAT3 signaling axis,resulting in inhibition of PCa growth in vitro and in vivo.

Marsdenia tenacissima extract induces G_0/G_1 cell cycle arrest in human esophageal carcinoma cells by inhibiting mitogen-activated protein kinase(MAPK) signaling pathway

Marsdenia tenacissima extract(MTE, trade name: Xiao-Ai-Ping injection) is an extract of a single Chinese plant medicine. It has been used for the treatment of cancer in China for decades, especially for esophageal cancer and other cancers in the digestive tract. In the present study, the potential mechanism for MTE's activity in esophageal cancer was explored. The effects of MTE on the proliferation of human esophageal cancer cells(KYSE150 and Eca-109) were investigated by the MTT assay, the Brd U(bromodeoxyuridine) incorporation immunofluorescence assay, and flow cytometric analysis. MTE inhibited cell proliferation through inducing G0/G1 cell cycle arrest in KYSE150 and Eca-109. Western blot analysis was employed to determine protein levels in the MTE treated cells. Compared with the control cells, the expression levels of the cell cycle regulatory proteins cyclin D1/D2/D3, cyclin E1, CDK2/4/6(CDK: cyclin dependent kinase), and p-Rb were decreased significantly in the cells treated with MTE at 40 mg·m L-1. In addition, MTE had an inhibitory effect on the MAPK(mitogen-activated protein kinase) signal transduction pathway, including ERK(extracellular signal-regulated kinase), JNK(c-Jun N-terminal kinase), and p38 MAPK. Moreover, MTE showed little additional effects on the regulation of cyclin D1/D3, CDK4/6, and p-Rb when the ERK pathway was already inhibited by the specific ERK inhibitor U0126. In conclusion, these data suggest that MTE inhibits human esophageal cancer cell proliferation through regulation of cell cycle regulatory proteins and the MAPK signaling pathways, which is probably mediated by the inhibition of ERK activation.

Pregnane Glycosides from Stems of Marsdenia tenacissima

Objective To study the chemical constituents from the stems of Marsdenia tenacissima.Methods The chemical constituents were isolated by various column chromatography and their structures were identified by spectral and chemical analysis.Results Two pregnane glycosides were isolated from the stems of M.tenacissima and identified as 3-O-β-D-glucopyranosyl-(1→4)-6-deoxy-3-O-methyl-β-D-allopyranosyl-(1→4)-β-D-oleandropyranosyl-11α-O- tigloyltenacigenin B,named as tenacigenoside I(1)and 3-O-β-D-glucopyranosyl-(1→4)-6-deoxy-3-O-methyl-β-D- allopyranosyl-(1→4)-β-D-oleandropyranosyl-11α,12β-di-O-acetyltenacigenin B,named as tenacigenoside K(2).Conclusion Compound 1 is a new compound,1H-NMR and 13C-NMR data of compound 2 are reported for the first time.

Chemical constituents from Xiaoaiping injection

Xiaoaiping injection is a single-prescription of Marsdenia tenacissima. One new compound, tenacigenoside L （1）, along with ten known compounds were isolated from the CHCl3-soluble fraction of Xiaoaiping injection extract by silica gel, ODS, Sephadex LH-20 and semi-preparative HPLC chromatography. The new compound was characterized as 3-O-β-D-oleandropyranosyl- 17β-tenacigenin B. The ten known compounds were identified as tenacigenin B （2）, 17β-tenacigenin B （3）, marsdenoside F （4）, tenacissoside G （5）, tenacissoside I （6）, tenacissoside H （7）, marsdenoside D （8）, tenacigenin A （9）, marsdenoside I （10）, and tenacigenin C （11）. The structures of 11 compounds were elucidated on the basis of extensive analyses of spectroscopic data including MS, 1D NMR and 2D NMR, and comparison of their spectroscopic data with those reported in the literatures.

Polyoxypregnanes as safe, potent, and specific ABCB1-inhibitory pro-drugs to overcome multidrug resistance in cancer chemotherapy in vitro and in vivo

Multidrug resistance(MDR)mediated by ATP binding cassette subfamily B member 1(ABCB1)is significantly hindering effective cancer chemotherapy.However,currently,no ABCB1-inhibitory drugs have been approved to treat MDR cancer clinically,mainly due to the inhibitor specificity,toxicity,and drug interactions.Here,we reported that three polyoxypregnanes(POPs)as the most abundant constituents of Marsdenia tenacissima(M.tenacissima)were novel ABCB1-modulatory pro-drugs,which underwent intestinal microbiota-mediated biotransformation in vivo to generate active metabolites.The metabolites at non-toxic concentrations restored chemosensitivity in ABCB1-overexpressing cancer cells via inhibiting ABCB1 efflux activity without changing ABCB1 protein expression,which were further identified as specific non-competitive inhibitors of ABCB1 showing multiple binding sites within ABCB1 drug cavity.These POPs did not exhibit ABCB1/drug metabolizing enzymes interplay,and their repeated administration generated predictable pharmacokinetic interaction with paclitaxel without obvious toxicity in vivo.We further showed that these POPs enhanced the accumulation of paclitaxel in tumors and overcame ABCB1-mediated chemoresistance.The results suggested that these POPs had the potential to be developed as safe,potent,and specific pro-drugs to reverse ABCB1-mediated MDR.Our work also provided scientific evidence for the use of M.tenacissima in combinational chemotherapy.