Soybean isoflavone genistein has multiple anticancer properties and its pro-apoptotic and anti-proliferative effects have been studied in different cancer Cells. However, the mechanisms of action of genistein and its molecular targets on human colon Cells have not been fully elucidated. Therefore, caspase-3 and p38 mitogen activated protein kinase (p38 MAPK) as the main therapeutic targets were investigated in this study at both gene expression and protein levels in HT29 colon cancer Cells. The caspase-3 andp38 MAPK gene expression levels were examined by real time PCR whereas flow cytometry technique was performed to determine their intracellular protein levels. The caspase-3 enzyme activity was obtained by colorimetric method while the gelatinase activity of matrix metalloproteinase-2 (MMP2) was determined by zymography. In addition, MTT test, wound healing assay and clonogenic assay were carried out to determine the effect of genistein on HT29 cell viability, migration, and proliferation, respectively. Genistein induced apoptotic death in HT29 Cells through activation of caspase-3 pathway at the transcriptional, protein, and enzymatic levels. Moreover, genistein inhibited the proliferation of HT29 Cells by reducing of bothp38 MAPK gene expression and its active phosphorylated protein level. Also, we showed that genistein strongly suppressed the metastatic potency of HT29 colon cancer Cells via the reduction of MMP2 activity. Based on the results of this study, we conclude that genistein may exhibit its anticancer properties on HT29 colon cancer Cells by modulating caspase-3 and p38 MAPK pathway at different transcriptional and protein levels.

Aims: Nanoparticles due to their wide applications in medicine,industry,and biotechnology, have attracted many scientists’ attentions. Recently, nanoparticles especially selenium nanoparticles are widely used to diagnosis and cancer treatment. The aim of this study was to evaluate the cytotoxic and anticancer effects of selenium nanoparticles on colon cancer cell line and analysis of CAD (Caspase Activated DNase) gene expression.Material and methods: In this study, colon cancer HT29 and normal HEK293 cell lines were purchased from the Pasteur Institute Cell Bank of Tehran and treated with selenium nanoparticles overnight. The Cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) (Gibco, Scotland) medium with 10% FBS serum and 1% streptomycin antibiotic (Gibco, Scotland). The Cells were then stored at 37 ° C. In this study, cytotoxic effect of Selenium NPs was evaluated on HT29 and HEK293 Cells using MTT (3-(4, 5-Dimethyltetrazollium Bromide) assay. Subsequently, they were treated with selenium nanoparticles in different concentrations (0, 7.81, 15.62, 31.25, 62.5, 125, 250 and 500 mg/mL) for 24 hours. To solubilize the viable Cells formazan crystals production, we added 100 μl/well of dimethyl sulfoxide (DMSO) to them. After treatment of HT29 Cells with IC50 concentration, the total RNA was extracted and cDNA synthesized. Moreover, CAD gene expression was evaluated using Real Time PCR method. The data was evaluated by ABI StepOne utilizing the Applied Biosystems qRT-PCR (ABI 7300 system, Applied Biosystems). The quantification of the mode of Selenium NPs -induced cell death in the HT29 Cells were ascertained using flow cytometry followed by staining with fluorescein isothiocyanate (FITC)‐Annexin V and propidium iodide (PI) staining. Finally, the study of apoptosis and necrosis of Selenium NPs was evaluated using flow cytometry method. Data analysis was statistically determined by using One-way analysis of variance (ANOVA) with SPSS/22 software followed by a Tukey test.Results: The result showed that the treatment of Selenium NPs at 31.25 to 500 µg/mL concentration had maximum cytotoxic effect, revealed statistically significant (P˂0.001). The IC50 value for Selenium NPs were measured at 75 µg/mL after 24 hours. In order to determine the effect of Selenium NPs on cancerous Cells, alterations in the mRNA expression levels of CAD gene in HT29 Cells were done by qRT-PCR technique followed by the exposure to nanoparticle. The CAD gene expression comparing to reference gene was up-regulated 4.04±0.125 fold. To determine the mechanism of cell death in the cancer Cells, annexin V/PI flow cytometry was carried out. In the treatment of HT29 Cells by IC50 of selenium NPs, 10.43%, and, 24.28% of early and late stages’ apoptosis were observed, respectivelyConclusion: Our results suggest that selenium NPs can display some promising cytotoxic properties through inducing apoptosis pathway. Based on the results, up-regulated gene expression involved in apoptosis (CAD) and activating apoptosis, it can be concluded that the selenium NPs can be used as drug candidate in colon cancer treatment, but more studies are needed regarding the medicinal importance of nanoparticles.

Background: Nowadays, investigations on the role of nanoparticles for diagnosis, and treatment of cancers are being increased. This study aimed at examining the cytotoxic effect of green synthesized silver nanoparticles (AgNPs) on human colon cancer (HT29) and normal (HEK293) Cells using the Artemisa tournefortiana extract. Materials and Methods: In the current study, the green synthesis of AgNPs was conducted using the A. tournefortiana extract. Then, the characterization of fabricated AgNPs was performed by ultraviolet-visible (UV-vis) spectroscopy, X-ray diffraction (XRD), and transmission electron microscopy (TEM) methods. The HT29 and HEK293 cell lines were treated with different concentrations of synthesized AgNPs for 24 hours and the viability of Cells and half-maximal inhibitory concentration (IC50) were calculated by the MTT assay. Results: The fabricated AgNPs showed the characteristic surface plasmon resonance peak at around 425 nm. The crystallographic shapes from the XRD and TEM tests showed that the AgNPs were mostly spherical in shape, having an average diameter of 22 nm. The MTT results revealed that AgNPs significantly decreased the viability of Cells in dose-and timedependent manner. The IC50 values of nanoparticles for HT29 and HEK293 cell lines, during the 24 hours, were 40. 71 and 61. 38 μ g/mL, respectively. Conclusion: According to the results of the current study, the green fabricated AgNPs can have a more cytotoxic effect on colon cancer Cells compared to the normal Cells. Thus, they can be considered as a promising strategy for the treatment of colon cancer.

Background: There aresomehypotheses about radiation-sensitizingandradiation-protective effects of antioxidants. Saffron, dried stigmas of Crocus sativus L., is a precious medicinal plant that contains an impressive variety of plant compounds such as crocin, crocetin, and safranal that act as antioxidants. The present study examined the cytotoxic effects of saffron extracts with different polarity and their synergism or protective effects with radiation on a colorectal cancer cell line (HT-29) and normal human fibroblasts. Objectives: The aim was to find a natural agent to improve radiotherapy efficacy. Methods: HT-29 colorectal cancer Cellsand normalhumanfibroblasts were cultured in RPMI1640medium, incubated with different concentrations of different saffron extracts (50-250  g/ml), and then were exposed to a dose of 8 Gy of X-rays. The cytotoxicity effect was determined by the MTT assay. Results: Saffron extracts decreased cell viability in HT-29 colorectal cancer Cells and normal human fibroblasts as a concentrationdependent manner. Combination radiotherapy with polar saffron extract in most doses showed synergistic effects on HT-29 cell death while it did not show any distinctive synergistic effect in normal Cells. Semi-polar and non-Polar extracts just in low doses had synergistic effects on tumor Cells. These two extracts did not show any protective effects on normal Cells. Conclusions: Among the various saffron extracts, polar saffron extract and low doses of non-polar saffron extract in combination with radiation increase radiation sensitivity and cell death in tumor Cells, while they do not increase radiation sensitivity in normal Cells and even protect normal Cells to some extent.

Background and Aim: Colorectal cancer is one of the most common types of cancer in the world, causing many deaths worldwide each year. 5-Fluorouracil is the first choice for the treatment of this type of cancer. Drug resistance and its many side effects are among the weaknesses of this drug. Silibinin, the main bioactive component of silymarin, is originally extracted from Silybum marianum, which is commonly used as an anti-hepatic agent. In recent years, the anticancer effect of silibinin has been observed on different cancer Cells. This study aimed to investigate the combined cytotoxic effects of silibinin and 5-fluorouracil on cell viability and p53 expression in the colon cancer HT29 cell line. Material and Methods: Survival of HT29 Cells was evaluated by the MTT method after 48 hours of treatment with different concentrations of silibinin and 5 fluorouracil alone and in combination. The expression level of the P53 protein was measured by Western blot method. Results: Silibinin and 5-fluorouracil significantly inhibited HT29 cell survival in a dose-dependent manner. The simultaneous use of silibinin and 5-fluorouracil showed that silibinin enhances the cytotoxic effects of 5-fluorouracil. In addition, 100 μg/ml silibinin enhanced the impact of 100 μM 5-fluorouracil on P53 protein expression. Conclusion: This study suggested that silibinin-5-fluorouracil combination may be a valuable candidate for colon cancer patients. Of course, further studies are needed to reach definitive conclusion.

Backgrond: Todays, there are considerable efforts to find plant substances as new drugs in cancer therapy. Scrophularia striata plants are a well-known in traditional medicine in Iran. This prompted us to investigate the antitumor effects of Scrophularia striata (an Iranian species) extract on human colorectal cancer (HT29) and normal (HEK 293) cell lines.Materials and Methods: In this experimental study, an extract of Scrophularia striata was prepared by maceration method. Then, the cytotoxicity effect of different concentrations of the Scrophularia striata extract was an analysis by MTT assay on HT29 and HEK293 Cells and IC50 for extracts was determined. Statistical analysis was done using SPSS software. The resulting data were analyzed using ANOVA test.Findings: Results of this study showed a significant cytotoxicity activity of Scrophularia striata extract against HT29 Cells in a dose-dependent manner. So, Scrophularia striata extract at a concentration of 10 mg/ml to 0.1 mg/ml have toxic effects on the HT29 Cells (p<0.05). However, this extract doesn’t have any cytotoxic effect on normal Cells. Conclusion: The Scrophularia striata extract has toxic effects on tumor HT29 cell line, so it can be used as an anticancer drug with specific concentration and low side-effects.

Background: Cancer stem Cells (CSCs) have critical roles in tumor initiation, progression, metastasis, resistance to chemoradiotherapy, and recurrence in various cancers such as colon cancer. Therefore, extensive studies are required to understand the CSCs mechanism of action and design novel cancer therapies. Successful isolation and characterization of CSCs from tumor tissues or cancer cell lines provide the proper opportunity for these kinds of studies. This study aimed to isolate and characterize CSCs from HT29 colon cancer cell lines. Methods: Sphere formation assay was used to isolate CSCs from the HT29 cell line. The expression of four stemness genes, including LGR5, SOX2, c-Myc, and Oct4, was assessed by real-time PCR. The CSC markers, CD 44 and CD 24, were also evaluated and compared between the parental HT29 cell line and HT29-derived spheres by flow cytometry. Statistical analysis was performed using GraphPad Prism version 6. 0 and the MannWhitney U test. Results: HT29-derived sphere Cells were successfully formed in serum-free media. The proportion of CD 24 and CD 44-expressing Cells and the expression of LGR5, SOX2, c-Myc, and Oct4 stemness genes were significantly higher in CSCs isolated from HT29 spheres compared with parental HT29 Cells. Conclusion: Sphere formation assay is a proper method for the isolation of CSCs from the HT29 cell line. The stemness markers LGR5, SOX2, c-Myc, Oct4, CD 44, and CD 24 are suitable for the characterization of these Cells.