Project description:free fatty acids(palmitate, oleate, linoleate) 0.7mM and tnf-alpha (0 20,100 ng/ml) were subjected to HepG2 cell line to study the cytotoxicity induced by these two factors Keywords: stress response
Project description:The regulation of complex cellular activities in palmitate treated HepG2 cells, and the ensuing cytotoxic phenotype, involves cooperative interactions between genes. While previous approaches have largely focused on identifying individual target genes, elucidating interacting genes has thus far remained elusive. We applied the concept of information synergy to reconstruct a ?gene-cooperativity? network for palmititate-induced cytotoxicity in liver cells. Our approach integrated gene expression data with metabolic profiles to select a subset of genes for network reconstruction. Subsequent analysis of the network revealed insulin signaling as the most significantly enriched pathway, and desmoplakin (DSP) as its top neighbor. We determined that palmitate significantly reduces DSP expression, and treatment with insulin restores the lost expression of DSP. Insulin resistance is a common pathological feature of fatty liver and related ailments, whereas loss of DSP has been noted in liver carcinoma. Reduced DSP expression can lead to loss of cell-cell adhesion via desmosomes, and disrupt the keratin intermediate filament network. Our findings suggest that DSP expression may be perturbed by palmitate and, along with insulin resistance, may play a role in palmitate induced cytotoxicity, and serve as potential targets for further studies on non-alcoholic fatty liver disease (NAFLD). free fatty acids(palmitate, oleate, linoleate) 0.7mM and tnf-alpha (0 20,100 ng/ml) were subjected to HepG2 cell line to study the cytotoxicity induced by these two factors. control(Hepg2 medium and BSA medium) treatment(combinations of TNFa+FFAs) two biological replicates for each condition, color swap for each sample
Project description:We report the transcriptome of human endothelial cells (cell line: TIME) after supplementation with polyunsaturated fatty acids (PUFA) and/or stimulation with pro-inflammatory cytokines. The fatty acids docosahexaenoic acid (DHA, C22:6n3) or arachidonic acid (AA, C20:4n6) were included in the culture medium in concentrations of 15 µmol/L using ethanol as a vehicle (0.2 % v/v final ethanol concentration). Cells were cultured in the enriched media totaling 144 h. Stimulation of cells was performed in the last 24 h of fatty acid supplementation by addition of the cytokines IL-1β, TNF-α, and IFN-γ each in a concentration of 5 ng/ml.
Project description:TNF-alpha has a number of pro-atherogenic effects in macrovascular endothelial cells, including induction of leukocyte adhesion molecules and chemokines. We investigated the role of acyl-CoA synthetase 3 (ACSL3) in the response of cultured human macrovascular endothelial cells to TNF-alpha. TNF-alpha induced ACSL3 both in human umbilical vein endothelial cells (HUVECs) and in human coronary artery endothelial cells (HCAECs). RNA sequencing demonstrated that knockdown of ACSL3 had no marked effects on the TNF-alpha transcriptome in HCAECs. Instead, ACSL3 was required for TNF-alpha-induced lipid droplet formation from fatty acids.
Project description:The response of HepG2-SF cells exposed to 100 µM fatty acids during 7 days was investigated using stable isotope labeling by amino acids in cell culture (SILAC) for analysis of the full proteome
Project description:We report the miRNA profiles of macrophages (cell line: RAW264.7) and endothelial cells (cell line: TIME) after supplementation with polyunsaturated fatty acids (PUFA) and stimulation with LPS/pro-inflammatory cytokines. The fatty acids docosahexaenoic acid (DHA, C22:6n3) or arachidonic acid (AA, C20:4n6) were included in the culture medium in concentrations of 15 µmol/L using ethanol as a vehicle (0.2 % v/v final ethanol concentration). Cells were cultured in the enriched media totaling either 72 h (RAW264.7) or 144 h (TIME). Stimulation of cells was performed in the last 24 h of fatty acid supplementation by addition of either LPS (1 µg/mL; from E. coli serotype 0111:B4) for cell line RAW264.7 or the cytokines IL-1β, TNF-α, and IFN-γ each in a concentration of 5 ng/ml for cell line TIME.
Project description:There is growing evidence that proteolytic cleavage of histone N-terminal tails, known as histone clipping, influences nucleosome dynamics and functional properties. Using middle-down MS analysis we characterized intact and proteolytically processed histone H3 proteoforms isolated from human hepatocarcinoma cell line HepG2/C3A grown in 3D culture.
Project description:Maternal obesity during pregnancy leads to a pro-inflammatory milieu in the placenta. We conducted a global transcriptomic profiling in BeWo cells following palmitic acid (PA, 500 uM) and/or TNF-alpha (10 ng/ml) treatment for 24 h. Microarray analysis revealed that placental cytotrophoblasts increased expression of genes related to inflammation, stress response and immediate-early factors in response to plamitic acid, TNF-alpha or a combination of both. Our results suggest that fatty acids and inflammatory cytokines induce inflammation in placental cells via activation of JNK-Egr-1 signaling. global transcriptomic profiling in BeWo cells following palmitic acid (PA, 500 uM) and/or TNF-alpha (10 ng/ml) treatment for 24 h
Project description:Elaidic acid is an abundant industrial produced trans fatty acid in foodstuffs. Trans fatty acid intake has been correlated to an unfavorable plasma lipoprotein profile and an increased cardiovascular disease risk. The present study aimed to identify a plasma protein biomarker panel related to human intake of elaidic acid. The human liver cell line HepG2-SF was used as a model system, and the cells were maintained for seven days in serum-free medium containing 100 µM elaidic acid (trans∆9-C18:1), oleic acid (cis∆9-C18:1) or stearic acid (C18:0). The secretomes were analyzed by stable isotope labeling of amino acids in cell culture (SILAC), difference in gel electrophoresis (DIGE) and gene expression microarray analysis. Twelve proteins were found to be differentially regulated based on SILAC data (>1.3 fold change, P-value < 0.05), 13 proteins were found to be differentially regulated based on DIGE analysis (>1.3 fold change, P-value < 0.05), and 17 mRNA transcripts encoding extracellular proteins were determined to be affected (>1.3 fold change, P-value < 0.01) following the addition of elaidic acid compared to oleic acid or stearic acid. The results revealed that 37 proteins were regulated specifically in response to elaidic acid exposure, and nine of these proteins were confirmed to be regulated in this manner by using selected reaction monitoring mass spectrometry. HepG2 cells kept used in serum free conditions. The cells were incubated for seven days in either 100 µM oleic, elaidic or stearic acid, before total RNA was extracted. For each group four biological replicates were made and from the each RNA extraction two technical replicates were made. There is a control group without fatty acid incubation.