A growing interest in apoptosis, programmed cell death, in the last years is observed and leads to better understanding of molecular mechanisms during cell-cell signaling, cell-environment interaction and screening of drugs. This in turn results in emerging of new assays and development of more accurate kits for fast and early detection of apoptosis. However, their sensitivity and reliability have often been scrutinized. Here we introduce a rapid and improved method of DNA ladder apoptosis assay for evaluating apoptosis in mammalian cells. NIH-3T3 cell line was used in this study. After treatment of cells with apoptotic agent, 500 μM H2O2 at 48 hours, DNA was extracted. Then an update protocol of DNA ladder assay was applied for detection of apoptosis. Flow cytometry and DAPI staining were performed to verify apoptosis. Primary and late apoptosis in the H2O2-treated cells was determined by flow cytometry analysis. DAPI Staining used to show DNA damage and DNA ladder assay using 1.5% gel electrophoresis showed fragmentation in the DNA of treated cells. In this research we aimed to improve DNA ladder assay to the high quality detection of apoptosis in mammalian cells. In our strategy, employing a practical DNA extraction protocol, DNA ladder assay could be applied as an easy/fast method for apoptosis detection. This improved method is able to detect apoptosis in a cost effective/timely manner without need for commercial kits and special equipment.

Introduction: The oral tumor is the sixth most prevalent type of cancer worldwide and the second leading cause of cancer-related mortality. Although chemotherapy and immunotherapy are the main strategies for the treatment of oral cancer, an emergence of inevitable resistance to these treatment modalities is the major drawback that causes recurrence of the disease. Nowadays, probiotics have been suggested as adjunctive and complementary treatment modalities for improving the impacts of chemotherapy and immunotherapy agents. Probiotics, the friendly microflora in our bodies, contribute to the production of useful metabolites with positive effects on the immune system against various diseases such as cancer. Methods: Lactobacillus plantarum is one of the most important bacteria, which commensally live in the human oral system. In the current study, the impacts of L. plantarum on maintaining oral system health were investigated, and the molecular mechanisms of inhibition of oral cancer KB cells mediated by L. plantarum were evaluated using real-time polymerase chain reaction (PCR) and FACS flow cytometry analyses. Results: Our findings showed that L. plantarum is effective in the signal transduction of the oral cancer cells through upregulation and downregulation of PTEN and MAPK pathways, respectively. Conclusion: Based on the biological effects of oral candidate probiotics candidate bacterium L. plantarum on functional expression of PTEN and MAPK pathways, this microorganism seems to play a key role in controlling undesired cancer development in the oral system. Taken all, L. plantarum is proposed as a potential candidate for probiotics cancer therapy.

Butylated hydroxyanisole (BHA) is a synthetic phenolic food additive that is used to prevent oils, fats, and shortenings from oxidative deterioration and rancidity. It was tested for potential cytotoxicity and genotoxicity upon A549 lung cancer cells. MTT assay and flow cytometry analysis were used for cytotoxicity assessment, whereas genotoxicity was evaluated in vitro by DAPI staining, alkaline comet, and DNA fragmentation assays. BHA dose- and time-dependently decreased the growth of A549 cells with an IC₅₀ of ∼0.55, 0.4, and 0.3 mM BHA at 24, 48, and 72 h, respectively. Primary and late apoptosis in the treated cells was determined using the flow cytometry analyses. In addition, single-strand DNA breakage has been observed through the comet assay technique. In addition, the morphology of DAPI-stained cells and DNA fragmentation assay using gel electrophoresis showed clear fragmentation in the chromatin and DNA rings within the nucleus of cells treated with BHA.