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Fetuin-A influences vascular cell growth and production of proinflammatory and angiogenic proteins by human perivascular fat cells

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Abstract

Aims/hypothesis

Fetuin-A (alpha2-Heremans-Schmid glycoprotein), a liver-derived circulating glycoprotein, contributes to lipid disorders, diabetes and cardiovascular diseases. In a previous study we found that perivascular fat cells (PVFCs) have a higher angiogenic potential than other fat cell types. The aim was to examine whether fetuin-A influences PVFC and vascular cell growth and the expression and secretion of proinflammatory and angiogenic proteins, and whether TLR4-independent pathways are involved.

Methods

Mono- and co-cultures of human PVFCs and endothelial cells were treated with fetuin-A and/or palmitate for 6–72 h. Proteins were quantified by ELISA and Luminex, mRNA expression by real-time PCR, and cell growth by BrDU-ELISA. Some PVFCs were preincubated with a nuclear factor κB NFκBp65 inhibitor, or the toll-like receptor 4 (TLR4) inhibitor CLI-095, or phosphoinositide 3-kinase (PI3K)/Akt inhibitors and/or stimulated with insulin. Intracellular forkhead box protein O1 (FoxO1), NFκBp65 and inhibitor of κB kinase β (IKKβ) localisation was visualised by immunostaining.

Results

PVFCs expressed and secreted IL-6, IL-8, plasminogen activator inhibitor 1 (PAI-1), basic fibroblast growth factor (bFGF), platelet-derived growth factor (PDGF)-BB, monocyte chemotactic protein-1 (MCP-1), vascular endothelial growth factor (VEGF), placental growth factor (PLGF) and hepatocyte growth factor (HGF). Fetuin-A upregulated IL-6 and IL-8, and this was potentiated by palmitate and blocked by CLI-095. Immunostaining and electrophoretic mobility shift assay (EMSA) showed partial NFκBp65 activation. MCP-1 was upregulated and blocked by CLI-095, but not by palmitate. However, HGF was downregulated, which was slightly potentiated by palmitate. This effect persisted after TLR4 pathway blockade. Stimulation of insulin–PI3K–Akt signalling by insulin resulted in nuclear FoxO1 extrusion and HGF upregulation. Fetuin-A counteracted these insulin effects.

Conclusions/interpretation

Fetuin-A and/or palmitate influence the expression of proinflammatory and angiogenic proteins only partially via TLR4 signalling. HGF downregulation seems to be mediated by interference with the insulin-dependent receptor tyrosine kinase pathway. Fetuin-A may also influence angiogenic and proinflammatory proteins involved in atherosclerosis.

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Abbreviations

bFGF:

Basic FGF

EC:

Endothelial cell

EMSA:

Electrophoretic mobility shift assays

FGF:

Fibroblast growth factor

FoxO1:

Forkhead box protein O1

HGF:

Hepatocyte growth factor

IKKβ:

Inhibitor of κB kinase β

LPS:

Lipopolysaccharide

MCP-1:

Monocyte chemotactic protein-1

NFκB:

Nuclear factor κB

PAI-1:

Plasminogen activator inhibitor-1

PDGF:

Platelet-derived growth factor

PI3K:

Phosphoinositide 3-kinase

PVAT:

Perivascular adipose tissue

PVFC:

Perivascular fat cell

PLGF:

Placental growth factor

SMC:

Smooth muscle cell

TLR4:

Toll-like receptor 4

TSP-1:

Thrombospondin-1

VEGF:

Vascular endothelial growth factor

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Funding

The present study was funded by the Federal Ministry for Education and Research (FkZ 01GI 0925), a grant from the German Federal Ministry of Education and Research to the German Center for Diabetes Research and a grant of from the German Federal Ministry for Education and Research (FkZ 01KQ0902F), joint research project ‘Gesundheitsregion REGINA’.

Duality of interest

The authors declare that there is no duality of interest associated with this manuscript.

Contribution statement

DIS-A substantially contributed to conception and design, acquisition of data, and analysis and interpretation of data, drafted and wrote the article, finally approved the version to be published and is responsible for the integrity of the work as a whole. FG, CW and SU contributed substantially to conception and design, data analysis and interpretation of data and revised the article critically for important intellectual content. NS, KR, US, BS and ER contributed to data acquisition, data analysis, interpretation of data and critically revised the manuscript. CK contributed to data acquisition and critically revised the manuscript. H-ES, UAS and AK contributed to data acquisition and critically revised the manuscript. H-UH designed the study and critically revised the manuscript. All authors approved the final version of the manuscript to be published.

Author information

Author notes

  1. Ulrich A. Stock

    Present address: Department of Thoracic, Cardiac and Thoracic Vascular Surgery, University Hospital Frankfurt, Frankfurt, Germany

Authors and Affiliations

  1. Department of Internal Medicine, Division of Endocrinology, Diabetology, Angiology, Nephrology and Clinical Chemistry, University Hospital Tübingen, Otfried-Müller Str. 10, 72076, Tübingen, Germany

    Dorothea I. Siegel-Axel, Susanne Ullrich, Norbert Stefan, Kilian Rittig, Felicia Gerst, Christian Klingler, Ulrike Schmidt, Birgit Schreiner, Elko Randrianarisoa, Cora Weigert & Hans-Ulrich Häring

  2. Institute for Diabetes Research and Metabolic Diseases of the Helmholz Centre Munich at the University of Tübingen, Tübingen, Germany

    Dorothea I. Siegel-Axel, Susanne Ullrich, Norbert Stefan, Felicia Gerst, Cora Weigert & Hans-Ulrich Häring

  3. German Center for Diabetes Research (DZD), Tübingen, Germany

    Dorothea I. Siegel-Axel, Susanne Ullrich, Norbert Stefan, Felicia Gerst, Cora Weigert & Hans-Ulrich Häring

  4. Department of Plastic, Hand and Reconstructive Surgery, BG-Trauma-Center, Eberhard Karls University Tübingen, Tübingen, Germany

    Hans-Eberhard Schaller

  5. Department of Thoracic, Cardiac and Vascular Surgery, University Hospital Tübingen, Tübingen, Germany

    Ulrich A. Stock

  6. Department of General Visceral and Transplantation Surgery, University Hospital Tübingen, Tübingen, Germany

    Alfred Königsrainer

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  1. Dorothea I. Siegel-Axel
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Correspondence to Dorothea I. Siegel-Axel.

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Siegel-Axel, D.I., Ullrich, S., Stefan, N. et al. Fetuin-A influences vascular cell growth and production of proinflammatory and angiogenic proteins by human perivascular fat cells. Diabetologia 57, 1057–1066 (2014). https://doi.org/10.1007/s00125-014-3177-0

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