RESUMEN
A combination of a radiotherapeutic regimen with telomerase inhibition is valuable when tumor cells are to be sensitized to radiation. Here, we describe cell clones unresponsive to radiosensitization after telomere shortening. After extensive division of individual transformed clones of mTERC-/- cells, about 22% of clones were unresponsive to radiosensitization even though telomerase action was inhibited. The telomere lengths of unsensitized mTERC-/- clones were reduced, as were those of most sensitized clones. However, the unsensitized clones did not exhibit chromosomal end-to-end fusion to the extent noted in all sensitized clones. Thus, a defense mechanism preventing telomere erosion is operative even when telomeres become shorter under conditions of telomerase deficiency, and results in unresponsiveness to the radiosensitization generally mediated by telomere shortening.
Asunto(s)
Tolerancia a Radiación , Telomerasa/antagonistas & inhibidores , Telómero/metabolismo , Animales , Células Clonales , Fibroblastos/enzimología , Fibroblastos/efectos de la radiación , Ratones , Ratones Mutantes , Telomerasa/genéticaRESUMEN
A galactose- and N-acetyl-D-galactosamine-specific lectin (Viscum album L. var. coloratum agglutinin, VCA), which is known for its anti-cancer activity, was isolated from Korean mistletoe. In this study, IEC-6 rat intestinal epithelial cells and IM-9 human B-cells were cultured to determine the effect of VCA on cytokine and immunoglobulin (Ig) secretion. In lipopolysaccharide (LPS)-stimulated IEC-6 cells, VCA significantly shifted the interleukin (IL)-2, IL-5, IL-6, and tumor necrosis factor-alpha (TNF-alpha) secretion toward a more immunostimulatory response. Since intestinal epithelial cell-derived secretions may be capable of affecting local B cell Ig production in a variety of ways, we mimicked this condition by deriving a 2-day culture supernatant from IEC-6 cell line which was treated VCA in the presence or absence of LPS, and adding these supernatants to cultures of IM-9 human B cells. As a result, IgA secretion was significantly enhanced at in the presence of VCA at 10(-8)-10(-4) microg/mL. This study suggests that cytokines derived from IEC by VCA may create an environment which may contribute to the enhancement of IgA secretion seen in mucosal tissues. Overall, the induction of cytokines in intestinal epithelial cells, and IgA in B cells by Korean mistletoe lectin could indicate an enhanced immunosurveillance to prevent intestinal infections or other intestinal pathologies.
Asunto(s)
Linfocitos B/efectos de los fármacos , Citocinas/metabolismo , Inmunoglobulina A/metabolismo , Factores Inmunológicos/farmacología , Mucosa Intestinal/efectos de los fármacos , Lectinas de Plantas/farmacología , Viscum album , Animales , Linfocitos B/inmunología , Línea Celular , Supervivencia Celular/efectos de los fármacos , Medios de Cultivo Condicionados/metabolismo , Relación Dosis-Respuesta a Droga , Humanos , Inmunidad Mucosa/efectos de los fármacos , Factores Inmunológicos/aislamiento & purificación , Interleucinas/metabolismo , Mucosa Intestinal/inmunología , Lipopolisacáridos/farmacología , Lectinas de Plantas/aislamiento & purificación , Ratas , Factor de Necrosis Tumoral alfa/metabolismo , Viscum album/químicaRESUMEN
Purified mistletoe lectins are known to have cytotoxic and stimulating activities in the immune system. Mistletoe extract has been given subcutaneously because of its unstablity and poor absorption in the Gastrointestinal (GI) tract. A hallmark of M cells is their capacity to internalize material from the lumen and to transfer it efficiently to the underlying lymphoid cells. Although lectins are the prime candidates for oral vaccine delivery, the mechanisms whereby lectins are taken up, transported by M cells, and affect underlying immune cells remain poorly understood. In this study, uptake mechanism of Korean mistletoe lectin (Viscum album L. var. coloratum aggulutinin, VCA) across the human FAE (follicle associated epithelium) was investigated. An inverted FAE model of co-culture was obtained by a co-culture system of Caco-2 cells and human Raji B lymphocytes, and VCA transport across the in vitro model of human FAE was investigated. There was a greater transport of VCA across FAE monolayer cells than that of Caco-2 monolayer cells. These observations will be useful to assess the transport of other orally administered material in the GI tract.
Asunto(s)
Mucosa Intestinal/metabolismo , Lectinas/metabolismo , Muérdago/química , Linfocitos B/efectos de los fármacos , Linfocitos B/metabolismo , Western Blotting , Células CACO-2 , Línea Celular , Técnicas de Cocultivo , Ensayos de Selección de Medicamentos Antitumorales , Ensayo de Inmunoadsorción Enzimática , Galectinas/biosíntesis , Galectinas/genética , Humanos , Mucosa Intestinal/citología , Sales de Tetrazolio , TiazolesRESUMEN
AIM: To investigate the anti-cancer mechanisms of Korean mistletoe lectin (Viscum album coloratum agglutinin, VCA) using a human colon cancer cell line (COLO). METHODS: Cytotoxic effects of VCA on COLO cells were determined by 3- (4, 5-dimethylthiazol-2-yl) -2, 5-diphenyltetrazolium bromide (MTT) assay in vitro and tumor-killing effects in vivo. To study the mechanisms involved, the expression of various pro-caspases, anti-apoptotic proteins, and death receptors was determined by western blot. To determine which death receptor is involved in VCA-induced apoptosis of COLO cells, cytotoxicity was examined by MTT assay after treatment with agonists or antagonists of death receptors. RESULTS: VCA killed COLO cells in a time- and dose-dependent manner and induced complete regression of tumors in nude mice transplanted with COLO cells. Treatment of COLO cells with VCA activated caspase-2, -3, -8, and -9 and decreased expression of anti-apoptotic molecules including receptor interacting protein, nuclear factor-kappaB, X-linked inhibitor of apoptosis protein, and Akt/protein kinase B. We then examined the involvement of death receptors in VCA-induced apoptosis. Only tumor necrosis factor receptor 1, among the death receptors examined, was involved in apoptosis of COLO cells, evidenced by inhibition of VCA-induced apoptosis and decreased activation of caspases, particularly caspase-8, by tumor necrosis factor receptor 1 antagonizing antibody. CONCLUSION: VCA-induced apoptotic COLO cell death is due to the activation of caspases and inhibition of anti-apoptotic proteins, in part through the tumor necrosis factor receptor 1 signaling pathway.
Asunto(s)
Apoptosis/efectos de los fármacos , Neoplasias del Colon/patología , Neoplasias del Colon/fisiopatología , Preparaciones de Plantas/farmacología , Proteínas de Plantas/farmacología , Toxinas Biológicas/farmacología , Apoptosis/fisiología , Proteínas Reguladoras de la Apoptosis/genética , Proteínas Reguladoras de la Apoptosis/fisiología , Caspasas/genética , Caspasas/fisiología , Línea Celular Tumoral , Neoplasias del Colon/genética , Relación Dosis-Respuesta a Droga , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/fisiología , Proteínas Proto-Oncogénicas c-akt/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/fisiología , Proteínas Inactivadoras de Ribosomas Tipo 2 , Transducción de Señal/fisiología , Factor 1 Asociado a Receptor de TNF/genética , Factor 1 Asociado a Receptor de TNF/fisiología , Factores de TiempoRESUMEN
The anti-cancer activity of mistletoe has been ascribed to a combination of cytotoxic and immunological effects. We previously showed that Korean mistletoe lectin (Viscum album L. var. coloratum agglutinin, VCA) can stimulate IFN-gamma production and modulate proliferation in murine splenocytes. In this study, we investigated the effects of VCA on human peripheral blood mononuclear cells (hPBMC) and T-lymphocytes. The addition of VCA resulted in a significant inhibition of proliferation at higher concentrations (at 2-8 ng/mL, 1-8 ng/mL in hPBMC and T-lymphocytes, respectively) but an induction at lower concentrations (at 4-16 pg/mL, 4-32 pg/mL in hPBMC and T-lymphocytes, respectively). Further studies were carried out to determine if the pro-proliferative or anti-proliferative activity exhibited by VCA was correlated with apoptosis and cytokine secretion. As a result, the apoptotic cell number increased to 26% after 48 h of VCA treatment (10 ng/mL) in the presence of anti-CD3/CD28 antibodies. On the other hand, without anti-CD3/CD28 antibody stimulants, VCA did not arrest cell cycle. In addition, it was shown that VCA could induce IL-2 secretion was dose-dependently increased by VCA in stimulated (anti-CD3/CD28 antibodies) (at 0.25-2 ng/mL) and non-stimulated (at 3-25 pg/mL) human T-lymphocytes. Also, at low and non-toxic concentrations of VCA, the RT-PCR result confirmed the release of pro-inflammatory cytokines such as IL-1alpha, IL-1beta, IL-6, IL-8, and IFN-gamma. These data may suggest new perspective to modulate the balance between cell growth, cytokine production and programmed cell death therapeutically.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Proliferación Celular/efectos de los fármacos , Citocinas/metabolismo , Leucocitos Mononucleares/efectos de los fármacos , Activación de Linfocitos/efectos de los fármacos , Lectinas de Plantas/farmacología , Linfocitos T/efectos de los fármacos , Viscum album , Anticuerpos , Antineoplásicos Fitogénicos/química , Antineoplásicos Fitogénicos/aislamiento & purificación , Apoptosis/efectos de los fármacos , Antígenos CD28/inmunología , Complejo CD3/inmunología , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Citocinas/genética , Relación Dosis-Respuesta a Droga , Hemaglutinación/efectos de los fármacos , Pruebas de Hemaglutinación , Humanos , Interleucina-2/metabolismo , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Leucocitos Mononucleares/patología , Peso Molecular , Lectinas de Plantas/química , Lectinas de Plantas/aislamiento & purificación , Preparaciones de Plantas/farmacología , ARN Mensajero/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/patología , Factores de Tiempo , Viscum album/químicaRESUMEN
Loss of p53 tumor suppressor facilitates acquisition of telomerase activity. In fact, both p53 inactivation and telomerase activation are frequently found in human cancers. p53 inactivation, however, eliminates or attenuates the biological responses to telomerase inhibition and the eventual telomere erosion. We show that telomere erosion can increase the susceptibility to radiation, irrespective of p53 status. Both telomerase inhibition and critically shortened telomere with significant change of chromosomal end-to-end fusion were essential for the enhancement of radiosensitivity. The enhancement was correlated with greater formation of multinucleated cells. p53 inactivation did not eliminate the observed generation of chromosomal fusion and multinucleation, and the resulting increased susceptibility to radiation, as opposed to the previously proved role of p53 in mediating cellular responses to telomere dysfunction. The present findings suggest the importance of chromosomal end fusion in modulating radiosensitivity rather than p53 DNA damage signaling. Thus, the suggested anticancer radiotherapeutic strategy combined with telomerase inhibition could clinically be applicable to cancers, irrespective of p53 status.
Asunto(s)
Cromosomas/genética , Tolerancia a Radiación , Recombinación Genética , Telomerasa/antagonistas & inhibidores , Telómero/metabolismo , Animales , Muerte Celular , Inestabilidad Genómica , Ratones , Mitosis , Tolerancia a Radiación/genética , Telomerasa/genética , Telomerasa/metabolismo , Telómero/genética , Proteína p53 Supresora de Tumor/antagonistas & inhibidores , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
It is well documented that an extract of European mistletoe has a variety of biological effects, such as the stimulation of cytokine production from immune cells, and additional immunoadjuvant activities. While the European mistletoe has been studied intensively, we know less about Korean mistletoe as a therapeutic plant, especially as a possible immunomodulating drug. This study will investigated the effects of Korean mistletoe lectin (Viscum album L. var. coloratum agglutinin, VCA) on murine splenocytes to investigate whether VCA acts as an immunomodulator, which could lead to improved immune responses in these cells. The results showed that VCA inhibited cell proliferation at higher concentrations (at 1-8 ng/ml) and enhanced cell proliferation at lower concentrations (at 4-32 pg/ml). Further studies were carried out to determine if the proproliferative or anti-proliferative activity exhibited by VCA was correlated with cytokine secretion. Consequently, interferon (IFN)-gamma secretion was decreased in concanavalin A (ConA)-stimulated murine splenocytes by VCA (4-64 ng/ml), but there was no change in IL-4 levels. This suggests that VCA has the ability to modulate murine splenocyte proliferation and can possibly act on the balance of Th1/Th2 cellular immune responses.
Asunto(s)
Proliferación Celular/efectos de los fármacos , Citocinas/metabolismo , Expresión Génica/efectos de los fármacos , Lectinas de Plantas/farmacología , Esplenosis/patología , Viscum album/química , Animales , Células Cultivadas , Concanavalina A/farmacología , Citocinas/genética , Electroforesis en Gel de Poliacrilamida , Expresión Génica/fisiología , Pruebas de Hemaglutinación , Ratones , Muérdago/químicaRESUMEN
Flavonoids, a group of low molecular weight phenylbenzopyrones, have various pharmacological properties including antioxidant activity, anticancer, and immunomodulatory effects. In the present study, lipopolysaccharide (LPS) and phorbol 12-myristate 13-acetate/phytohemagglutinin (PMA/PHA) were used as stimulants for RAW 264.7 macrophages and human peripheral blood mononuclear cell (hPBMC), and tumor necrosis factor (TNF)-alpha and interleukin (IL)-2 productions were measured. In addition, flavonoids were examined for their effects on LPS-induced NO production in RAW 264.7 macrophages. The results showed that all compounds were not strongly cytotoxic at the tested concentrations on hPBMC and RAW 264.7 macrophages. On immunomodulatory properties, catechin, epigallocatechin (EGC), naringenin, and fisetin repressed NO production and TNF-alpha secretion. Furthermore, catechin, epigallocatechin gallate (EGCG), epicatechin (EC), luteolin, chrysin, quercetin, and galangin increased IL-2 secretion while EGC, apigenin, and fisetin inhibited the secretion. These results indicated that flavonoids have the capacity to modulate the immune response and have a potential anti-inflammatory activity. There was no obvious structure-activity relationship regard to the chemical composition of the flavonoids and their cell biological effects.
Asunto(s)
Citocinas/biosíntesis , Flavonoides/farmacología , Leucocitos Mononucleares/metabolismo , Macrófagos/metabolismo , Óxido Nítrico/biosíntesis , Adulto , Animales , Células Cultivadas , Humanos , Interleucina-2/biosíntesis , Leucocitos Mononucleares/efectos de los fármacos , Macrófagos/efectos de los fármacos , Ratones , Persona de Mediana Edad , FN-kappa B/fisiología , Acetato de Tetradecanoilforbol/farmacología , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Flavonoids, a group of low molecular weight phenylbenzopyrones, have various pharmacological properties including antioxidant, anticancer, bactericidal, and anti-inflammatory. We carried out anti-herpetic assays on 18 flavonoids in five classes and a virus-induced cytopathic effect (CPE) inhibitory assay, plaque reduction assay, and yield reduction assay were performed. When flavonoids were applied at various concentrations to Vero cells infected by HSV-1 and 2, most of the flavonoids showed inhibitory effects on virus-induced CPE. Among the flavonoids, EC, ECG (flavanols), genistein (isoflavone), naringenin (flavanone), and quercetin (flavonol) showed a high level of CPE inhibitory activity. The antiviral activity of flavonoids were also examined by a plaque reduction assay. EC, ECG, galangin, and kaempferol showed a strong antiviral activity, and catechin, EGC, EGCG, naringenin, chrysin, baicalin, fisetin, myricetin, quercetin, and genistein showed moderate inhibitory effects against HSV-1. In these experiments, flavanols and flavonols appeared to be more active than flavones. Furthermore, treatment of Vero cells with ECG and galangin (which previously showed strong antiviral activities) before virus adsorption led to a slight enhancement of inhibition as determined by a yield reduction assay, indicating that an intracellular effect may also be involved.
Asunto(s)
Antivirales , Flavonoides/farmacología , Herpesvirus Humano 1/efectos de los fármacos , Herpesvirus Humano 2/efectos de los fármacos , Animales , Recuento de Células , Chlorocebus aethiops , Efecto Citopatogénico Viral/efectos de los fármacos , Humanos , Indicadores y Reactivos , Células Vero , Ensayo de Placa ViralRESUMEN
The extract of European mistletoe (Viscum album, L) has been used in adjuvant chemotherapy of cancer and mistletoe lectins are considered to be major active components. The present work was performed to investigate the effects of Korean mistletoe lectin (Viscum album L. coloratum agglutinin, VCA) on proliferation and apoptosis of human hepatoma cells as well as the underlying mechamisms for these effects. We showed that VCA induced apoptosis in both SK-Hep-1 (p53-positive) and Hep 3B (p53-negative) cells through p53- and p21-independent pathways. VCA induced apoptosis by down-regulation of Bcl-2 and by up-regulation of Bax functioning upstream of caspase-3 in both cell lines. In addition, we observed down-regulation of telomerase activity in both VCA-treated cells. Our results provide direct evidence of the anti-tumor potential of this biological response which comes from inhibition of telomerase and consequent inducing apoptosis. VCA-induced apoptosis is regulated by mitochondrial controlled pathway independently of p53. These findings are important for the therapy with preparation of mistletoe because they show that telomerase-dependent mechanism can be targeted by VCA in human hepatocarcinoma. Taken together, our results suggest that the VCA, considered as a telomerase-inhibitor, can be envisaged as a candidate for enhancing sensitivity of conventional anticancer drugs.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Apoptosis/efectos de los fármacos , Carcinoma Hepatocelular/patología , Inhibidores Enzimáticos/farmacología , Genes p53/efectos de los fármacos , Mitocondrias/enzimología , Preparaciones de Plantas , Proteínas de Plantas , Proteínas Proto-Oncogénicas c-bcl-2 , Telomerasa/antagonistas & inhibidores , Toxinas Biológicas/farmacología , Western Blotting , Carcinoma Hepatocelular/enzimología , Caspasa 3 , Inhibidores de Caspasas , Núcleo Celular/patología , Núcleo Celular/ultraestructura , Fragmentación del ADN/efectos de los fármacos , ADN de Neoplasias/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Citometría de Flujo , Genes bcl-2/efectos de los fármacos , Genes bcl-2/genética , Humanos , Corea (Geográfico) , Proteínas de Neoplasias/química , Proteínas de Neoplasias/aislamiento & purificación , Proteína Oncogénica p21(ras)/genética , Proteínas Proto-Oncogénicas/genética , Proteínas Inactivadoras de Ribosomas Tipo 2 , Células Tumorales Cultivadas , Proteína X Asociada a bcl-2RESUMEN
Mistletoe lectin has been reported to induce apoptosis in different cancer cell lines in vitro and to show antitumor activity against a variety of tumors in animal models. We previously demonstrated the Korean mistletoe lectin (Viscum album var. coloratum, VCA)-induced apoptosis by down-regulation of Bcl-2 and telomerase activity and by up-regulation of Bax through p53- and p21-independent pathway in hepatoma cells. In the present study, we observed the induction of apoptotic cell death through activation of caspase-3 and the inhibition of telomerase activity through transcriptional down-regulation of hTERT in the VCA-treated A253 cells. We also observed the inhibition of telomerase activity and induction of apoptosis resulted from dephosphorylation of Akt in the survival signaling pathways. In addition, combining VCA with the inhibitors of phosphatidylinositol 3-kinase (PI3-kinase) upstream of Akt, wortmannin and LY294002 showed an additive inhibitory effect of telomerase activity. In contrast, the inhibitor of protein phosphatase 2A (PP2A), okadaic acid inhibited VCA-induced dephosphorylation of Akt and inhibition of telomerase activity. Taken together, VCA induces apoptotic cell death through Akt signaling pathway in correlated with the inhibition of telomerase activity and the activation of caspase-3. From these results, together with our previous studies, we suggest that VCA triggers molecular changes that resulting in the inhibition of cell growth and the induction of apoptotic cell death of cancer cells, which suggest that VCA may be useful as chemotherapeutic agent for cancer cells.
Asunto(s)
Apoptosis , Línea Celular Tumoral , Lectinas/efectos adversos , Lectinas/química , Muérdago/química , Fosfatidilinositol 3-Quinasas/metabolismo , Telomerasa/antagonistas & inhibidores , Caspasa 3 , Caspasas/efectos adversos , Caspasas/metabolismo , Supervivencia Celular/efectos de los fármacos , Cromonas/farmacología , Ensayos de Selección de Medicamentos Antitumorales/métodos , Sinergismo Farmacológico , Humanos , Corea (Geográfico) , Lectinas/antagonistas & inhibidores , Lectinas/aislamiento & purificación , Morfolinas/farmacología , Fosfatidilinositol 3-Quinasas/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3 , Monoéster Fosfórico Hidrolasas/antagonistas & inhibidores , Monoéster Fosfórico Hidrolasas/farmacología , Fosforilación/efectos de los fármacos , Extractos Vegetales/química , Preparaciones de Plantas/efectos adversos , Preparaciones de Plantas/antagonistas & inhibidores , Preparaciones de Plantas/química , Proteínas de Plantas/efectos adversos , Proteínas de Plantas/antagonistas & inhibidores , Proteínas de Plantas/química , Proteínas Inactivadoras de Ribosomas Tipo 2 , Telomerasa/genética , Toxinas Biológicas/efectos adversos , Toxinas Biológicas/química , Regulación hacia ArribaRESUMEN
The aqueous extract of European mistletoe (Viscum album, L.) has been used in cancer therapy. The purified mistletoe lectins, main components of mistletoe, have demonstrated cytotoxic and immune-system-stimulating activities. Korean mistletoe (Viscum album L. coloratum), a subspecies of European mistletoe, has also been reported to possess anticancer and immunological activities. A galactose- and N-acetyl-D-galactosamine-specific lectin (Viscum album L. coloratum agglutinin, VCA) with Mr 60 kDa was isolated from Korean mistletoe. Mistletoe preparations have been given subcutaneously due to the low stability of lectin in the gastrointestinal (GI) tract. In the present study, we investigated the possibility of alginate/chitosan microcapsules as a tool for oral delivery of mistletoe lectin. In addition, our strategy has been to develop a system composed of stabilizing cores (granules), which contain mistletoe lectin, extract or powder, coated by a biodegradable polymer wall. Our results indicated that successful incorporation of VCA into alginate/chitosan microcapsules has been achieved and that the alginate/chitosan microcapsule protected the VCA from degradation at acidic pH values. And coating the VCA with polyacrylic polymers, Eudragit, produced outstanding results with ideal release profiles and only minimal losses of cytotoxicity after manufacturing step. The granules prepared with extract or whole plant produced the best results due to the stability in the extract or whole plant during manufacturing process.
Asunto(s)
Alginatos/química , Cápsulas , Quitina/análogos & derivados , Quitina/química , Preparaciones de Acción Retardada , Composición de Medicamentos/métodos , Ácido Glucurónico/química , Ácidos Hexurónicos/química , Muérdago , Lectinas de Plantas/aislamiento & purificación , Administración Oral , Alginatos/farmacocinética , Cloruro de Calcio/química , Cloruro de Calcio/farmacocinética , División Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Quitina/farmacocinética , Quitosano , Sistemas de Liberación de Medicamentos/métodos , Ensayos de Selección de Medicamentos Antitumorales , Ácido Glucurónico/farmacocinética , Ácidos Hexurónicos/farmacocinética , Humanos , Concentración de Iones de Hidrógeno , Componentes Aéreos de las Plantas/química , Extractos Vegetales/química , Lectinas de Plantas/química , Lectinas de Plantas/farmacología , Preparaciones de Plantas/química , Preparaciones de Plantas/aislamiento & purificación , Preparaciones de Plantas/farmacología , Proteínas de Plantas/química , Proteínas de Plantas/aislamiento & purificación , Proteínas de Plantas/farmacología , Plantas Medicinales/química , Ácidos Polimetacrílicos/química , Ácidos Polimetacrílicos/farmacocinética , Proteínas Inactivadoras de Ribosomas Tipo 2 , Toxinas Biológicas/química , Toxinas Biológicas/aislamiento & purificación , Toxinas Biológicas/farmacología , AguaRESUMEN
Recently, Viscum album var. coloratum agglutinin (VCA) was shown to have various effects on cancer cells. However, most researchers are focused on high concentrations (1-1000 ng/ml) of VCA and its anti-cancer effects. Therefore, we wanted to know whether low concentrations of VCA have an effect on proliferation and invasion of human trophoblast cells (HTR-8/SVneo cell line). Cell proliferations at low concentration of VCA (1-10 pg/ml) were increased over 2-fold relative to the control. Also, gelatinolytic activities of matrix metalloproteinase-2 were increased after VCA treatment, while TIMP-1 expression was reduced. Furthermore, the expression of integrin subunits α5 and ß1 were increased 1.5-fold when cells were treated with low dose of VCA (10 pg/ml). Lastly, VCA was able to promote trophoblast invasion through activation of the Akt signaling pathway. In conclusion, low concentrations of VCA can stimulate the ability of trophoblast cells to invade through the extracellular matrix in vitro.
Asunto(s)
Lectinas de Plantas/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Trofoblastos/efectos de los fármacos , Viscum album , Línea Celular , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Cromonas/farmacología , Humanos , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/genética , Morfolinas/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3 , ARN Mensajero/metabolismo , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Trofoblastos/fisiologíaRESUMEN
Ultraviolet (UV) A penetrates deeply into the skin and induces the generation of reactive oxygen species (ROS) causing damage to fibroblasts, which leads to aging of the skin. However, the body has developed an antioxidant defence system against the harmful effects of ROS. Enzymes such as superoxide dismutase (SOD) and catalase (CAT) play critical roles on the removal of excess ROS in living organisms. In this study, the antioxidant activities of anthocyanins (cyanidin 3-galactoside and cyanidin 3-lathyroside) from Acanthopanax divaricatus var. albeofructus (ADA) fruits were investigated by xylenol orange, thiobarbituric acid reactive substances (TBARS), and antioxidant enzyme assay. As a result, generation of H2O2 and lipid peroxide induced by UVA-irradiation in human dermal fibroblast (HDF-N) cells was reduced by treatment of anthocyanins. Also, augmented enzyme (SOD and CAT) activities were observed in UVA-irradiated cells when treated with anthocyanin. In conclusion, the results obtained show that anthocyanins from ADA fruits are potential candidates for the protection of fibroblast against the damaging effects of UVA irradiation. Furthermore, anthocyanin may be a good candidate for antioxidant agent development.
RESUMEN
A galactose- and N-acetyl-D-galactosamine-specific lectin (Viscum album L. var. coloratum agglutinin, VCA), which is known for its anticancer activity, was isolated from Korean mistletoe. This study reports a microarray analysis of the effects of VCA on an activated human T cells under various times and concentrations. A total of over 3000 genes were identified whose expression levels were significantly altered against controls after treatment with VCA and anti-CD3/CD28 antibody stimulation on human T-cells over an 8 h period. An analysis of the gene expression profile induced by VCA following incubation in human T cells revealed the activation and inhibition of genes involved in a wide range of immune functions in line with the broad mechanisms of action of VCA. These functions include cytokine gene expression, cell adhesion, cell motility, cell growth and maintenance, cell death, and the response to stress and to external stimulus. This report is aimed at providing the mistletoe research community with a robust database on which further studies could be built.
Asunto(s)
Anticuerpos/farmacología , Antígenos CD28/inmunología , Complejo CD3/inmunología , Redes Reguladoras de Genes/genética , Proteínas Inactivadoras de Ribosomas/química , Linfocitos T/efectos de los fármacos , Toxinas Biológicas/química , Viscum album/química , Adhesión Celular/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Línea Celular , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Separación Celular , Citocinas/biosíntesis , Citocinas/genética , Fragmentación del ADN , ADN Complementario/biosíntesis , ADN Complementario/genética , Electroforesis en Gel de Poliacrilamida , Ensayo de Inmunoadsorción Enzimática , Expresión Génica/efectos de los fármacos , Perfilación de la Expresión Génica , Análisis por Micromatrices , Monocitos/efectos de los fármacos , Monocitos/metabolismo , ARN Mensajero/biosíntesis , Proteínas Inactivadoras de Ribosomas Tipo 2 , Transducción de Señal/efectos de los fármacos , Estimulación QuímicaRESUMEN
A galactose- and N-acetyl-D-galactosamine-specific lectin (Viscum album L. var. coloratum agglutinin, VCA), which is known for its anti-tumor activity, was isolated from Korean mistletoe. Mistletoe preparations have been given subcutaneously because of the unstability and poor absorption in the GI tract. In this study, we investigated the effect of incubation time and glucose on the VCA transport across the in vitro model of human FAE (follicle-associated epithelium) by two different cell models: Caco-2 cell monolayers mimicking human enterocytes and a model of the human FAE which is mainly composed of M-cells and enterocytes. As a result, the VCA transport across the FAE monolayer cells was higher and faster than the transport across the Caco-2 monolayer cells, and glucose increased VCA transport across both monolayer cells. In addition, IL-12 was secreted from myeloid DC1 and lymphoid DC2.4 cells which were co-treated simultaneously with LPS and VCA. Furthermore, the FAE model associated with underlying immune cells was established and VCA was added to the inserts apically. There was a greater IL-12 secretion in dendritic cells situated below FAE monolayer than Caco-2 monolayer. The results from this study provide important insight into the possible oral application of mistletoe on anti-tumor therapeutics.
Asunto(s)
Antineoplásicos Fitogénicos/farmacocinética , Células Dendríticas Foliculares/metabolismo , Interleucina-12/metabolismo , Absorción Intestinal , Ganglios Linfáticos Agregados/citología , Lectinas de Plantas/farmacocinética , Viscum album/química , Inmunidad Adaptativa/efectos de los fármacos , Animales , Transporte Biológico , Células CACO-2 , Línea Celular , Supervivencia Celular/efectos de los fármacos , Técnicas de Cocultivo , Células Dendríticas Foliculares/efectos de los fármacos , Enterocitos/efectos de los fármacos , Enterocitos/metabolismo , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Glucosa/metabolismo , Humanos , Inmunidad Innata/efectos de los fármacos , Lipopolisacáridos/toxicidad , Linfocitos/efectos de los fármacos , Linfocitos/metabolismo , Ratones , Células Mieloides/efectos de los fármacos , Células Mieloides/metabolismo , Ganglios Linfáticos Agregados/inmunologíaRESUMEN
Viscum album L. coloratum agglutinin (VCA), isolated from Korean mistletoe, is a strong inducer of apoptosis in a variety of tumor cells; however, the underlying molecular mechanisms responsible are not clear. Here, we show that VCA induces apoptotic killing, as demonstrated by DNA fragmentation, Hoechst 33258 staining, terminal deoxynucleotidyl transferase dUTP nick-end labeling assay, and flow cytometry analysis in hepatocarcinoma Hep3B cells. VCA treatment results in a significant increase in reactive oxygen species (ROS) and loss of mitochondrial membrane potential (DeltaPsim). Furthermore, treatment with the antioxidant N-acetyl-L-cysteine reduces ROS induction by VCA, preventing apoptosis in Hep3B cells, indicating that oxidative stress is involved in VCA-mediated cell death. Our results also show rapid changes in mitochondrial transition permeability, Bax translocation, cytochrome c release, caspase-3 activity, and poly(ADP-ribose) polymerase degradation in Hep3B cells occurring in VCA-induced apoptosis. There is much evidence that implicates c-Jun NH2-terminal kinase (JNK) activation with apoptosis in a variety of cellular and animal models. In this study, we show that VCA induces JNK phosphorylation, which is abolished with pretreatment with a JNK inhibitor. Moreover, Hep3B cells overexpressing JNK1 or stress-activated protein kinase kinase (SEK1) seem to be more susceptible to cell death from ROS and loss of DeltaPsim induced by VCA, whereas expression of dominant-negative JNK1 or SEK1 in Hep3B cells do not. These data suggest that JNK phosphorylation may be a major regulator involved in VCA-induced apoptosis. Together, these results suggest that VCA induces apoptosis by inducing ROS production and a loss of DeltaPsim, in which JNK phosphorylation plays a critical role in these events.