Steatohepatitis/Metabolic Liver Disease
Differential pattern of lipid droplet-associated proteins and de novo perilipin expression in hepatocyte steatogenesis†
Version of Record online: 7 FEB 2008
Copyright © 2008 American Association for the Study of Liver Diseases
Volume 47, Issue 6, pages 1936–1946, June 2008
How to Cite
Straub, B. K., Stoeffel, P., Heid, H., Zimbelmann, R. and Schirmacher, P. (2008), Differential pattern of lipid droplet-associated proteins and de novo perilipin expression in hepatocyte steatogenesis. Hepatology, 47: 1936–1946. doi: 10.1002/hep.22268
Potential conflict of interest: Nothing to report.
- Issue online: 28 MAY 2008
- Version of Record online: 7 FEB 2008
- Accepted manuscript online: 7 FEB 2008 12:00AM EST
- Manuscript Accepted: 28 JAN 2008
- Manuscript Received: 15 SEP 2007
- Ministry of Science, Research, and the Arts of Baden-Württemberg. Grant Number: Az: 23-7532.22-23-12/1
Supplementary material for this article can be found on the H EPATOLOGY Web site ( http://interscience.wiley.com/jpages/0270-9139/suppmat/index.html ).
|hep22268-Suppl1final.tif||357K||Suppl. fig.1 Scheme of supposed human perilipin splice variants and specific primer pairs used for rtPCR.A) Human perilipin splice variants A-D are deduced from Lu and coauthors (19), the sequence dimensions are according to the NCBI Map Viewer (http://www.ncbi.nlm.nih.gov/mapview/; released August 23rd, 2007). B) Primer pairs used for amplification of perilipin splice variants. The molecular size of the respective PCR products are 687 bp for PeriA (with PanPeri); 496 bp for PeriB; 373 bp for PeriC and 508 bp for PeriD. Abbreviations are as follows: E: exon; black boxes: ATG-codon; brown boxes: coding sequences (cds); white boxes: non coding sequences; f, forward primer (green); r, reverse primer (red).|
|hep22268-Suppl2final.tif||350K||Suppl. fig.2 rtPCR analysis of perilipin splice variants present in human liver.Using specific primer pairs that detected TIP47, adipophilin as well as different splice variants of human perilipin (Peri A, B, C and D; cf. suppl. fig.1), cDNAs obtained from human adult liver tissues (hLiver, lane 2) as well as primary human hepatocytes of patients with different BMI (phH 1: BMI <25, lane 3; phH2: 25<BMI<30, lane 4; phH3: BMI: >30, lane 5) were analyzed in comparison to human adult adipose tissue (hAdipo, lane 1) as well as human adult adrenal gland (hAdrenGl, lane 6) and human adult testis (hTestis, lane 7) used as positive controls: Gene products specific for TIP47 and adipophilin (AP) are detectable in all tissues analysed; their presence in adipose tissue is most likely due to the presence of blood vessels within the tissue. Gene products specific for perilipin splice variant A and B were detected in human adipose tissue, in liver tissue and primary human hepatocytes, as well as in human testis, according to the calculated size of 678 bp for perilipin A and 496 bp for perilipin B, whereas both are absent in human adrenal gland. For perilipin C and D, however, a specific gene product of 373 bp for perilipin C and 508 bp for perilipin D was amplified in all total cDNAs tested.|
|hep22268-Suppl3final.tif||167K||Suppl. fig.3 Specificity of antibodies against PAT proteins.The specificity of the antibodies against TIP47, adipophilin (AP) and perilipin (Peri) was confirmed by immunoblotting following SDS-PAGE of recombinant human TIP47 (lane 1, recTIP47), adipophilin (lane 2, recAP) and perilipin (lane 3, recPeri). Molecular mass markers are indicated on the right margin.|
|hep22268-Suppl4final.tif||4051K||Suppl. fig.4 Immunofluorescence microscopy of PAT proteins in models of steatosisin vivoand in vitro.A) Confocal laser scanning microscopy of TIP47 (green) together with nile red as well as adipophilin (AP; green) together with perilipin (Peri, red) in hepatocellular carcinoma-derived cells of the line PLC/PRF-5, after treatment with BSA-coupled oleate to enhance LD-formation. B) Confocal laser scanning microscopy of adipophilin (green) with perilipin (red) in steatotic liver specimens of obese mice. Note the absence of perilipin in oleate-treated cells as well as murine steatotic liver. Respective DIC images are given on the right side. Bars: 20 μm.|
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