ascorbic-acid and Sarcoma--Ewing

Current treatment of osteosarcoma is associated with poor prognosis, especially due to the increased risk of developing other cancers with chemotherapy. Therefore, new, safe and effective treatment strategies are needed. We investigated the effect of a unique mixture of nutrients containing lysine, proline, arginine, ascorbic acid, and epigallocatechin gallate (EGCG) on human osteosarcoma cell lines U-2OS, MNNG-HOS, and Ewing's sarcoma SK-ES-1 by measuring: cell proliferation, expression of matrix metalloproteinase-2 (MMP-2), MMP-9, and invasive and angiogenesis potential. Cell proliferation was evaluated by MTT assay, matrix metalloproteinases (MMP) expression by gelatinase zymography, VEGF expression by ELISA, and invasion through Matrigel. Cells were also treated with phorbol 12-myristate 13-acetate (PMA) to study enhanced MMP and VEGF expression. The invasion of osteosarcoma U-2OS and MNNG-HOS cells through Matrigel was significantly reduced in a dose-dependent fashion, with 100% inhibition of invasion of U-2OS cells at 100 microg/ml, and MNNG cells at 50 microg/ml concentration of the synergistically acting nutrient mixture. Ewing's sarcoma SK-ES-1 cells were not invasive. Nutrient synergy (NS) exhibited a dose response antiproliferative effect on osteosarcoma U-2OS cells, reaching 67% at 1000 microg/ml of NS; no significant suppression of cell proliferation was seen with MNNG or Ewing's sarcoma cells. Zymography showed dose-dependent inhibition of MMP secretion by all three cell lines in the presence of NS. VEGF secretion by U-2OS cells was completely blocked at 500 microg/ml of NS. Our results suggest NS is an excellent candidate for therapeutic use in the treatment of osteosarcoma, by inhibiting cancer cell invasion, and secretion of MMPs and VEGF, all critical parameters for cancer control and prevention.

Topics: Arginine; Ascorbic Acid; Bone Neoplasms; Catechin; Cell Proliferation; Dose-Response Relationship, Drug; Humans; Lysine; Matrix Metalloproteinases; Neovascularization, Pathologic; Nutritional Physiological Phenomena; Osteosarcoma; Proline; Sarcoma, Ewing; Tetradecanoylphorbol Acetate; Tumor Cells, Cultured; Vascular Endothelial Growth Factor A

Betulinic acid (BA) is known to induce apoptosis in melanoma neuroectodermal and malignant brain cancer cell lines. Present report describes the role of antioxidants on the BA-induced toxicity to human cell line SK-N-MC. Hydrophilic antioxidants viz., L-ascorbic acid (VitC) and N-acetyl-L-cysteine (l-NAC) had no protective effect on BA-induced apoptosis at the maximal concentrations tested. The lipophilic antioxidant, alpha-DL-tocopherol (VitE) showed a concentration and a time dependent effect on the protection of SK-N-MC cells from BA-induced apoptosis. The apoptotic parameters were analyzed using FACS analysis of propidium iodide (PI) stained nuclei, PS externalization using Annexin-V assay and changes in mitochondrial membrane potential. Generation of superoxide radical was monitored by the fluorescent dye hydroethidium (HE). Cells showed Annexin-V positivity and an increase in the propidium iodide (PI) uptake in the early hours of treatment with BA, which was concomitant with the loss of mitochondrial membrane potential. Addition of alpha-DL-tocopherol to the cell cultures 1-h prior to the treatment with BA abolished all the effects of BA-induced apoptosis. These observations suggest that BA initiates events at membrane level leading to induction of apoptosis. The observed ineffectiveness of hydrophilic antioxidants and substantial protection by lipophilic antioxidants indicate involvement of membrane-associated damages that form the basis of BA-induced cytotoxicity.

Topics: Acetylcysteine; alpha-Tocopherol; Annexin A5; Antioxidants; Apoptosis; Ascorbic Acid; Betulinic Acid; Cell Line; Cell Line, Tumor; Cell Survival; Humans; Membrane Potentials; Mitochondria; Pentacyclic Triterpenes; Reactive Oxygen Species; Sarcoma, Ewing; Triterpenes