RESUMEN
Oxidative stress contributes to cancer pathologies and to apoptosis. Marine algae exhibit cytotoxic, antiproliferative and apoptotic effects; their metabolites have been used to treat many types of cancer. We investigated in culture extracts of Petalonia fascia, Jania longifurca and Halimeda tuna to determine their effects on mouse neuroblastoma cell line, NA2B. NA2B cells were treated with algae extracts, and the survival and proliferation of NA2B cells were assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The effects of algae extracts on oxidative stress in NA2B cells also were investigated using nitric oxide synthase (NOS) immunocytochemistry and apoptosis was assessed using terminal deoxynucleotidyl transferase dUTP nick end labeling. We observed significant neurite inhibition with moderate damage by the neurotoxicity-screening test (NST) at IC50 dilutions of the extracts. MTT demonstrated that J. longifurca extracts were more toxic than P. fascia and H. tuna extracts. We found an increase of endothelial and inducible NOS immunostaining for oxidative stress and TUNEL analysis revealed increased apoptosis after application of extract. Our findings suggest that the algae we tested may have potential use for treatment of cancer.
Asunto(s)
Extractos Celulares , Neuritas/efectos de los fármacos , Phaeophyceae/química , Animales , Apoptosis/efectos de los fármacos , Extractos Celulares/química , Extractos Celulares/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Inmunohistoquímica , Concentración 50 Inhibidora , Biología Marina , RatonesRESUMEN
AIM: The apoptotic effect of geldanamycin derivative may be important for the colorectal cancer therapy. The mechanisms of apoptosis require understanding of the behavior of colon cancer cell line Colo-205 which mimics colon adenocarcinoma. Therefore, the effect of IC50 dose of 17-allylamino-17-demethoxygeldanamycin (17-AAG) on the colon cancer cells in vitro was studied for its anti-apoptotic activity. METHOD: Apoptotic ratio of the Colo-205 cells was determined after 17-AAG application with terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining and apoptosis related genes. Apoptosis signal path related key mitochondrial proteins, cytochrome c, bcl-2, caspase 9 and Apaf-1 expression were examined with RT-PCR method. RESULTS: 17-AAG caused induction of cell death. Apoptotic related genes such as cytochrome-c, Apaf-1 and caspase-9 protein expressions were increased significantly (p < 0.05) and anti-apoptotic bcl-2 expression was decreased significantly (p < 0.05). Our results indicated that the application of 17-AAG on Colo-205 cells showed anticancer effect by the apoptosis due to alteration of apoptotic genes. CONCLUSION: The apoptotic effect of 17-AAG as an natural product for alternative medicine would be very important for the success and quality of life during the treatment of colon carcinoma with the combination of anticancer drugs (Tab. 1, Fig. 2, Ref. 32).
Asunto(s)
Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Benzoquinonas/farmacología , Neoplasias del Colon/tratamiento farmacológico , Lactamas Macrocíclicas/farmacología , Neoplasias del Colon/patología , Humanos , Células Tumorales Cultivadas/efectos de los fármacosRESUMEN
We investigated the effects of different implant biomaterials on cultured canine bone marrow stromal cells (BMSC) undergoing differentiation into osteoblasts (dBMSC). BMSC were isolated from canine humerus by marrow aspiration, cultured and differentiated on calcium phosphate scaffold (CPS), hydroxyapatite, hydroxyapatite in gel form and titanium mesh. We used the MTT method to determine the effects of osteogenic media on proliferation. The characteristics of dBMSC were assessed using alizarin red (AR), immunocytochemistry and osteoblastic markers including alkaline phosphatase/von Kossa (ALP/VK), osteocalcin (OC) and osteonectin (ON), and ELISA. The morphology of dBMSC on the biomaterials was investigated using inverted phase contrast microscopy and scanning electron microscopy. We detected expression of ALP/VK, AR, OC and ON by day 7 of culture; expression increased from day 14 until day 21. CPS supported the best adhesion, cell spreading, proliferation and differentiation of BMSCs. The effects of the biomaterials depended on their surface properties. Expression of osteoblastic markers showed that canine dBMSCs became functional osteoblasts. Tissue engineered stem cells can be useful clinically for autologous implants for treating bone wounds.
Asunto(s)
Materiales Biocompatibles/farmacología , Células de la Médula Ósea/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Osteoblastos/citología , Osteoblastos/efectos de los fármacos , Animales , Antraquinonas/química , Antraquinonas/farmacología , Materiales Biocompatibles/química , Biomarcadores/análisis , Células de la Médula Ósea/citología , Modelos Animales de Enfermedad , Perros , Inmunohistoquímica , Coloración y EtiquetadoRESUMEN
OBJECTIVE: We aimed to differentiate dental pulp stem cells (DPSC) to odontoblast-like cells (ODPSC) and to investigate their attachment and growth on dentin in the presence of extra calcium by colorimetric assay and scanning electron microscopy (SEM). METHODS: After isolation of DPSC, they were differentiated to ODPSC. Standard dentin discs from human molar teeth were prepared. While the dentin discs in Group 1 did not receive any extra treatment, the discs in Group 2 were treated with acidic calcium phosphate precipitation (CPP) solution. In Group 3, the discs were suspended in phosphate buffered saline containing calcium. DPSC or ODPSC (3×10(4) cells/mL) were seeded on all discs and incubated for 7, 14 or 21 days. Attachment and growth of 7-day cell cultures on extra dentin samples were examined by SEM. MTT assay showed that number of cells on dentin surfaces was increased by time periods regardless of type of treatment and cells (p<0.05). RESULTS: While DPSC and ODPSC showed similar proliferation rates at 7 and 14days (p>0.05), the number of ODPSC was higher than DPSC in 21-day samples (p=0.039). MTT assay showed that number of cells on dentin surfaces was increased by time periods regardless of type of treatment and cells (p<0.05). Calcium-treated dentin surfaces always had lower number of cells; being significant for only CPP-treated surfaces (p<0.01). Both types of cells demonstrated good attachment and proliferation on dentin surfaces regardless of type of dentin treatment. CONCLUSIONS: Because the nature of dentin surface itself showed good adhesive characteristics with ODPSC and DPSC, additional calcium treatment of dentin surfaces may not be necessary.
Asunto(s)
Calcio/farmacología , Pulpa Dental/citología , Dentina/citología , Células Madre/citología , Fosfatos de Calcio/química , Adhesión Celular/efectos de los fármacos , Adhesión Celular/fisiología , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , Proliferación Celular/efectos de los fármacos , Proliferación Celular/fisiología , Células Cultivadas , Pulpa Dental/efectos de los fármacos , Pulpa Dental/metabolismo , Dentina/ultraestructura , Humanos , Inmunohistoquímica , Microscopía Electrónica de Rastreo , Diente Molar/citología , Odontoblastos/citología , Odontoblastos/efectos de los fármacos , Odontoblastos/metabolismo , Odontoblastos/ultraestructura , Cloruro de Sodio , Células Madre/efectos de los fármacos , Células Madre/metabolismo , Propiedades de SuperficieRESUMEN
Bone marrow contains mesenchymal stem cells that form many tissues. Various scaffolds are available for bone reconstruction by tissue engineering. Osteoblastic differentiated bone marrow stromal cells (BMSC) promote osteogenesis on scaffolds and stimulate bone regeneration. We investigated the use of cultured autologous BMSC on different scaffolds for healing defects in tibias of adult male canines. BMSC were isolated from canine humerus bone marrow, differentiated into osteoblasts in culture and loaded onto porous ceramic scaffolds including hydroxyapatite 1, hydroxyapatite gel and calcium phosphate. Osteoblast differentiation was verified by osteonectine and osteocalcine immunocytochemistry. The scaffolds with stromal cells were implanted in the tibial defect. Scaffolds without stromal cells were used as controls. Sections from the defects were processed for histological, ultrastructural, immunohistochemical and histomorphometric analyses to analyze the healing of the defects. BMSC were spread, allowed to proliferate and differentiate to osteoblasts as shown by alizarin red histochemistry, and osteocalcine and osteonectine immunostaining. Scanning electron microscopy showed that BMSC on the scaffolds were more active and adhesive to the calcium phosphate scaffold compared to the others. Macroscopic bone formation was observed in all groups, but scaffolds with stromal cells produced significantly better results. Bone healing occurred earlier and faster with stromal cells on the calcium phosphate scaffold and produced more callus compared to other scaffolds. Tissue healing and osteoblastic marker expression also were better with stromal cells on the scaffolds. Increased trabecula formation, cell density and decreased fibrosis were observed in the calcium phosphate scaffold with stromal cells. Autologous cultured stromal cells on the scaffolds were useful for healing of canine tibial bone defects. The calcium phosphate scaffold was the best for both cell differentiation in vitro and bone regeneration in vivo. It may be possible to improve healing of bone defects in humans using stem cells from bone marrow.
Asunto(s)
Células de la Médula Ósea/citología , Regeneración Ósea/fisiología , Huesos/citología , Diferenciación Celular/fisiología , Células Madre Mesenquimatosas/citología , Cicatrización de Heridas/efectos de los fármacos , Animales , Células de la Médula Ósea/efectos de los fármacos , Fosfatos de Calcio/farmacología , Células Cultivadas , Perros , Durapatita/farmacología , Masculino , Osteoblastos/citología , Osteogénesis/efectos de los fármacos , Osteogénesis/fisiología , Ingeniería de Tejidos/métodosRESUMEN
Abstract We investigated the cytotoxic, neurotoxic, apoptotic and antiproliferative effects of extracts from Petalonia fascia, Jania longifurca and Halimeda tuna on the MCF-7 breast cancer cell line. J. longifurca extracts were more toxic than those of P. fascia and H. tuna. The algal extracts showed significant toxic effects at different dilutions. The toxic effects were due to increased oxidative stress and resulted in apoptosis. Algal toxicity may exert negative effects through the food chain or by direct interaction. Algal toxicity also has potential for cancer therapy. The toxic effects that we observed may be especially important for therapy for breast tumors.
Asunto(s)
Antineoplásicos/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Chlorophyta/química , Phaeophyceae/química , Extractos Vegetales/farmacología , Rhodophyta/química , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Femenino , Humanos , Dosificación Letal Mediana , Células MCF-7 , Neurotoxinas/farmacología , Extractos Vegetales/toxicidad , Agua de Mar/microbiologíaRESUMEN
Bone marrow is a complex tissue composed of hematopoietic and stromal stem cells with the potential to differentiate into adipogenic, fibroblastic, reticular, osteogenic and chondrogenic lineages. Identification of differentiation markers during transformation of stromal cells into osteoblasts in a time-dependent manner may be informative for cell-based tissue engineering. Therefore, we investigated the effects of osteogenic medium (OM) on the proliferation and differentiation of rat bone marrow stromal cells (BMSCs). BMSCs from adult male rat tibia and femur were collected and cultured in alpha-MEM medium with 10% fetal bovine serum, penicillin, streptomycin and gentamycin. After three days of culture, the medium covering the adherent cells in culture was changed to OM containing dexamethasone, Na-beta-glycerophosphate and ascorbic acid. As a control, cell culture was also continued in the original medium for the same time period. Differentiated osteoblast cells were collected after 7, 10, 14, 21 and 30 days of culture, fixed with 4% paraformaldehyde and their immunolabelling for osteoblast markers osteonectin (ON) and osteocalcin (OC) was assessed using an indirect immunoperoxidase technique. Immunolabelling of ON and OC was detectable from day 10 of culture, began to increase on day 14, and increased steadily through to day 21. Labelling was highest on day 30 and was more intense in cells cultured with OM compared to the culture without OM. The control cells cultured in the absence of OM produced negligible levels of both markers. In conclusion, our culture system facilitated differentiation of BMSCs into osteoblasts featuring osteoblast markers, and these cells may be useful in autologous bone implant for the treatment of bone wound healing.