Examples: histone, BN000065

Project: PRJNA561818

Diabetes and breast cancer are common diseases with a major impact on the health sector in Mexico and worldwide. Epidemiological and experimental works support the link between type 2 diabetes and breast cancer; these data support that insulin resistance, hyperglycemia, hyperinsulinemia, and elevated levels of IGF-1 in patients with type II diabetes mellitus promote growth and invasiveness of tumor cells. The aim of the present work was to determine, by microarray, the mechanisms of action and signaling of a hyperglycemic microenvironment in the cell line (MDA-MB-231) and its effect to treatment with cisplatin (CCP). MDA-MB-231 breast cancer cells were cultured in DMEM medium, supplemented with 10% fetal bovine serum and antibiotics at 5% CO2, at 37 ˚C. We proceeded to extract total RNA for the analysis of microarrays under LG (low glucose) and HG (high glucose) conditions; for the cDNA synthesis, it was labeled with dUTP-Cy3 or dUTP-Cy5 fluorophores, using a CyScribe Firs-Strand cDNA kit. The analysis was carried out through the KEGG pathways program; some bioenergetic metabolism processes (glycolysis, biosynthesis of purines and pyrimidines, and metabolism of glycerol phospholipids) were found altered, which fulfill the feedback function to the cellular microenvironment, activating some signaling processes, such as the Hippo route, PI3K-Akt, Jak-STAT, MAPK, Ras, Wnt/β-catenin, apoptosis, and favoring an aggressive phenotype and drug resistance in a hyperglycemic microenvironment. The microarray analysis was validated by qRT-PCr of the tetraspanin and Frizzled genes. Overall design: The cells were cultured in BG and HG media; 10 μg of total RNA were obtained and used for cDNA synthesis and labeling with a Super Script II kit Invitrogen, using the dUTP-Cy3 incorporation array for BG (control) and dUTP-Cy5 for HG. Incorporation efficiency was analyzed measuring absorbance at 555 nm for Cy3 and 655 nm for Cy5. Similar quantities of fluorophores labeled cDNA were hybridized on the oligonucleotides collection 50-mer Human10K from MWG Biotech Oligo Bio Sets (Germany). Images of the microarrays were acquired and quantified in a Scan Array 4000 using the Quant Array software from Packard BioChips (USA). A first analysis of the images and the data was performed using the Array-Pro Analyzer software from Media Cybernetics (USA). Microarray data analysis was performed using the free software program genArise, allowing background correction, normalization, intensity filter, replicate analysis, and selection of differentially expressed genes; this program was developed in the Computing Unit of the Cellular Physiology Institute of the UNAM.

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