Dataset Information

Human Hepatocyte Proteome

ABSTRACT: Here we report a comprehensive proteomic analysis of purified human hepatocytes and a human hepatoma cell line HepG2. The complete dataset comprises 9,400 proteins and provides quantitative depiction of the proteomes of hepatocytes and HepG2 cells at the protein titer and copy number dimensions.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Dr Wisniewski Jacek R.

PROVIDER: MSV000079562 | MassIVE | Mon Mar 07 16:18:00 GMT 2016

SECONDARY ACCESSION(S): PXD001874

REPOSITORIES: MassIVE

ACCESS DATA

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Publications

In-depth quantitative analysis and comparison of the human hepatocyte and hepatoma cell line HepG2 proteomes.

Wiśniewski Jacek R JR Vildhede Anna A Norén Agneta A Artursson Per P

Journal of proteomics 20160126

Hepatocytes play a pivotal role in human homeostasis. They are essential in regulation of glucose and lipid levels in blood and play a central role in metabolism of amino acids, lipids, drugs and xenobiotic-compounds. In addition, hepatocytes produce a major portion of proteins circulating in the blood. Hepatocytes were isolated from liver tissue obtained from surgical resections. Proteins were extracted and processed using filter aided sample preparation protocol and were analyzed by LC-MS/MS u ...[more]

PMID: 26825538

Similar Datasets

Project description:Triglyceride accumulation in nonalcoholic fatty liver (NAFL) results from unbalanced lipid metabolism which, in the liver, is controlled by several transcription factors. The Foxa subfamily of winged helix/forkhead box (Fox) transcription factors comprises three members which play important roles in controlling both metabolism and homeostasis through the regulation of multiple target genes in the liver, pancreas and adipose tissue. In the mouse liver, Foxa2 is repressed by insulin and mediates fasting responses. Unlike Foxa2, however, the role of Foxa1 in the liver has not yet been investigated in detail. In this study, we evaluate the role of Foxa1 in two human liver cell models, primary cultured hepatocytes and HepG2 cells, by adenoviral infection. Moreover, human and rat livers were analyzed to determine Foxa1 regulation in NAFL. Results demonstrate that Foxa1 is a potent inhibitor of hepatic triglyceride synthesis, accumulation and secretion by repressing the expression of multiple target genes of these pathways (e.g., GPAM, DGAT2, MTP, APOB). Moreover, Foxa1 represses the fatty acid transporter FATP2 and lowers fatty acid uptake. Foxa1 also increases the breakdown of fatty acids by inducing HMGCS2 and ketone body synthesis. Finally, Foxa1 is able to largely up-regulate UCP1, thereby dissipating energy and consistently decreasing the mitochondria membrane potential. We also report that human and rat NAFL have a reduced Foxa1 expression, possibly through a protein kinase C-dependent pathway. We conclude that Foxa1 is an antisteatotic factor that coordinately tunes several lipid metabolism pathways to block triglyceride accumulation in hepatocytes. However, Foxa1 is down-regulated in human and rat NAFL and, therefore, increasing Foxa1 levels could protect from steatosis. Altogether, we suggest that Foxa1 could be a novel therapeutic target for NAFL disease and insulin resistance. To determine the global impact of Foxa1 on human liver gene transcription, microarray expression analyses were performed in HepG2 cells transfected with Ad-Foxa1 or Ad-Control. We used microarrays to detail the global programme of gene expression in HepG2 cells infected with Ad-Foxa1 or control adenovirus (insertless Ad-pACC).

Project description:Triglyceride accumulation in nonalcoholic fatty liver (NAFL) results from unbalanced lipid metabolism which, in the liver, is controlled by several transcription factors. The Foxa subfamily of winged helix/forkhead box (Fox) transcription factors comprises three members which play important roles in controlling both metabolism and homeostasis through the regulation of multiple target genes in the liver, pancreas and adipose tissue. In the mouse liver, Foxa2 is repressed by insulin and mediates fasting responses. Unlike Foxa2, however, the role of Foxa1 in the liver has not yet been investigated in detail. In this study, we evaluate the role of Foxa1 in two human liver cell models, primary cultured hepatocytes and HepG2 cells, by adenoviral infection. Moreover, human and rat livers were analyzed to determine Foxa1 regulation in NAFL. Results demonstrate that Foxa1 is a potent inhibitor of hepatic triglyceride synthesis, accumulation and secretion by repressing the expression of multiple target genes of these pathways (e.g., GPAM, DGAT2, MTP, APOB). Moreover, Foxa1 represses the fatty acid transporter FATP2 and lowers fatty acid uptake. Foxa1 also increases the breakdown of fatty acids by inducing HMGCS2 and ketone body synthesis. Finally, Foxa1 is able to largely up-regulate UCP1, thereby dissipating energy and consistently decreasing the mitochondria membrane potential. We also report that human and rat NAFL have a reduced Foxa1 expression, possibly through a protein kinase C-dependent pathway. We conclude that Foxa1 is an antisteatotic factor that coordinately tunes several lipid metabolism pathways to block triglyceride accumulation in hepatocytes. However, Foxa1 is down-regulated in human and rat NAFL and, therefore, increasing Foxa1 levels could protect from steatosis. Altogether, we suggest that Foxa1 could be a novel therapeutic target for NAFL disease and insulin resistance. To determine the global impact of Foxa1 on human liver gene transcription, microarray expression analyses were performed in human hepatocytes transfected with Ad-Foxa1 or Ad-Control. We used microarrays to detail the global programme of gene expression in human hepatocytes infected with Ad-Foxa1 or control adenovirus (insertless Ad-pACC).

Project description:Primary human hepatocytes (PHH) are a main instrument in drug metabolism research and in the prediction of drug-induced phase I/II enzyme induction in humans. The HepG2 liver-derived cell line is commonly used as a surrogate for human hepatocytes, but their use in ADME and toxicity studies can be limited because of lowered basal levels of metabolizing enzymes. Despite their widespread use, the transcriptome of HepG2 cells compared to PHH is not well characterized. In this study, microarray analysis was conducted to ascertain the differences and similarities in mRNA expression between HepG2 cells and human hepatocytes before and after exposure to a panel of fluoroquinolone compounds. Comparison of the naÃ¯ve HepG2 cell and PHH transcriptomes revealed a substantial number of basal gene expression differences. When HepG2 cells were dosed with a series of fluoroquinolones, trovafloxacin, which has been associated with human idiosyncratic hepatotoxicity, induced substantially more gene expression changes than the other quinolones, similar to previous observations with PHH. While TVX-treatment resulted in many gene expression differences between HepG2 cells and PHH, there were also a number of TVX-induced commonalities, including genes involved in RNA processing and mitochondrial function. Taken together, these results provide insight for interpretation of results from drug metabolism and toxicity studies conducted with HepG2 cells in lieu of PHH, and could provide further insight into the mechanistic evaluation of TVX-induced hepatotoxicity. Experiment Overall Design: HepG2 cells were obtained from the American Type Culture Collection (Rockville, MD). The cells were cultured in Minimum Essential Medium (Invitrogen Life Technologies, Carlsbad,CA) with 10% Fetal Bovine Serum under a humidified 5% CO2 atmosphere using T-162 plastic culture flasks. The cells were split when they reached approximately 70-85% confluence after washing with sterile phosphate buffered saline and detachment of the cells with trypsin (Invitrogen Life Technologies, Carlsbad, CA). The HepG2 cells were cultured in 6-well plastic plates upon exposure to quinolone compounds. Primary human hepatocytes, obtained from In Vitro Technologies (IVT, Baltimore, MD) in 6-well type I collagen coated plates, were cultured with 2 mL of Hepatocyte Incubation Media (IVT) at 37Â°C with 5% CO2 for 24 hours after receipt. Experiment Overall Design: For the genomic experiments, quinolone compounds, dissolved in 0.1 N KOH (Sigma Chemical Co., St. Louis, MO), were added to the wells with fresh media at levels of 100 ÂµM (HepG2) or 400 ÂµM (primary human hepatocytes) for 24 hours using at least two technical replicates. Trovafloxacin was dosed using two separate preparations of HepG2 cells and two separate donors of human hepatocytes. Vehicle control cells were dosed with an equivalent volume of 0.1N KOH as the experimental samples. For intracellular comparison (HepG2 cells vs PHH), naÃ¯ve cells were harvested using TRIzolâ?¢ reagent (Invitrogen Life Technologies, Carlsbad, CA). Experiment Overall Design: Total RNA was isolated from the TRIzolâ?¢ extracts using the standard procedure from the manufacturer. O.D. at 260 nm determined RNA concentrations. RNA quality was accessed using an Agilent Technologies bioanalyzer before proceeding to microarray sample preparation. Microarray analysis was performed using the standard protocol provided by Affymetrix Inc. (Santa Clara, CA) and as previously described, starting with 5 Âµg of total RNA (Richert et al., 2006). Experiment Overall Design: Fragmented, labeled cRNA was hybridized to an Affymetrix human genome U133A array, which contains sequences corresponding to roughly 22,200 transcripts at 45Â°C overnight. The arrays were washed, developed, and scanned.