Transcriptomic profiling in cardiovascular disease with renal failure in human

Abstract

BackgroundChronic kidney disease (CKD) is a major health issue globally; it affects 50 million people worldwide and over 26 million Adults in the United States. Studies indicate that up to 50% of deaths in patients with ens‐stage renal failure are due to cardiovascular disease (CVD) and there is a 10 to 30‐fold higher cardiovascular mortality in CKD patients than in the age‐matched general population, despite adjustment for classical cardiovascular risk factors. To profile the exhaustive signatures in calcifited human endotelials, we performed whole transcriptome analysis to explore novel genes in primary human aortic endothelial cells (HAECs) and human brain microvascular endothelial cells (HBMECs).MethodsHAECs and HBECs were treated with calcification medium, and total RNA was extracted using the Nucleospin RNA isolation kit (Clontech). cDNA was synthesized and fragmented using SMART‐seq2 and Nextera XT (Illumina), respectively to generate libraries for sequencing on Hi‐seq 2000 (Illumina). Five million single‐ended raw reads from each sample were mapped to the human reference genome consortium GRCh38 by Burrows‐Wheeler Aligner (BWA); the abundancies of each gene, (FPKM, fragment per kilo‐base exon per million mapped reads), were estimated using RNA‐seq by Expectation Maximization (RSEM). Log 2 transformed FPKM were normalized by quantile normalization, which were then subjected to differential expression analysis to calculate significant fold change (Gene‐E, Broad Institute). Gene selections were performed by the combinations of fold‐changes on log 2 ratio, and p value < 0.05. Data were analyzed using Kyoto Encyclopedia of Genes and Genomes (KEGG, http://www.genome.jp/kegg/) and The Database for Annotation, Visualization and Integrated Discovery v6.7 (DAVID, http://david.abcc.ncifcrf.gov/).ResultsPathway analysis among the top 500 up‐regulated genes in comparing untreated HAECs and HBMECs to those treated with calcification medium revealed that these genes are mainly involved in cell metabolic, proliferation, and apoptosis processes. Among these genes, they were found to play roles in cell cycle regulation, apoptosis, oxidative stress, and stress resistance such as heat shock protein 70 families (HSPA1, HSPA5, and HSP40), NF‐kappa‐B inhibitor alpha, and aryl hydrocarbon receptor nuclear translocator (ARNT) in HIF‐1, NF‐kappa B and c‐AMP signaling pathways. In addition, MT‐RNR1 and cannabinoid receptor 1 are also detected that involved in amount of ATP produced in mitochondria and co‐chaperone of HSP70.ConclusionWe have identified a group of genes differentially expressed in healthy and calcified endothelial cells. Our data contribute to our understanding of the direct effects in the presence of hyperphosphatemia and hypocalcemia at endothelial cells. This gene set may have pathophysiological significance and may serve to inform rational development of therapeutic strategies to prevent CVD adverse effects in CKD patients.