Abstract Background Propolis and Hypericum perforatum L. are natural products which contain many active compounds and have numerous beneficial effects, including an antitumor effect. Gliobmastoma multiforme (GBM) is a common primary brain tumor with poor prognosis and limited treatment options. In this study, the effect of propolis (EEP) combined with H. perforatum L. (HPE) on glioblastoma cell line U87MG was investigated for the first time.
Methods Anti-proliferative activity of EEP, HPE and their combination (EEP + HPE) was determined by a cytotoxicity test, DNA binding by [3H]-thymidine incorporation and cell migration assay. Anti-metastatic properties in U87MG treated with EEP, HPE and EEP + HPE were estimated on cells migration test (scratch assay) and metalloproteinases (MMP2 and MMP9) secretion (gelatin zymography).
Results Combination of HPE and EEP extracts was found to have a time- and dose-dependent inhibitory effect on the viability of U87MG cells. This effect was significantly higher (p < 0.05) when compared to these two extracts applied separately, which was confirmed by the significant reduction of DNA synthesis and significantly higher mitochondrial membrane permeabilization. A significant decreasing in migration cells and in pro-MMP9 and pro-MMP2 secretion in U87MG cells were demonstrated after exposure to combination of EEP (30 μg/ml) with HPE (6.25 μg/ml).
Conclusions In this study, the combination of ethanolic extract from propolis and ethanolic extract of fresh-cut H. perforatum L. was proved the ability to reduce invasiveness of glioma cells through the inhibition of MMP2 and MMP9 secretion and suppression of cell migration. It has a more potent anti-proliferative effect on U87MG glioma cell line compared to using propolis and H. perforatum L. separately. Further studies are required to verify whether the examined extracts can activate apoptotic pathways.
Bee products prevent VEGF-induced angiogenesis in human umbilical vein endothelial cells
Hiroshi Izuta,1 Masamitsu Shimazawa,1 Kazuhiro Tsuruma,1 Yoko Araki,2 Satoshi Mishima,2 and Hideaki Hara11 Department of Biofunctional Evaluation, Molecular Pharmacology, Gifu Pharmaceutical University, 5-6-1 Mitahora-higashi, Gifu 502-8585, Japan2Nagaragawa Research Center, Api Co. Ltd., 692-3 Nagara, Gifu 502-0071, Japan
Background Vascular endothelial growth factor (VEGF) is a key regulator of pathogenic angiogenesis in diseases such as cancer and diabetic retinopathy. Bee products [royal jelly (RJ), bee pollen, and Chinese red propolis] from the honeybee, Apis mellifera, have been used as traditional health foods for centuries. The aim of this study was to investigate the anti-angiogenic effects of bee products using human umbilical vein endothelial cells (HUVECs).
Methods In an in vitro tube formation assay, HUVECs and fibroblast cells were incubated for 14 days with VEGF and various concentrations of bee products [RJ, ethanol extract of bee pollen, ethanol extract of Chinese red propolis and its constituent, caffeic acid phenethyl ester (CAPE)]. To clarify the mechanism of in vitro angiogenesis, HUVEC proliferation and migration were induced by VEGF with or without various concentrations of RJ, bee pollen, Chinese red propolis, and CAPE.
Results RJ, bee pollen, Chinese red propolis, and CAPE significantly suppressed VEGF-induced in vitro tube formation in the descending order: CAPE > Chinese red propolis >> bee pollen > RJ. RJ and Chinese red propolis suppressed both VEGF-induced HUVEC proliferation and migration. In contrast, bee pollen and CAPE suppressed only the proliferation.
Conclusion Among the bee products, Chinese red propolis and CAPE in particular showed strong suppressive effects against VEGF-induced angiogenesis. These findings indicate that Chinese red propolis and CAPE may have potential as preventive and therapeutic agents against angiogenesis-related human diseases.
More ..... In a previous study, we found that Brazilian green propolis and its constituent caffeoylquinic acid derivatives suppressed VEGF-induced cell proliferation, migration, and in vitro tube formation in HUVECs . In the present study, Chinese red propolis and its chemical constituent, CAPE, suppressed VEGF-induced in vitro tube formation. Caffeic acid is a known suppressor of tumor angiogenesis that acts in human retinal carcinoma cells by blocking STAT3-mediated VEGF expression . These studies indicate that caffeoyl groups included in propolis may be important components responsible for its anti-angiogenic activities.The pharmacological effects of bee products have been reported in many different diseases. In particular, there are many reports about the anti-tumor effects of propolis and CAPE. Our previous studies indicate that baccharin and drupanin, constituents of Brazilian propolis, inhibit tumor growth both in vitro and in vivo . Likewise, CAPE induces growth arrest and apoptosis of colon cancer cells via the beta-catenin/T-cell factor signaling pathway . Tumor angiogenesis is a very important step in the growth and metastasis of tumor development. Combined with these findings in the present study, Chinese propolis and its CAPE constituent suppressed VEGF-induced angiogenesis in HUVECs indicates that the anti-tumor effects of propolis and CAPE may be dependent both on direct inhibition of tumor cell growth and on angiostatic effects on the vessels supplying nutrients to the neoplasm.
a Department of Pharmacology and Toxicology, Cancer Research Institute, Tohoku Pharmaceutical University; b Department of Pharmacology, Tohoku Pharmaceutical University; and c Department of Educational Center, Tohoku Pharmaceutical University; Sendai 981–8558, Japan. Received November 6, 2003; accepted January 30, 2004
We have investigated the effect of propolis (CB Propolis) on the growth of human histiolytic lymphoma U937cells. We found that propolis strongly inhibited the growth of the cells and macromolecular synthesis in a dose and time-dependent manner by apoptosis. Propolis at 0.015—0.5m l/ml showed antitumor activity with an IC50 of 0.18m l/ml for 3 d. It also inhibits DNA, RNA and protein synthesis with an IC50 of 0.08, 0.17 and 4.3m l/ml, respectively. The inhibitory effect on DNA synthesis was partially irreversible. Moreover, an apoptotic DNA ladder and chromatin condensation were observed in the same concentration range in which cell growth was inhibited. The caspase inhibitor, Z-Asp-CH2-DCB, prevented DNA fragmentation. These results suggest that the antitumor activity of propolis occurs through the induction of apoptosis. Propolis may be useful as a cancer chemopreventive and chemotherapeutic agent.
Comparison of Radical Scavenging Activity, Cytotoxic Effects and Apoptosis Induction in Human Melanoma Cells by Taiwanese Propolis from Different Sources
Chia-Nan Chen1, Meng-Shih Weng1, Chia-Li Wu2 and Jen-Kun Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, Section 1 Jen-Ai Road, Taipei 100, Taiwan, and 2Department of Chemistry, Tamkang University Tamsui 251, Taiwan
Propolis is a sticky substance that is collected from plants by honeybees. We previously demonstrated that propolins A, B,C, D, E and F, isolated from Taiwanese propolis (TP), could effectively induce human melanoma cell apoptosis and were strong antioxidant agents. In this study, we evaluated TP for freeradical scavenging activity by DPPH (1,2-diphenyl-2-picrylhydrazyl).The phenolic concentrations were quantified by the Folin–Ciocalteu method. The apoptosis trigger activity in human melanoma cells was evaluated. TP contained a higher level of phenolic compoundsand showed strong capability to scavenge free radicals. Additionally,TP1g, TP3, TP4 and TP7 exhibited a cytotoxic effect on human melanoma cells, with an IC50 of 2.3, 2.0, 3.3 and 3.3 µg/ml,respectively. Flow cytometric analysis for DNA fragmentation indicated that TP1g, TP2, TP3 and TP7 could induce apoptosis in human melanoma cells and there is a marked loss of cells from the G2/M phase of the cell cycle. To address the mechanism of the apoptosis effect of TP, we evaluated its effects on induction of apoptosis-related proteins in human melanoma cells. The levels of procaspase-3 and PARP [poly(ADP-ribose) polymerase] were markedly decreased. Furthermore, propolins A, B, C, D, E andF in TP were determined using HPLC. The results indicate that TP is a rich source of these compounds. The findings suggestthat TP induces apoptosis in human melanoma cells due to its high level of propolins.
Inhibitory effects of caffeic acid phenethyl ester (CAPE) on 12-0-tetradecanoylphorbol-13-acetate-induced tumor promotion in mouse skin and the synthesis of DNA, RNA and protein in HeLa cells
Mou-Tuan Huang, Wei Ma, Patricia Yen, Jian-Guo Xie, Jingkang Han1, Krystyna Frenkel1, Dezider Grunberger, and Allan H-Conney
Laboratory for Cancer Research, Department of Chemical Biology, College of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, NJ 08855-0789, 'Department of Environmental Medicine, New York University Medical Center, New York, NY 10016-6451 and institute of Cancer Research, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA
Topical application of caffeic acid phenethyl ester (CAPE), a constituent of the propolis of honeybee hives, to the backs of CD-I mice previously initiated with 7,12- dimethylbenz[<i]anthracene (DMBA) inhibited 12-0-tetradecanoylphorbol-13-acetate (TPA)-induced tumor promotion and the formation of 5-hydroxymethyl-2'-deoxyuridine (HMdU) in epidermal DNA. Topical application of 5 nmol TPA twice weekly for 20 weeks to mice previously initiated with 200 nmol of DMBA resulted in 18.8 skin papillomas per mouse. Topical application of 1, 10, 100 or 3000 nmol of CAPE together with 5 nmol of TPA twice a week for 20 weeks inhibited the number of skin papillomas per mouse by 24, 30, 45 or 70%, respectively, and tumor size per mouse was decreased by 42, 66, 53 or 74%, respectively.Topical application of 5 nmol of TPA twice weekly for 20 weeks to mice previously initiated with DMBA produced an average of 12.6 HMdU residues per 104 normal bases in epidermal DNA. Topical application of 1, 10, 100 or 3000 nmol of CAPE with 5 nmol of TPA twice weekly for 20 weeks to DMBA-initiated mice decreased the level of HMdU in epidermal DNA by 40-93%. The in vitro addition of 1.25, 2.5, 5, 10 or 20 jiM CAPE to cultured HeLa cells inhibited the synthesis of DNA by 32, 44, 66, 79 or 95%, respectively, the synthesis of RNA was inhibited by 39, 43,58, 64 or 75%, respectively, and the synthesis of protein was inhibited by 29, 30, 37, 32 or 47%, respectively. The results indicate a potent inhibitory effect of CAPE on TPAinduced tumor promotion and TPA-induced formation of HMdU in DNA of mouse skin as well as an inhibitory effect of CAPE on the synthesis of DNA, RNA and protein in cultured HeLa cells.