Colchicine is an alkaloid that has been widely used to treat gout. It also has a curative effect on cancer. Although many studies have shown that its effect on cell apoptosis was mediated by the activation of caspase-3, the pathways involved in the process remained obscure. Here we show some evidence regarding the missing information using human normal liver cells L-02 in our study. The effect of colchicine on apoptosis in L-02 cells and the apoptosis-associated signaling pathways were determined using different tests including cell viability assay, Annexin V and propidium idodide binding, PI staining, Hoechst 33342 staining, mitochondrial membrane potential assay, caspase activity assay and Western blot analysis. We found that colchicine-induced a dose-dependent drop of cell viability in L-02 cells; early apoptosis happened when cells were treated with 0.1μM of colchicine. The colchicine-induced loss of mitochondrial membrane potential, activation of caspase-3 and 9, up-regulation of Bax and down-regulation of Bcl-2 showed an evidence for the colchicine activity on apoptosis, at least, by acting via the intrinsic apoptotic pathway.
"At this concentration it was also not highly effective against clonogenic cells. It has been shown recently that colchicine exerted anti-proliferative effect in hepatocellular carcinoma cell lines , but the significance of this finding is diminished by observation that it also reduced viability in normal liver cells . Given these results, colchicine is probably not a good drug candidate for the treatment of melanoma. "
[Show abstract][Hide abstract] ABSTRACT: Accumulating evidence supports the concept that melanoma is highly heterogeneous and sustained by a small subpopulation of melanoma stem-like cells. Those cells are considered as responsible for tumor resistance to therapies. Moreover, melanoma cells are characterized by their high phenotypic plasticity. Consequently, both melanoma stem-like cells and their more differentiated progeny must be eradicated to achieve durable cure. By reevaluating compounds in heterogeneous melanoma populations, it might be possible to select compounds with activity not only against fast-cycling cells but also against cancer stem-like cells. Natural compounds were the focus of the present study.
We analyzed 120 compounds from The Natural Products Set II to identify compounds active against melanoma populations grown in an anchorage-independent manner and enriched with cells exerting self-renewing capacity. Cell viability, cell cycle arrest, apoptosis, gene expression, clonogenic survival and label-retention were analyzed.
Several compounds efficiently eradicated cells with clonogenic capacity and nanaomycin A, streptonigrin and toyocamycin were effective at 0.1 µM. Other anti-clonogenic but not highly cytotoxic compounds such as bryostatin 1, siomycin A, illudin M, michellamine B and pentoxifylline markedly reduced the frequency of ABCB5 (ATP-binding cassette, sub-family B, member 5)-positive cells. On the contrary, treatment with maytansine and colchicine selected for cells expressing this transporter. Maytansine, streptonigrin, toyocamycin and colchicine, even if highly cytotoxic, left a small subpopulation of slow-dividing cells unaffected. Compounds selected in the present study differentially altered the expression of melanocyte/melanoma specific microphthalmia-associated transcription factor (MITF) and proto-oncogene c-MYC.
Selected anti-clonogenic compounds might be further investigated as potential adjuvants targeting melanoma stem-like cells in the combined anti-melanoma therapy, whereas selected cytotoxic but not anti-clonogenic compounds, which increased the frequency of ABCB5-positive cells and remained slow-cycling cells unaffected, might be considered as a tool to enrich cultures with cells exhibiting melanoma stem cell characteristics.
PLoS ONE 03/2014; 9(3):e90783. DOI:10.1371/journal.pone.0090783 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: 2,4-Dichlorophenoxyacetic acid (2,4-D), the most widely used herbicide in the world, has been previously reported to induce lung damage. Here in this study we have investigated the molecular mechanism of 2,4-D induced lung toxicity in A549 and WI38 cell lines. Cell viability experiments indicate the IC50 values in A549 and WI38 cells for 72 h are 126 ± 2.25 μM and 115 ± 4.39 μM, respectively. Although not arresting a particular phase of the cell cycle of A549 cells, 2,4-D dose dependently increased the subG1 population, indicative of cell death, and the mode of cell death is apoptosis in both cell lines, as observed by annexin V/PI flow cytometric analysis and the expression status of pro and anti apoptotic proteins by western blot analysis. Dose dependent shrinkage of A549 cells indicates that 2,4-D can disrupt the microtubule network, and this is confirmed by immunofluorescence studies in vitro. In a cell free system, we found that 2,4-D depolymerises the microtubule network in vitro (IC50 = 212 ± 1.63 μM). 2,4-D can quench the intrinsic tryptophan fluorescence of tubulin in both a time and a dose dependent manner; the stoichiometry of 2,4-D–tubulin binding is 1 : 1 and the dissociation constant is 22.82 ± 1.29 μM. In silico studies indicate that 2,4-D binds to tubulin between the α and β subunits, very close to the colchicine binding site, and there is very little conformational change of the tubulin structure, as we also confirmed by circular dichroism studies. So, in brief, these results suggest that disruption of the cellular tubulin-microtubule network is one of the key mechanisms in the induction of lung cytotoxicity by 2,4-D.
Toxicology Research 01/2014; 3(2). DOI:10.1039/c3tx50082a · 3.98 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Studies suggest an important role of autophagy as a target for cancer therapy. We constructed a "disease-gene-drug" network using the modular approach of bioinformatics and screened herbal monomers demonstrating functions related to autophagy regulation.
Based on the microarray results of the gene expression omnibus (GEO) database (GSE2435 and GSE31040, starvation-induced autophagy model), we used the human protein reference database (HPRD) to obtain the protein-protein interaction (PPI) network. In addition, we used the CFinder software to identify several functional modules, performed gene ontology-biological process (GO-BP) functional enrichment analysis using the DAVID software, constructed a herbal monomer-module gene regulatory network using literature search and the Cytoscape software, and then analyzed the relationships between autophagy, genes, and herbal monomers.
We screened 544 differentially expressed genes related to autophagy, 375 pairs of differentially expressed genes, and 7 gene modules, wherein the functions of module 3 (composed of 7 genes) were enriched in "cell death". Using the constructed herbal monomer-module gene regulatory network, we found that 30 herbal monomers can simultaneously regulate these 7 genes, indicating a potential regulatory role in autophagy.
Database screening using the disease-gene-drug network can provide new strategies and ideas for the application of herbal medicines in cancer therapy.
BMC Complementary and Alternative Medicine 12/2014; 14(1):466. DOI:10.1186/1472-6882-14-466 · 2.02 Impact Factor
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