mtt-formazan and Mouth-Neoplasms

Toothpastes contain three main components: detergents, abrasives, and fluoride. Detergents, particularly sodium lauryl sulfate, have been proposed as components that enable toothpastes to produce cytotoxic effects in vitro. However, not all toothpastes contain sodium lauryl sulfate, and almost no studies have found an association between detergents and the in vitro cytotoxicity of toothpastes. The present study examined the in vitro cytotoxicity of nine commercially available toothpastes containing four different detergents. Toothpastes were diluted in serum-free medium, centrifuged, and filter sterilized. The half-lethal concentration of the toothpaste-conditioned medium (TCM) was calculated based on the formation of formazan by gingival fibroblasts, oral squamous cell carcinoma HSC-2 cells, and L929 cells. Cell proliferation was analyzed, and live-dead staining was performed, after exposure of cells to conditioned medium prepared with 1% toothpaste (1% TCM). It was found that toothpastes containing sodium lauryl sulfate and amine fluoride strongly inhibited cell viability with the half-lethal concentration being obtained with conditioned medium prepared with approximately 1% toothpaste (1% TCM). Toothpastes containing cocamidopropyl betaine and Steareth-20 showed higher half-lethal concentration values, with the half-lethal concentration being obtained with conditioned medium prepared with 10% (10% TCM) and 70% (70% TCM) toothpaste, respectively. Proliferation and live-dead data were consistent with the cell-viability analyses. These results demonstrate that the type of detergent in toothpastes can be associated with changes in in vitro cell toxicity.

Topics: Animals; Betaine; Carcinoma, Squamous Cell; Cariostatic Agents; Cell Culture Techniques; Cell Line; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cells, Cultured; Culture Media, Conditioned; Culture Media, Serum-Free; Detergents; Diamines; Epithelial Cells; Fibroblasts; Fluorescent Dyes; Fluorides; Formazans; Gingiva; Humans; Indicators and Reagents; Materials Testing; Mice; Mouth Neoplasms; Polyethylene Glycols; Sodium Dodecyl Sulfate; Tetrazolium Salts; Toothpastes

To investigate the effect of metformin on the proliferation and cell apoptosis of oral squamous cell carcinoma (OSCC) (HSC-3, HSC-4) in vitro and in vivo.. HSC-3, HSC-4 cells were treated with metformin at different concentration (2-50 mmol/L) for 24, 48 or 72 hours. In vitro cell proliferation ability was determined by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay and colony formation assay. Cell cycle progression was assessed by flow cytometry. Cell apoptosis was tested by both TdT-mediated dUTP nick-end labeling (TUNEL) assay and flow cytometry. The activation of related cell markers was examined by immunohistochemistry. Xenograft mouse model was used to demonstrate the in vivo anti-tumor effect of metformin. A total of 30 BALB/c mice were randomly divided into control groups (water + phosphate buffer saline, PBS) and treatment groups (pre-oral, oral or intraperitoneal injection). Each group had 6 mice. The tumor size was measured once every three days until endpoint (35 days). After sacrificing the mice, tumor tissue was removed, sectioning and then analyzed by TUNEL or immunohistochemistry (IHC) assays.. Metformin inhibited proliferation and colony formation of HSC-3, HSC-4 in a time- and dose-dependent manner. The cell proliferation was significantly reduced when treated with 5, 10, 20 and 40 mmol/L metformin for 48 and 72 hours (P < 0.05).The colony formation of OSCC cells treated with metformin for 72 hours in vitro had the same result. Treated with 2, 20 and 50 mmol/L metformin for 24 hours increased the ratio of G0/G1 1.2-1.8 fold compared with the control group on HSC-4 cell. The percentage of apoptosis cell rose from 10% (control) to around 30% (treatment) in vitro. Metformin also decreased the size of xenografts by 82.5% (pre-oral), 63.9% (oral), and 62.8% (oral or intraperitoneal injection). The percentage of apoptosis cell rose from lower than 10% (control) to 70% (pre-oral), 50% (oral), and 25% (oral or intraperitoneal injection). The percentage of PCNA positive cell was lower than 60% (control group was normalized to 100%).. Metformin could inhibit the growth of OSCC cell line (HSC-3, HSC-4) by reducing cell proliferation and increasing cell apoptosis in vitro and in vivo. Therefore metformin could be a potential new treatment candidate for human OSCC.

Topics: Animals; Apoptosis; Carcinoma, Squamous Cell; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Flow Cytometry; Formazans; Humans; In Vitro Techniques; Metformin; Mice; Mice, Inbred BALB C; Mouth Neoplasms; Random Allocation; Tetrazolium Salts; Time Factors; Tumor Stem Cell Assay

Molecular-receptor-targeted imaging of folate receptor positive oral carcinoma cells using folic-acid-conjugated fluorescent Au(25) nanoclusters (Au NCs) is reported. Highly fluorescent Au(25) clusters were synthesized by controlled reduction of Au(+) ions, stabilized in bovine serum albumin (BSA), using a green-chemical reducing agent, ascorbic acid (vitamin-C). For targeted-imaging-based detection of cancer cells, the clusters were conjugated with folic acid (FA) through amide linkage with the BSA shell. The bioconjugated clusters show excellent stability over a wide range of pH from 4 to 14 and fluorescence efficiency of approximately 5.7% at pH 7.4 in phosphate buffer saline (PBS), indicating effective protection of nanoclusters by serum albumin during the bioconjugation reaction and cell-cluster interaction. The nanoclusters were characterized for their physico-chemical properties, toxicity and cancer targeting efficacy in vitro. X-ray photoelectron spectroscopy (XPS) suggests binding energies correlating to metal Au 4f(7/2) approximately 83.97 eV and Au 4f(5/2) approximately 87.768 eV. Transmission electron microscopy and atomic force microscopy revealed the formation of individual nanoclusters of size approximately 1 nm and protein cluster aggregates of size approximately 8 nm. Photoluminescence studies show bright fluorescence with peak maximum at approximately 674 nm with the spectral profile covering the near-infrared (NIR) region, making it possible to image clusters at the 700-800 nm emission window where the tissue absorption of light is minimum. The cell viability and reactive oxygen toxicity studies indicate the non-toxic nature of the Au clusters up to relatively higher concentrations of 500 microg ml(-1). Receptor-targeted cancer detection using Au clusters is demonstrated on FR(+ve) oral squamous cell carcinoma (KB) and breast adenocarcinoma cell MCF-7, where the FA-conjugated Au(25) clusters were found internalized in significantly higher concentrations compared to the negative control cell lines. This study demonstrates the potential of using non-toxic fluorescent Au nanoclusters for the targeted imaging of cancer.

Topics: Carrier Proteins; Cell Line; Cell Line, Tumor; Cell Survival; Flow Cytometry; Folate Receptors, GPI-Anchored; Folic Acid; Formazans; Gold; Humans; Luminescent Measurements; Microscopy, Atomic Force; Microscopy, Electron, Transmission; Microscopy, Fluorescence; Mouth Neoplasms; Nanocomposites; Photoelectron Spectroscopy; Reactive Oxygen Species; Receptors, Cell Surface; Serum Albumin, Bovine; Spectroscopy, Fourier Transform Infrared; Tetrazolium Salts

To investigating the relation between the expression of P-glycoprotein and Glutathione transferase-pi and the chemoresistance.. The expressions of these two proteins in patients with oral and maxillofacial squamous carcinoma and normal oral tissues were detected by immunohistochemistry.. The positive expression rate of P-gp and GST-pi in oral and maxillofacial malignant tumor was 57.1% and 53.6% respectively, and no expression in normal oral tissues; the expression of GST-pi was relevant to the resistance to cisplatin, while the expression of P-gp was relevant to the resistance to chemotherapeutic drug in general.. The method of immunohistochemistry combining MTT assay in vitro may become an efficient way to predict the sensitivity to chemotherapeutic drug.

Topics: ATP Binding Cassette Transporter, Subfamily B, Member 1; Carcinoma, Squamous Cell; Drug Resistance, Neoplasm; Facial Neoplasms; Formazans; Glutathione S-Transferase pi; Glutathione Transferase; Humans; Immunohistochemistry; Isoenzymes; Maxillary Neoplasms; Mouth Neoplasms; Tetrazolium Salts