The silencing of Bcl‑xL in the non‑small cell lung cancer (NSCLC) cell line, A549, downregulates miR‑361‑5p expression. This study aimed to determine the biological effects of miR‑361‑5p on NSCLC, and to elucidate the molecular mechanisms through which apoptosis is regulated. MicroRNA (miRNA or miR) functional analyses were performed via transfection of miR‑361‑5p mimics and inhibitors, demonstrating that the inhibition of miR‑361‑5p induced the apoptosis of NSCLC cells. To elucidate the function of miR‑361‑5p in vivo, cells transfected with miR‑361‑5p inhibitors were microinjected into zebrafish embryos, and immunostained using antibodies to detect the active form of caspase‑3. Co-transfection with siBcl‑xL and miR‑361‑5p mimics illustrated the association between Bcl‑xL, miR‑361‑5p and apoptosis; miR‑361‑5p mimics blocked the apoptosis initiated by siBcl‑xL. Luciferase reporter assays identified mothers against decapentaplegic homolog 2 (SMAD2) as a novel target of miR‑361‑5p and the reduction of its protein level was validated by western blot analysis. To confirm the molecular mechanisms through which apoptosis is regulated, gene rescue experiments revealed that the ectopic expression of SMAD2 attenuated the inhibitory effects on apoptosis induced by miR‑361‑5p. In this study, to the best of our knowledge, we provide the first evidence that miR‑361‑5p functions as an oncomiR in A549 and SK‑LU‑1 cells through the regulation of SMAD2, suggesting that miR‑361‑5p may be employed as a potential therapeutic target for the miRNA-based therapy of NSCLC.
Bcl-xL is an anti-apoptotic protein that is frequently found to be overexpressed in non-small cell lung cancer leading to an inhibition of apoptosis and poor prognosis. Recently, the role of miRNAs in regulating apoptosis and cell survival during tumorigenesis has become evident, with cancer cells showing perturbed expression of various miRNAs. In this study, we utilized miRNA microarrays to determine if miRNA dysregulation in bcl-xL silenced lung adenocarcinoma cells could be involved in regulating cell death. Short interfering RNA-based transfection of A549 and SK-LU1 lung adenocarcinoma cells was successful in inducing a reduction in bcl-xL expression levels, resulting in a decrease in cell viability. A total of 10 miRNAs were found to be significantly differentially expressed when compared between siRNA-transfected and non-transfected cells including hsa-miR-181a, hsa-miR-769-5p, hsa-miR-361-5p, hsa-miR-1304 and hsa-miR-608. When overexpression studies on hsa-miR-608 was performed via transfection of miRNA mimics, cell death was found to be induced in A549 and SK-LU1 cells in comparison to untreated cells. This effect was reversed when knockdown studies involving anti-sense inhibitors were introduced. Combination of siRNA based silencing of bcl-xL (siBcl-xL) followed by anti-sense inhibitor transfection led to a decrease in the apoptotic population of A549 and SK-LU1 cells in comparison to cells only treated with siBcl-xL, illustrating the connection between bcl-xL, hsa-miR-608 and cell death. Gene target prediction analysis implicated the PI3K/AKT, WNT, TGF-β, and ERK signaling pathways as targets of bcl-xL induced miRNA alterations. We have demonstrated that bcl-xL silencing in A549 and SK-LU1 cells leads to the occurrence of cell death through the dysregulation of specific miRNAs. This study also provides a platform for anti-sense gene therapy whereby miRNA expression can be exploited to increase the apoptotic properties in lung adenocarcinoma cells.
Inositol hexaphosphate (IP6), or phytic acid is a natural dietary ingredient and has been described as a "natural cancer fighter", being an essential component of nutritional diets. The marked anti-cancer effect of IP6 has resulted in our quest for an understanding of its mechanism of action. In particular, our data provided strong evidence for the induction of apoptotic cell death, which may be attributable to the up-regulation of Bax and down-regulation of Bcl-xl in favor of apoptosis. In addition, the up-regulation of caspase-3 and -8 expression and activation of both caspases may also contribute to the apoptotic cell death of human colorectal adenocarcinoma HT-29 cells when exposed to IP6. Collectively, this present study has shown that rice bran IP6 induces apoptosis, by regulating the pro- and anti-apoptotic markers; Bax and Bcl-xl and via the activation of caspase molecules (caspase-3 and -8).
The aims of this study are to determine the mutagenicity of a locally produced polyhydroxybutyrate (PHB) using Salmonella mutagenicity test and to find out if PHB altered the expression of p53 and c-myc proto-oncogenes and bcl-xl and bcl-xs anti-apoptotic genes in the human fibroblast cell line, MRC-5. Different concentrations of PHB were incubated with special genotypic variants of Salmonella strains (TA1535, TA1537, TA1538, TA98 and TA100) carrying mutations in several genes both with and without metabolic activation (S9) and the test was assessed based on the number of revertant colonies. The average number of revertant colonies per plate treated with PHB was less than double as compared to that of negative control. For the gene expression analyses, fibroblast cell lines were treated with PHB at different concentrations and incubated for 1, 12, 24 and 48 h separately. The total RNA was isolated and analysed for the expression of p53, c-myc, bcl-xl and bcl-xs genes. The PHB did not show over or under expression of the genes studied. The above tests indicate that the locally produced PHB is non-genotoxic and does not alter the expression of the proto-oncogenes and anti-apoptotic genes considered in this study.
The interconnected Ras/ERK and PI3K/AKT pathways play a central role in colorectal tumorigenesis, and they are targets for elucidating mechanisms involved in attempts to induce colon cancer cell death. Both ginger (Zingiber officinale) and honey have been shown to exhibit anti-tumor and anti-inflammation properties against many types of cancer, including colorectal cancer. However, there are currently no reports showing the combined effect of these two dietary compounds in cancer growth inhibition. The aim of this study was to evaluate the synergistic effect of crude ginger extract and Gelam honey in combination as potential cancer chemopreventive agents against the colorectal cancer cell line HT29.