Hypoxia-induced gene expression pattern in doxorubicin resistant MCF7 cells

Purpose: To investigate hypoxia-induced gene expression pattern in doxorubicin-resistant human breast cancer cells (MCF7). Methods: Human breast cancer cells (MCF7) were exposed to 60 episodes of 8 h hypoxia thrice a week for three months. Chemo-resistance to doxorubicin was assessed using 3-(4,5-dimethylthiazol-2- yl)-2, 5-diphenyl tetrazolium bromide (MTT) cell proliferation assay. Real-time quantitative polymerase chain reaction (qRT-PCR) assay was performed to assess gene expression pattern in doxorubicinresistant cells on exposure to hypoxia. Results: Hypoxia significantly increased the resistance of MCF7 cells to doxorubicin, with a maximum of 16.42-fold enhancement after 25 episodes of 8-h hypoxia, while the resistance thereafter significantly decreased with prolonged episodes of hypoxia (p < 0.05). Gene expression analysis revealed significant changes in 42 genes. The expressions of 10 of these genes were significantly upregulated, while those of 32 genes were significantly down-regulated (p < 0.05). Cytochrome P450 family 1, subfamily A, member1 (CYP1A1) was the most conspicuous upregulated gene (13.32-fold), while breast cancer gene 1 (BRCA1) was the most down-regulated (8.23-fold). Gene expression analysis after 60 episodes of 8-h hypoxia revealed the upregulation of CYP1A1 (5.77-fold). Similarly, 27 genes were significantly down-regulated, with BRCA2 as the most down-regulated gene (8.11-fold). Topoisomerase (DNA) II alpha (TOP2A) was the most down-regulated among genes involved in drug metabolism and resistance (6.37-fold), while cyclin-dependent kinase 2 (CDK2) was the most profoundly downregulated among genes involved in cell cycle regulation (3.56-fold). Conclusion: These results indicate that development of resistance to doxorubicin by MCF7 cells after short-term hypoxia results from the upregulation of genes responsible for the metabolism of doxorubicin and for shifting the cells to alternative pathway driven principally by EGF and ESR2. The observed down-regulation is an adaptation of the MCF7 cells to survive under long-term hypoxia

The effect of cycling hypoxia on MCF-7 cancer stem cells and the impact of their microenvironment on angiogenesis using human umbilical vein endothelial cells (HUVECs) as a model

Background Breast cancer is the most common type of cancer among females. Hypoxia mediates cancer hallmarks and results from reduced oxygen level due to irregularities in tumor vascularization or when the tumor size prevents oxygen diffusion and triggers angiogenesis to compensate for low oxygen. Cancer stem cells (CSCs) are a rare subpopulation, able to self-renew and to give rise to tumor-initiating cells. It is proposed that CSCs’ secretions help to recruit endothelial cells via angiogenic factors to establish tumor vascularization. In the tumor microenvironment, the effect of hypoxia on CSCs and the impact of their secretions on triggering angiogenesis and tumor vascularization remain questionable. In this study, three-dimensional (3D) CSCs derived from MCF-7 were directly exposed to repetitive long-term cycles of hypoxia to assess its effect on CSCs and then to evaluate the role of the hypoxic CSCs’ (CSCsHYP) secretions in angiogenesis using (HUVECs) as a model for tumor neovascularization response. Methods CSCs derived from MCF-7 cell-line were expanded under repetitive, strictly optimized, long-term/continuous and intermittent hypoxic shots for almost four months to assess hypoxic effect on CSCs, sorted based on CD44+/CD24− biomarkers. Hypoxic phenotype of CSCsHYP was evaluated by assessing the acquired chemoresistance using MTT assay and elevated stemness properties were assessed by flow cytometry. To evaluate the effect of the secretions from CSCsHYP on angiogenesis, HUVECs were exposed to CSCsHYP conditioned-medium (CdM)—in which CSCs had been previously grown—to mimic the tumor microenvironment and to assess the effect of the secretions from CSCsHYP on the HUVECs’ capability of tube formation, migration and wound healing. Additionally, co-culture of CSCsHYP with HUVECs was performed. Results CSCsHYP acquired higher chemoresistance, increased stemness properties and obtained greater propagation, migration, and wound healing capacities, when compared to CSCs in normoxic condition (CSCsNOR). HUVECs’ tube formation and migration abilities were mediated by hypoxic (CSCs) conditioned media (CdM). Discussion This study demonstrates that chemoresistant and migrational properties of CSCs are enhanced under hypoxia to a certain extent. The microenvironment of CSCsHYP contributes to tumor angiogenesis and migration. Hypoxia is a key player in tumor angiogenesis mediated by CSCs

Synthesis, and Antitumor Activity of Some N1-(Coumarin-7-yl) Amidrazones and Related Congeners

A series of new N1-(coumarin-7-yl)amidrazones incorporating N-piperazines and related congeners were synthesized by reacting the hydrazonoyl chloride derived from 7-amino-4-methylcoumarin with the appropriate piperazines. The chemical structures of the newly prepared compounds were supported by elemental analyses, 1H-NMR, 13C-NMR, and ESI-HRMS spectral data. The antitumor activity of the newly synthesized compounds was evaluated. Among all the compounds tested, 7-{2-[1-(4-(1-benzyl-2-ethyl-4-nitro-1H-imidazol-5-yl)piperazin-1-yl)-2-oxopropylidene]hydrazinyl}-4-methyl-2H-chromen-2-one (3n) was the most potent against MCF-7 and K562 cells, with IC50 values of 20.2 and 9.3 μM, respectively

An intronic single-nucleotide polymorphism (rs13217795) in FOXO3 is associated with asthma and allergic rhinitis: a case–case–control study

Abstract Background Asthma and allergic rhinitis are respiratory diseases with a significant global burden. Forkhead box O3 (FOXO3) is a gene involved in the etiology of a number of respiratory diseases. The objective of this study is to assess the association of rs13217795, an intronic FOXO3 single-nucleotide polymorphism, with asthma and allergic rhinitis. Methods In this case–case–control genetic association study, genotyping was conducted using the PCR–RFLP method. Genotype-based associations were investigated under the general, recessive, and dominant models of disease penetrance using binomial logistic regression; and, allele-based associations were tested using Pearson’s chi-squared test. Results The final study population consisted of 94 controls, 124 asthmatics, and 110 allergic rhinitis patients. The general and recessive models of disease penetrance were statistically significant for both case–control comparisons. Under the general model, the odds of the asthma phenotype were 1.46 (0.64 to 3.34) and 3.42 (1.37 to 8.57) times higher in heterozygotes and derived allele homozygotes, respectively, compared to ancestral allele homozygotes. The corresponding odds ratios for the allergic rhinitis phenotype were 1.05 (0.46 to 2.40) and 2.35 (0.96 to 5.73), respectively. The dominant model of disease penetrance was not statistically significant. The minor allele in all study groups was the ancestral allele, with a frequency of 0.49 in controls. There was no deviation from Hardy–Weinberg equilibrium in controls. Both case–control allele-based associations were statistically significant. Conclusions Herein we present the first report of the association between rs13217795 and allergic rhinitis, and the first independent verification of the association between rs13217795 and asthma. Marker selection in future genetic association studies of asthma and allergic rhinitis should include functional polymorphisms in linkage disequilibrium with rs13217795

Phytochemical Analysis and Anticancer Properties of Drimia maritima Bulb Extracts on Colorectal Cancer Cells

Cancer is a worldwide health problem and is the second leading cause of death after heart disease. Due to the high cost and severe side effects associated with chemotherapy treatments, natural products with anticancer therapeutic potential may play a promising role in anticancer therapy. The purpose of this study was to investigate the cytotoxic and apoptotic characteristics of the aqueous Drimia maritima bulb extract on Caco-2 and COLO-205 colorectal cancer cells. In order to reach such a purpose, the chemical composition was examined using the GC-MS method, and the selective antiproliferative effect was determined in colon cancer cell lines in normal gingival fibroblasts. The intracellular ROS, mitochondrial membrane potential, and gene expression changes in selected genes (CASP8, TNF-α, and IL-6 genes) were assessed to determine the molecular mechanism of the antitumor effect of the extract. GC-MS results revealed the presence of fifty-seven compounds, and Proscillaridin A was the predominant secondary metabolite in the extract. The IC50 of D. maritima bulb extract on Caco-2, COLO-205, and the normal human gingival fibroblasts were obtained at 0.9 µg/mL, 2.3 µg/mL, and 13.1 µg/mL, respectively. The apoptotic effect assay indicated that the bulb extract induced apoptosis in both colon cancer cell lines. D. maritima bulb extract was only able to induce statistically significant ROS levels in COLO-205 cells in a dose-dependent manner. The mitochondrial membrane potential (MMP) revealed a significant decrease in the MMP of Caco-2 and COLO-205 to various concentrations of the bulb extract. At the molecular level, RT-qPCR was used to assess gene expression of CASP8, TNF-α, and IL-6 genes in Caco-2 and COLO-205 cancer cells. The results showed that the expression of pro-inflammatory genes TNF-α and IL-6 were upregulated. The apoptotic initiator gene CASP8 was also upregulated in the Caco-2 cell line and did not reach significance in COLO-205 cells. These results lead to the conclusion that D. maritima extract induced cell death in both cell lines and may have the potential to be used in CRC therapy in the future

Phytochemical Analysis and Anticancer Properties of <i>Drimia maritima</i> Bulb Extracts on Colorectal Cancer Cells

Cancer is a worldwide health problem and is the second leading cause of death after heart disease. Due to the high cost and severe side effects associated with chemotherapy treatments, natural products with anticancer therapeutic potential may play a promising role in anticancer therapy. The purpose of this study was to investigate the cytotoxic and apoptotic characteristics of the aqueous Drimia maritima bulb extract on Caco-2 and COLO-205 colorectal cancer cells. In order to reach such a purpose, the chemical composition was examined using the GC-MS method, and the selective antiproliferative effect was determined in colon cancer cell lines in normal gingival fibroblasts. The intracellular ROS, mitochondrial membrane potential, and gene expression changes in selected genes (CASP8, TNF-α, and IL-6 genes) were assessed to determine the molecular mechanism of the antitumor effect of the extract. GC-MS results revealed the presence of fifty-seven compounds, and Proscillaridin A was the predominant secondary metabolite in the extract. The IC50 of D. maritima bulb extract on Caco-2, COLO-205, and the normal human gingival fibroblasts were obtained at 0.9 µg/mL, 2.3 µg/mL, and 13.1 µg/mL, respectively. The apoptotic effect assay indicated that the bulb extract induced apoptosis in both colon cancer cell lines. D. maritima bulb extract was only able to induce statistically significant ROS levels in COLO-205 cells in a dose-dependent manner. The mitochondrial membrane potential (MMP) revealed a significant decrease in the MMP of Caco-2 and COLO-205 to various concentrations of the bulb extract. At the molecular level, RT-qPCR was used to assess gene expression of CASP8, TNF-α, and IL-6 genes in Caco-2 and COLO-205 cancer cells. The results showed that the expression of pro-inflammatory genes TNF-α and IL-6 were upregulated. The apoptotic initiator gene CASP8 was also upregulated in the Caco-2 cell line and did not reach significance in COLO-205 cells. These results lead to the conclusion that D. maritima extract induced cell death in both cell lines and may have the potential to be used in CRC therapy in the future

Molecular Accounting and Profiling of Human Respiratory Microbial Communities: Toward Precision Medicine by Targeting the Respiratory Microbiome for Disease Diagnosis and Treatment

The wide diversity of microbiota at the genera and species levels across sites and individuals is related to various causes and the observed differences between individuals. Efforts are underway to further understand and characterize the human-associated microbiota and its microbiome. Using 16S rDNA as a genetic marker for bacterial identification improved the detection and profiling of qualitative and quantitative changes within a bacterial population. In this light, this review provides a comprehensive overview of the basic concepts and clinical applications of the respiratory microbiome, alongside an in-depth explanation of the molecular targets and the potential relationship between the respiratory microbiome and respiratory disease pathogenesis. The paucity of robust evidence supporting the correlation between the respiratory microbiome and disease pathogenesis is currently the main challenge for not considering the microbiome as a novel druggable target for therapeutic intervention. Therefore, further studies are needed, especially prospective studies, to identify other drivers of microbiome diversity and to better understand the changes in the lung microbiome along with the potential association with disease and medications. Thus, finding a therapeutic target and unfolding its clinical significance would be crucial

In vitro and in vivo evaluation of the antiangiogenic activities of Trigonella foenum-graecum extracts

Objective: To assess the antiangiogenic activity of fenugreek. Methods: Different fractions of fenugreek crude extracts were prepared and their antiangiogenic properties were assessed using the ex vivo rat aortic ring assay and in vivo chicken embryo chorioallantoic membrane (CAM) assay. They were investigated for their direct cytotoxic activity in the MCF7 cells using the MTT assay. Results: The ethanol extract showed 100% inhibition of blood vessel outgrowth from primary tissue explants in the rat aortic ring assay at a concentration of 100 μg/mL while the other extracts did not show significant antiangiogenic activity. The ethanol extract was therefore investigated at varying concentrations and exhibited a significant dose dependent effect. The CAM assay coincided with the results of the aortic ring assay as ethanol extract showed a significant inhibition of formation of new blood vessels. The extracts only showed anti-proliferative activity at the highest concentration of 400 μg/mL towards MCF7 breast cancer cell lines in the MTT assay. Conclusions: Findings of the both assays confirmed that the ethanol extract inhibited vascularization significantly. Further studies on the ethanol extract would be beneficial in isolating the active ingredient responsible for the inhibition