Veera L.D. Badisa

Protective effects of N-acetylcysteine against cadmium-induced damage in cultured rat normal liver cells

In this study, the protective effects of N-acetylcysteine (NAC), a precursor of reduced glutathione, were studied by measuring the viability, the levels of antioxidant enzymes, and by analyzing the cell cycle in cadmium (Cd)-treated rat liver cells. The cells were treated with 150 µM CdCl2 alone or co-treated with 150 µM CdCl2 and 5 mM NAC (2 h pre-, simultaneous or 2 h post-treatment) for 24 h. The viability of the cells treated with 150 µM CdCl2 alone decreased to 40.1%, while that of the cells co-treated with 5 mM NAC (pre-, simultaneous and post-treatment) significantly increased to 83.7, 86.2 and 83.7%, respectively in comparison to the control cells (100%). The catalase enzyme level decreased to undetectable level in the cells treated with CdCl2 alone, while it significantly increased in the co-treated cells (pre-, simultaneous and post-treatment) to 40.1, 34.3 and 13.2%, respectively. In the cells treated with CdCl2 alone, the glutathione peroxidase enzyme level decreased to 78.3%, while it increased in the co-treated cells (pre-, simultaneous, and post-treatment) to 84.5, 83.3 and 87.9%, respectively. The glutathione reductase enzyme level decreased to 56.1% in the cells treated with cadmium alone, but significantly increased in the cells co treated with NAC (pre-, simultaneous and post-treatment) to 79.5, 78.5 and 78.2%, respectively. Cd caused cell cycle arrest at the S and G2/M phases. The co-treatment with NAC inhibited cell cycle arrest by shifting the cells to the G1 phase. These results clearly show the protective effects of NAC against Cd-induced damage in rat liver cells.

Synthesis and in vitro evaluation of 3-(4-nitrophenyl)coumarin derivatives in tumor cell lines

Coumarins are naturally-occurring compounds that have attracted considerable interest due to their numerous biological activities depending on their pattern of substitution on the coumarin molecule. In this present investigation, we synthesized 3-(4-nitrophenyl)coumarin derivatives (9a-e) and evaluated their in vitro cytotoxic effect on human lung (A549), breast (MDA-MB-231) and prostate (PC3) cancer cell lines for 48 h using crystal violet dye binding assay. Cytotoxic effects of the most active compound on normal human lung (MRC-9) and breast (MCF-10A) cell lines, cell cycle analysis using flow cytometry and mitochondrial membrane potential (MMP) using Tetramethyl Rhodamine Methyl Ester (TMRM; rhodamine-123) fluorescent dye were also examined. Among the compounds that were evaluated, 9c showed cytotoxic effect (active), caused significant cells arrest (p<0.05) in G0/G1 and S phases of cell cycle and loss of MMP in A459, MDA-MB-231 and PC3 cell lines. Additionally, the cytotoxic effect of 9c was compared to reference drugs (Coumarin and Docetaxel) for comparative study. These results further demonstrate that acetoxy group at C-7 and C-8 positions of 9c are responsible for the observed cytotoxic effect in these cancer cell lines.

Effect of cadmium on the expression levels of interleukin-1α and interleukin-10 cytokines in human lung cells

Cadmium is an environmentally hazardous metal, which causes toxicity in humans. Inhalation of cigarette smoke and industrial fumes containing cadmium are sources of cadmium exposure. It is responsible for the malfunction of various organs, leading to disease particularly in the lungs, liver and kidneys. In the present study, the effect of cadmium chloride (CdCl2) on cell viability, and the expression levels of interleukin (IL)-1α and IL-10 cytokines at various concentrations and incubation durations were assessed in MRC-9 human normal lung and A549 human lung cancer cells to elucidate the mechanism of cadmium toxicity. Cell viability was measured using a crystal violet dye binding assay. The expression levels of the cytokines were measured by cytokine specific enzyme-linked immunosorbent assay kits. The viability assay results revealed higher sensitivity of the A549 lung cancer cells to CdCl2 compared with the normal MRC-9 lung cells. In the normal MRC-9 lung cells, higher expression levels of the cytokines were observed at the lowest CdCl2 concentration at a shorter exposure time compared with the lung cancer cells. Higher levels of the cytokines were observed in the A549 lung cancer cells at all other times and concentrations compared with the MRC-9 cells, indicating higher levels of inflammation. The cytokine levels were reduced at higher CdCl2 concentrations and longer exposure durations, demonstrating the toxic effect of cadmium. The results indicated that CdCl2 affected the expression levels of the cytokines and led to cytotoxicity in human lung cells, and suggested that compounds which reduce inflammation may prevent cadmium toxicity.

Mitigative action of monoisoamyl-2,3-dimercaptosuccinate (MiADMS) against cadmium-induced damage in cultured rat normal liver cells.

Cadmium is non-essential, carcinogenic and multitarget pollutant in the environment. Monoisoamyl-2,3-dimercaptosuccinate (MiADMS) is an ester of dimercaptosuccinic acid that acts as an antioxidant and chelator. Therefore, the mitigative action of MiADMS on viability, morphology, antioxidative enzymes and cell cycle were studied on rat liver cells treated with cadmium chloride (CdCl2). The cells were treated with 150 μM CdCl2 alone or cotreated with 300 μM MiADMS (concurrently, 2 h or 4 h post-CdCl2 treatment) for 24 h. The viability of cells treated with CdCl2 alone was decreased in comparison to the control cells. Cotreatment with MiADMS resulted in an increase in cell viability in comparison to the CdCl2 alone treated cells. The CdCl2 treatment altered the morphological shape of the cells, while cotreatment with MiADMS restored the shape. Antioxidative enzymes activities were decreased in the cells treated with CdCl2 alone, while MiADMS cotreatment resulted in an increase in enzyme activities. The CdCl2 arrested the cells in S phase of the cell cycle. Cotreatment with MiADMS alleviated cell cycle arrest by shifting to G1 phase. These results clearly show the mitigative action of MiADMS on CdCl2 toxicity and may suggest that MiADMS can be used as an antidote against cadmium.

Cytotoxic Activities of Snow Lotus Filtrates Against Human Cancer Cell Lines and Origin of Active Factor(s)

Snow Lotus (SL) filtrate was prepared by incubation of SL grains with pasteurized and unboiled 4% milk for 24 h at room temperature and tested on cultured human cancer cells, namely, human colon adenocarcinoma (HCA), HepG2, MCF-7 and compared with a normal mouse (NCTC clone 929) cell line. It was observed that this filtrate exhibited cytotoxic activities in HCA (LD50 16.6 µl/ml) and HepG2 (LD50 17.1 µl/ml) cells and to a lesser extent in MCF-7 (LD50 28.1 µl/ml) and normal mouse cells (LD50 24.3 µl/ml). Different SL filtrates were also prepared by incubation of grains with 2, 1 and 0% fat content milks. These filtrates exhibited moderate to high cytotoxic activities. Interestingly, the filtrate obtained from 2% milk showed significant differential cytotoxic activity in the range of 39 to 44% between human cancer and mouse cell lines. The cytotoxic studies were also carried out with filtrate of shorter incubation period, where no cytotoxic activity observed. Similarly, SL water filtrate was also inactive in all cell cultures. The origin of cytotoxic factor(s) in the filtrates is discussed.

7,8-Dihydroxy-3-(4-nitrophenyl)coumarin induces cell death via reactive oxygen species-independent S-phase cell arrest: MUSA et al.

We herein report the synthesis and in vitro cytotoxicity of 3‐arylcoumarin derivatives (6a‐f and 7a‐f) in human liver (HepG2), prostate (LNCap), and pancreatic (BxPC3) cancer cell lines. Among the tested compounds, 7,8‐dihydroxy‐3‐(4‐nitrophenyl) coumarin (7b) showed the highest cytotoxicity in the HepG2 cell line. The mechanism of cytotoxic action indicated that compound (7b) arrested HepG2 cells at the S phase of the cell cycle progression, induced loss of mitochondrial membrane potential, and caused reactive oxygen species (ROS)‐independent cell death. The cell viability result of pretreated HepG2 cells with antioxidant N‐acetylcysteine followed by compound (7b) treatment and the free radical scavenging activities of compound (7b) confirmed the ROS‐independent cell death. These results demonstrate that compound (7b) could serve as a valuable template for the development of novel synthetic compounds as potential anticancer agents for hepatocellular carcinoma treatment.

Comparative whole genome transcriptome analysis and fenugreek leaf extract modulation on cadmium‑induced toxicity in liver cells

Cadmium (Cd), an economically valuable metal, is widely used in various industrial processes. Although it is of economic value, it is hazardous to human health. Cd accumulates in vital organs where it causes various diseases. Natural compounds with chelating or antioxidant properties have been tested to reduce the toxic effect of Cd. The anti-oxidant, anti-diabetic and hypocholesterolemic properties of fenugreek (Trigonella foenum-graecum) leaves make it a candidate for investigation as protective agent against Cd-induced toxicity. In the present study, the protective effects of fenugreek leaf extract (FLE) on cell viability, morphology, and whole genomic transcription in cadmium chloride (CdCl2)-treated rat liver cells were analyzed. The cells were treated with 25 µM CdCl2 alone, or co-treated with 5 µg/ml FLE for 48 h. The co-treated cells were pretreated with FLE for 2 or 4 h, followed by CdCl2 treatment. Genomic transcription analysis was performed in the CdCl2-treated cells following treatment for 6 h. The CdCl2 caused a significant decrease in viability (35.8±4.1%) and morphological distortion of the cells, compared with the untreated control cells; whereas 4 h pretreatment with FLE (5 µg/ml) reversed the Cd-induced morphology alteration and increased the cell viability to 102±3.8%. Genomic transcription analysis of the CdCl2 only-treated cells showed 61 upregulated and 124 downregulated genes, compared with 180 upregulated and 162 downregulated genes in the FLE pretreated cells. Furthermore, 37 and 26% of the affected total genomic genes in the CdCl2 only-treated cells were involved in binding and catalytic activities, respectively, whereas 50 and 20% of the genes in the FLE pretreated cells were involved in binding and catalytic activities, respectively. In conclusion, these results suggested that genome transcriptome modulation may be important in the protective effect of FLE against Cd-induced toxicity in normal rat liver cells.

Abstract C51: In vitro cytotoxicity of 3-arylcoumarin derivatives in human prostate (PC3) cancer cell line

Coumarins are classified as a member of the benzopyrone family of compounds with diverse and interesting biological activities. They have been used as therapeutic agents in the treatment of various diseases. In the present study, we evaluated the in vitro cytotoxic activity of 3-arylcoumarins (1-11) in human prostate (PC3) cancer and (WPE1-NA22) normal cell lines at various concentrations (0, 10, 25, 50, 75 and 100 μM) after 48 h treatment using crystal violet dye binding assay. The most active compounds cytotoxicity was examined by cell cycle analysis, reactive oxygen species (ROS) production, mitochondrial membrane potential (MMP) measurement and Bax protein expression. Our findings indicate Result that 8-(acetyloxy)-3-(4-methanesulfonylphenyl)-2-oxo-2H-chromen-7-ylacetate (11) showed cytotoxic activity in PC3 cancer cell line (LD50 = 35.0 μM) and no cytotoxic activity in WPE1-NA22 normal cell line (inactive; LD50 = >100 μM) in comparison to the other synthesized analogs. Furthermore, It caused significant cells arrest (p Grant Support: Research supported by Florida A & M University Title III Program Citation Format: Musiliyu A. Musa, Moise Y. Joseph, Lekan M. Latinwo, Veera L. Badisa. In vitro cytotoxicity of 3-arylcoumarin derivatives in human prostate (PC3) cancer cell line. [abstract]. In: Proceedings of the Sixth AACR Conference: The Science of Cancer Health Disparities; Dec 6–9, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2014;23(11 Suppl):Abstract nr C51. doi:10.1158/1538-7755.DISP13-C51