ß-Escin Effectively Modulates HUVECS Proliferation and Tube Formation.

In the present study we evaluated the anti-angiogenic activities of ß-escin (the major active compound of Aesculus hippocastanum L. seeds). Human umbilical-vein endothelial cells (HUVECs) were used as an in vitro model for studying the molecular mechanism underlying the anti-angiogenic effect of ß-escin. We investigated the in vitro effects on proliferation, migration, and tube formation of HUVECs and in vivo anti-angiogenic activity was evaluated in a chick chorioallantoic membrane (CAM) angiogenesis assay. Moreover, the effect on gene expressions was determined by the RT2 ProfilerTM human angiogenesis PCR Array. It was found that ß-escin exerts inhibitory effect on the basic fibroblast growth factor (bFGF)-induced proliferation, migration and tube formation, as well as CAM angiogenesis in vivo. The inhibition of critical steps of angiogenic process observed with ß-escin could be partially explained by suppression of Akt activation in response to bFGF. Moreover, the anti-angiogenic effects of ß-escin could also be mediated via inhibition of EFNB2 and FGF-1 gene expressions in endothelial cells. In conclusion, ß-escin affects endothelial cells as a negative mediator of angiogenesis in vitro and in vivo and may therefore be considered as a promising candidate for further research elucidating its underlying mechanism of action.

Genistein Improves Skin Flap Viability in Rats: A Preliminary In Vivo and In Vitro Investigation.

Selective estrogen receptor modulators (SERMs) have been developed to achieve beneficial effects of estrogens while minimizing their side effects. In this context, we decided to evaluate the protective effect of genistein, a natural SERM, on skin flap viability in rats and in a series of in vitro experiments on endothelial cells (migration, proliferation, antioxidant properties, and gene expression profiling following genistein treatment). Our results showed that administration of genistein increased skin flap viability, but importantly, the difference is only significant when treatment is started 3 days prior the flap surgery. Based on our in vitro experiments, it may be hypothesized that the underlying mechanism may rather by mediated by increasing SOD activity and Bcl-2 expression. The gene expression profiling further revealed 9 up-regulated genes (angiogenesis/inflammation promoting: CTGF, CXCL5, IL-6, ITGB3, MMP-14, and VEGF-A; angiogenesis inhibiting: COL18A1, TIMP-2, and TIMP-3). In conclusion, we observed a protective effect of genistein on skin flap viability which could be potentially applied in plastic surgery to women undergoing a reconstructive and/or plastic intervention. Nevertheless, further research is needed to explain the exact underlying mechanism and to find the optimal treatment protocol.

Antiangiogenic Effect of Flavonoids and Chalcones: An Update.

Chalcones are precursors of flavonoid biosynthesis in plants. Both flavonoids and chalcones are intensively investigated because of a large spectrum of their biological activities. Among others, anticancer and antiangiogenic effects account for the research interest of these substances. Because of an essential role in cancer growth and metastasis, angiogenesis is considered to be a promising target for cancer treatment. Currently used antiangiogenic agents are either synthetic compounds or monoclonal antibodies. However, there are some limitations of their use including toxicity and high price, making the search for new antiangiogenic compounds very attractive. Nowadays it is well known that several natural compounds may modulate basic steps in angiogenesis. A lot of studies, also from our lab, showed that phytochemicals, including polyphenols, are potent modulators of angiogenesis. This review paper is focused on the antiangiogenic effect of flavonoids and chalcones and discusses possible underlying cellular and molecular mechanisms.

How Signaling Molecules Regulate Tumor Microenvironment: Parallels to Wound Repair.

It is now suggested that the inhibition of biological programs that are associated with the tumor microenvironment may be critical to the diagnostics, prevention and treatment of cancer. On the other hand, a suitable wound microenvironment would accelerate tissue repair and prevent extensive scar formation. In the present review paper, we define key signaling molecules (growth factors, cytokines, chemokines, and galectins) involved in the formation of the tumor microenvironment that decrease overall survival and increase drug resistance in cancer suffering patients. Additional attention will also be given to show whether targeted modulation of these regulators promote tissue regeneration and wound management. Whole-genome transcriptome profiling, in vitro and animal experiments revealed that interleukin 6, interleukin 8, chemokine (C-X-C motif) ligand 1, galectin-1, and selected proteins of the extracellular matrix (e.g., fibronectin) do have similar regulation during wound healing and tumor growth. Published data demonstrate remarkable similarities between the tumor and wound microenvironments. Therefore, tailor made manipulation of cancer stroma can have important therapeutic consequences. Moreover, better understanding of cancer cell-stroma interaction can help to improve wound healing by supporting granulation tissue formation and process of reepithelization of extensive and chronic wounds as well as prevention of hypertrophic scars and formation of keloids.

Soy and breast cancer: focus on angiogenesis.

Epidemiological studies have revealed that high consumption of soy products is associated with low incidences of hormone-dependent cancers, including breast and prostate cancer. Soybeans contain large amounts of isoflavones, such as the genistein and daidzain. Previously, it has been demonstrated that genistein, one of the predominant soy isoflavones, can inhibit several steps involved in carcinogenesis. It is suggested that genistein possesses pleiotropic molecular mechanisms of action including inhibition of tyrosine kinases, DNA topoisomerase II, 5α-reductase, galectin-induced G2/M arrest, protein histidine kinase, and cyclin-dependent kinases, modulation of different signaling pathways associated with the growth of cancer cells (e.g., NF-κB, Akt, MAPK), etc. Moreover, genistein is also a potent inhibitor of angiogenesis. Uncontrolled angiogenesis is considered as a key step in cancer growth, invasion, and metastasis. Genistein was found to inhibit angiogenesis through regulation of multiple pathways, such as regulation of VEGF, MMPs, EGFR expressions and NF-κB, PI3-K/Akt, ERK1/2 signaling pathways, thereby causing strong antiangiogenic effects. This review focuses on the antiangiogenic properties of soy isoflavonoids and examines their possible underlying mechanisms.

Agrimonia eupatoria L. and Cynara cardunculus L. Water Infusions: Phenolic Profile and Comparison of Antioxidant Activities.

Reactive oxygen species (ROS) are highly considered in the ethiopathogenesis of different pathological conditions because they may cause significant damage to cells and tissues. In this paper, we focused on potential antioxidant properties of two medical plants such as the Agrimonia eupatoria L. and Cynara cardunculus L. Both plants have previously been studied for their pharmacological activities, especially as hepatoprotective and hypoglycemic activities. It has been suggested, that their effects are related to the antioxidant properties of polyphenols, which are dominant compounds of the plants' extracts. In the present study HPLC-MS analysis of water infusion was performed allowing the identification of several phenolic constituents. Furthermore, antioxidant effects of the two extracts were compared showing higher effects for agrimony extract compared to artichoke. Thus, agrimony was selected for the in vivo study using the skin flap viability model. In conclusion, our results provide evidence that the A. eupatoria extract may be a valuable source of polyphenols to be studied for the future development of supplements useful in the prevention of diseases linked to oxidative stress.

[Basic biological roles of galectins in tissue repair and tumor growth]. / Základné biologické úlohy galektínov v reparácii tkanív a raste nádorov.

Galectins are representatives of endogenous lectins - molecules specifically recognizing distinct sugar motifs. They play an important role in the processes of cell proliferation, differentiation, migration and extracellular matrix formation. Furthermore, galectins are able to transfer cellular signals and to participate in intercellular interaction. It has been proven that galectins play an important role in the formation of tumor and/or wound healing microenvironment. This review contains an overview of experimental and clinical studies dealing with biological roles of galectins in tissue repair and in its parallel - the tumor growth.

The synthesis and anticancer activity of analogs of the indole phytoalexins brassinin, 1-methoxyspirobrassinol methyl ether and cyclobrassinin.

An effective synthesis of analogs of the indole phytoalexin cyclobrassinin with NR1R2 group instead of SCH3 was developed starting from indole-3-carboxaldehyde. The target compounds were prepared by spirocyclization of 1-Boc-thioureas with the formation of isolable spiroindoline intermediates, followed by the trifluoroacetic acid-induced cascade reaction consisting of methanol elimination, deprotection and rearrangement of the iminium ion. The structures of novel products were elucided by the (1)H and (13)C NMR spectroscopy, including HMBC, HSQC, COSY, NOESY and DEPT measurements. Several newly synthesized compounds demonstrated significant antiproliferative/cytotoxic activity against human leukemia and solid tumor cell lines, as well as remarkable selectivity of these effects against cancer cells relative to the non-malignant HUVEC cells.

Cyclic chalcone analogue KRP6 as a potent modulator of cell proliferation: an in vitro study in HUVECs.

In the present investigation a novel series of chalcone analogues were synthesized and evaluated for their anti-proliferative activity in human umbilical vein endothelial cells (HUVECs). Among 14 tested compounds, chalcone analogue (E)-3-(2'-methoxybenzylidene)-4-chromanone (KRP6) exhibited the most potent activity with IC50 19 µM. Moreover, HUVECs exhibited divergent, even opposing concentration-dependent responses to KRP6. This compound was the most potent inhibitor of cell proliferation and extracellular matrix formation (fibronectin and type IV collagen) at higher concentrations (20-50 µM). In contrast, KRP6 stimulated the compensatory increase in proliferative activity including extracellular matrix formation at low concentrations (1, 10 µM). KRP6 concentration-dependently modulated phosphorylation of Akt and mitogen-activated protein kinases such as extracellular signal-regulated kinase-1/-2 and p38 kinase, suggesting that these pathways play a role in the effect mediated by this compound. In addition, we found a selective effect on activated endothelial cells, in particular with resting endothelial cells. In conclusion, KRP6 is a potent modulator of selected steps of the angiogenic process in vitro. Accordingly, further in vivo research should be performed to facilitate its use in clinical practice.

Antiproliferative and antiangiogenic properties of horse chestnut extract.

This study was designed to examine the in vitro antiproliferative effect of the horse chestnut extract (HCE) on cancer cell lines. Furthermore, we have investigated the in vitro effect of HCE on some angiogenic events by using human umbilical vein endothelial cells. The cell proliferation was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and anchorage-independent growth by colony-forming assay. To understand the growth inhibitory effects, carcinoma cell lines (Jurkat, CEM, HeLa, and MCF-7) were treated with various concentrations of HCE. Incubation of Jurkat, CEM, HeLa, and MCF-7 cancer cells with HCE at 125 µg/mL for 72 h caused 93.7%, 32.3%, 20.4% and 40.4% reduction in cell survival. Colony-forming assay also confirmed growth-inhibitory effects of the compound studied. In HeLa HCE-treated cells, we found a significant increase in cells having sub-G(0) /G(1) DNA content which is considered to be a marker of apoptotic cell death. Apoptosis was also further confirmed by DNA fragmentation analysis.Furthermore, HCE inhibited migration of human umbilical vein endothelial cells as well as decreased secretion of matrix metalloproteinase and vascular endothelial growth factor.In conclusion, the present study has assessed the in vitro antiproliferative/antiangiogenic potential of HCE. These results generate a rationale for in vivo efficacy studies with horse chestnut in preclinical cancer models.

ER-α agonist induces conversion of fibroblasts into myofibroblasts, while ER-ß agonist increases ECM production and wound tensile strength of healing skin wounds in ovariectomised rats.

Oestrogen deprivation is one of the major factors responsible for many age-related processes, including poor wound healing in women. Previously, it has been shown that oestrogens have a modulatory effect in different wound-healing models. Therefore, in this study, the effect of selective oestrogen receptor (ER) agonists (PPT - ER-α agonist, DPN - ER-ß agonist) on excisional and incisional wound-healing models was compared in ovariectomised rats in vivo as well as on human dermal fibroblasts (HDF) and human umbilical endothelial cells (HUVEC) in vitro. In the in vivo study, 4 months after either ovariectomy or sham ovariectomy, Sprague-Dawley rats were randomly divided into four groups and subjected to two incisional and excisional wounds: (i) control - sham operated, vehicle-treated; (ii) ovariectomised, vehicle-treated; (iii) ovariectomised, PPT treated; (iv) ovariectomised, DPN treated. In the in vitro study, HDFs and HUVECs were used. After treatment with ER agonists, cells were processed for immunocytochemistry and gelatin zymography. Our study shows that stimulation of ER-α leads to the differentiation of fibroblasts into myofibroblasts both in vivo and in vitro. On the other hand, the formation of extracellular matrix was more prominent, and wound tensile strength (TS) was increased when ER-ß was stimulated. In contrast, stimulation of ER-α led to a more prominent increase in the expression of MMP-2 and decrease in wound TS. New information is presented in this investigation concerning oestrogen replacement therapy (ERT) in different wound-healing models. This study demonstrates that the ERT should be both wound and receptor-type specific.

Rate of oxidative modification of cytochrome c by hydrogen peroxide is modulated by Hofmeister anions.

Cytochrome c (cyt c) and other heme proteins are oxidatively modified in the presence of hydrogen peroxide in a concentration- and time-dependent manner. Cyt c modification has been monitored by several spectral probes by absorption spectroscopy (at wavelengths 410 nm, 530 nm), and circular dichroism (222, 268, 288 and 417 nm). Kinetics monitored with these spectral probes indicates that the oxidative modification of cyt c: i) proceeds in the order: heme --> aromatic amino acids --> secondary structure, and ii) the rate of the oxidative modification is proportional to the protein flexibility. The flexibility of cyt c was modulated by anions of Hofmeister series (sulfate, chloride, perchlorate) (Varhac et al. 2009). A minimalist scheme of the interaction of cyt c with hydrogen peroxide can be described by two steps: 1) interaction of hydrogen peroxide with heme iron forming the postulated ferryl intermediate, 2a) oxidation of another molecule of hydrogen peroxide and 2b) parallel oxidation of close amino acid residue(s) and/or heme. The catalase activity of cyt c is independent from the presence of Hofmeister anions, which indicates that both steps (1 and 2a) in the catalase reaction are independent from the flexibility of the heme region of the protein matrix. On the other hand, the flexibility of the polypeptide chain of the protein modulates the rate of parallel oxidative modification of the heme and amino acid residues.

Effect of selected flavones on cancer and endothelial cells.

In our study we used quercetin (3,3 ,4 ,5,7-pentahydroxyflavone) as the reference standard to compare antiproliferative and antiangiogenic effects of chrysin (5,7-dihydroxyflavone) and 3-hydroxyflavone. Our data indicates that chrysin and 3-hydroxyflavone showed significantly higher cytotoxic effect than reference standard quercetin. These tested agents significantly decreased cell survival with the efficacy of 65-85% at the concentration 100 micromol/l for HUVEC, lung carcinoma and leukemic cells being the most sensitive. Cell cycle analysis indicates that quercetin and 3-hydroxyflavone might affect the cell cycle of Jurkat cells by a similar or the same mechanism of action which lead to G2/M arrest as well as to an increase in sub-G0/G1 fraction. Treatment of Jurkat cells with chrysin resulted only increase in the fraction of cells with sub-G0/G1 DNA content, which is considered to be a marker of apoptotic cell death. Apoptosis was confirmed by DNA fragmentation and by staining with annexin V. All three tested flavones inhibited endothelial cell migration after 24 h of incubation at a concentration 100 micromol/l. At a lower concentration (10 micromol/l) only quercetin significantly inhibited migration of endothelial cells. Furthermore, in our experiments decreased secretion of matrix metalloproteinases (MMP-2 and MMP-9) was observed after a 72 h treatment with quercetin. No decrease in secretion of MMP-2 and MMP-9 was seen after chrysin and 3-hydroxyflavone treatment. On the other hand, our results showed that none of three flavonoids blocked microcapillary tube formation. Further studies are necessary to investigate the mechanism of action and to find out the relationship between the structure, character and position of substituents of natural substances and their biological activities.

Galectin-8 Favors VEGF-Induced Angiogenesis: In Vitro Study in Human Umbilical Vein Endothelial Cells and In Vivo Study in Chick Chorioallantoic Membrane.

BACKGROUND/AIM: Although it has been accepted that the tandem repeat galectin-8 (Gal-8) is linked to angiogenesis, the underlying mechanisms in endothelial cells has remained poorly understood. In this study we aimed to investigate the effect of Gal-8 on selected biological processes linked to angiogenesis in in vitro and in vivo models. MATERIALS AND METHODS: In detail, we assessed how exogenously added human recombinant Gal-8 (with or without vascular endothelial growth factor - VEGF) affects selected steps involved in vessel formation in human umbilical vein endothelial cells (HUVECs) as well as using the chick chorioallantoic membrane (CAM) assay. Gene expression profiling of HUVECs was performed to extend the scope of our investigation. RESULTS: Our findings demonstrate that Gal-8 in combination with VEGF enhanced cell proliferation and migration, two cellular events linked to angiogenesis. However, Gal-8 alone did not exhibit any significant effects on cell proliferation or on cell migration. The molecular analysis revealed that Gal-8 in the presence of VEGF influenced cytokine-cytokine receptor interactions, HIF-1 and PI3K/AKT signaling pathways. Gal-8 alone also targeted cytokine-cytokine receptor interactions, but with a different expression profile as well as a modulated focal adhesion and TNF signaling. CONCLUSION: Gal-8 promotes a pro-angiogenic phenotype possibly in a synergistic manner with VEGF.

Photodynamic effect of hypericin in primary cultures of human umbilical endothelial cells and glioma cell lines.

Hypericin is the most powerful naturally occurring photosensitizer and as such there is renaissant interest in the potentials of this compound for anticancer photodynamic therapy (PDT). The purpose of this study was to investigate the hypericin-mediated photodynamic therapy effects on normal human umbilical endothelial cells (HUVECs) in comparison with cancer human glioma cell lines U-87 MG and U-373 MG, in in vitro conditions. The data suggest that endothelial cells as well as glioma cell lines are sensitive only to photoactivated hypericin. The inhibitory effects of photoactivated hypericin did not differ in endothelial compared with tumor cells in cytotoxicity MTT and DNA fragmentation assays. However, an important difference in sensitivity was found between the above mentioned cell types in migration and metalloproteinases inhibition assays performed as cell function tests. The findings in both function tests were supported by the high sensitivity of endothelial cells in an additional angiogenesis test of tubular formation in vitro.

Inhibition of heat shock protein (Hsp) 90 potentiates the antiproliferative and pro-apoptotic effects of 2-(4'fluoro-phenylamino)-4H-1,3-thiazine[6,5-b]indole in A2780cis cells.

Ovarian carcinoma is initially sensitive to platinum-based therapy, but become resistant over time. The study of cancer sensitizing substance is therefore the major challenge for a number of scientific groups. Our experiments were carried out on human ovarian adenocarcinoma A2780cis cells resistant to cisplatin and their response to 2-(4'fluoro-phenylamino)-4H-1,3-thiazine[6,5-b]indole (thiazine[6,5-b]indole) and/or heat shock protein (Hsp) 90 inhibitor 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) using proliferation assay, cell cycle analysis and monitoring of apoptosis were examined. A2780cis cells revealed the same fold of resistance to Hsp90 inhibitor 17-DMAG as it is declared for cisplatin (18 times), but only 3.2 times for thiazine[6,5-b]indole. Our results showed that the combination of thiazine[6,5-b]indole and 17-DMAG significantly reduced proliferation of A2780cis cells and led to their accumulation in G2/M phase of the cell cycle. Moreover, both thiazine[6,5-b]indole as well as 17-DMAG increased the number of annexin V positive A2780cis cells in time dependent manner. Interestingly, thiazine[6,5-b]indole treatment significantly activated also caspase-3 compared to untreated or 17-DMAG-treated cells and reduced mitochondrial membrane potential (MMP) of A2780cis cells with more significant decline after combined treatment. In this regard, the incubation of A2780cis cells with thiazine[6,5-b]indole induced PARP protein cleavage as well as an increased level of Bad protein with more pronounced changes after combined treatment. Importantly, Hsp70 protein was not upregulated in A2780cis cells neither by individual treatment nor by mutual combination. Our results signify antiproliferative and pro-apoptotic effects of novel thiazine[6,5-b]indole potentiated by Hsp90 inhibitor 17-DMAG in ovarian adenocarcinoma cells resistant to cisplatin and therefore represents new strategy in cancer treatment.

Angiomodulators in cancer therapy: New perspectives.

The formation of new blood vessels plays a crucial for the development and progression of pathophysiological changes associated with a variety of disorders, including carcinogenesis. Angiogenesis inhibitors (anti-angiogenics) are an important part of treatment for some types of cancer. Some natural products isolated from marine invertebrates have revealed antiangiogenic activities, which are diverse in structure and mechanisms of action. Many preclinical studies have generated new models for further modification and optimization of anti-angiogenic substances, and new information for mechanistic studies and new anti-cancer drug candidates for clinical practice. Moreover, in the last decade it has become apparent that galectins are important regulators of tumor angiogenesis, as well as microRNA. MicroRNAs have been validated to modulate endothelial cell migration or endothelial tube organization. In the present review we summarize the current knowledge regarding the role of marine-derived natural products, galectins and microRNAs in tumor angiogenesis.

Pharmacological activation of estrogen receptors-α and -ß differentially modulates keratinocyte differentiation with functional impact on wound healing.

Estrogen deprivation is considered responsible for many age-related processes, including poor wound healing. Guided by previous observations that estradiol accelerates re­epithelialization through estrogen receptor (ER)­ß, in the present study, we examined whether selective ER agonists [4,4',4''-(4-propyl [1H] pyrazole-1,3,5-triyl)­trisphenol (PPT), ER­α agonist; 2,3-bis(4-hydroxyphenyl)-propionitrile (DPN), ER­ß agonist] affect the expression of basic proliferation and differentiation markers (Ki­67, keratin­10, ­14 and ­19, galectin­1 and Sox­2) of keratinocytes using HaCaT cells. In parallel, ovariectomized rats were treated daily with an ER modulator, and wound tissue was removed 21 days after wounding and routinely processed for basic histological analysis. Our results revealed that the HaCaT keratinocytes expressed both ER­α and ­ß, and thus are well-suited for studying the effects of ER agonists on epidermal regeneration. The activation of ER­α produced a protein expression pattern similar to that observed in the control culture, with a moderate expression of Ki­67 being observed. However, the activation of ER­ß led to an increase in cell proliferation and keratin­19 expression, as well as a decrease in galectin­1 expression. Fittingly, in rat wounds treated with the ER­ß agonist (DPN), epidermal regeneration was accelerated. In the present study, we provide information on the mechanisms through which estrogens affect the expression patterns of selected markers, thus modulating keratinocyte proliferation and differentiation; in addition, we demonstrate that the pharmacological activation of ER-α and -ß has a direct impact on wound healing.

Extracellular matrix of galectin-1-exposed dermal and tumor-associated fibroblasts favors growth of human umbilical vein endothelial cells in vitro: a short report.

BACKGROUND/AIM: Stromal cells in the tumor microenvironment are primarily considered as sources of promalignant factors. The objective of our study was to define the effect of extracellular matrix (ECM) produced by normal dermal or cancer-associated fibroblasts exposed to adhesion/growth-regulatory lectin galectin-1 on human umbilical vein endothelial cells (HUVECs). MATERIALS AND METHODS: Fibroblasts were cultured for 10 days with lectin, followed by removing cellular constituents after an osmotic shock. Freshly-isolated HUVECs were placed on the ECM. In parallel, HUVECs were seeded on untreated and gelatin-coated surfaces as controls. A positive control for growth of HUVECs culture using medium supplemented with vascular endothelial growth factor completed the test panel. Cells were kept in contact to the substratum for two days and then processed for immunocytochemistry. RESULTS: HUVECs seeded on fibroblast-generated ECM presented a comparatively high degree of proliferation. Furthermore, contact to substratum produced by tumor-associated fibroblasts led to generation of a meshwork especially rich in fibronectin. CONCLUSION: Galectin-1 is apparently capable to trigger ECM production favorable for growth of HUVECs, prompting further work on characterizing structural features of the ECM and in situ correlation of lectin presence, ECM constitution and neoangiogenesis.

In vitro toxicity of camalexin derivatives in human cancer and non-cancer cells.

The aim of the study was to investigate the cytotoxic activity of camalexin and its five synthetic derivatives in cancer and non-cancer cells. In cancer cells the benzocamalexin (BC) displayed the most potent activity with an IC value of 23.3-30.1 µmol/L. On the other hand, minimal toxicity (IC>100.0 µmol/L) in non-cancer cells was observed. Based on these results, BC was selected for further studies. Flow cytometric analysis revealed a BC-induced arrest of the cell cycle in the G2 phase associated with downregulation of α-tubulin, α1-tubulin, ß5-tubulin expression. These findings suggest that the inhibitory effect of BC is mediated via inhibition of microtubule formation. Moreover, BC downregulated the expression of microtubule-related protein indicating the effect of this compound on microtubule assembly. After treatment with BC increase of the sub-G DNA content fraction was noted which is considered to be a marker of apoptotic cell death. Apoptosis was also confirmed by DNA fragmentation assay. Moreover, quantitative real-time PCR showed that BC downregulated the expression of antiapoptotic genes Bcl-2 and Bcl-xL and upregulated the expression of proapoptotic Bax. Taken together, our study suggests that the blockade of cell cycle progression and initiation of apoptosis may play an important role in the antiproliferative activity of BC in human cancer cells.