OBJECTIVE:Obesity is associated with adipose tissue hypoxia, and is thought to be linked to the chronic low-grade inflammation of adipose tissue, although the precise mechanism has remained unclear. In this study, we investigated the effect of a prominent hypoxia on human primary adipocyte secretion and tumor necrosis factor alpha (TNFα)-induced nuclear factor-κB (NF-κB) signaling. RESULTS:Using cytokine array and ELISA analysis, we compared the secretion patterns of normoxic and hypoxic (1% O(2)) adipocytes and observed various alterations in adipokine release. We could reproduce known alterations like an induction of interleukin (IL)-6, vascular endothelial growth factor, leptin and a reduction in adiponectin release under hypoxia. Interestingly, we observed a significant reduction in the secretion of macrophage chemotactic protein (MCP)-1 and other NF-κB-related genes, such as growth-regulated oncogene-α, eotaxin and soluble TNF-Receptor1 (TNF-R1) under hypoxia. TNFα stimulation of hypoxic adipocytes resulted in a significantly reduced phosphorylation of NF-κB and its inhibitor IκBα compared with normoxic cells. Furthermore, chronic treatment of hypoxic adipocytes with TNFα resulted in an expected higher secretion of the chemokines MCP-1 and IL-8, but under hypoxia, the secretion level was substantially lower than that under normoxia. This reduction in protein release was accompanied by a reduced mRNA expression of MCP-1, whereas IL-8 mRNA expression was not altered. Additionally, we observed a significantly reduced expression of the TNF-receptor TNF-R1, possibly being one cause for the reduced responsiveness of hypoxic adipocytes towards TNFα stimulation. CONCLUSION:In conclusion, human primary adipocytes show a basal and TNFα-induced reduction of MCP-1 release under hypoxia. This effect may be due to a reduced expression of TNF-R1 and therefore attenuated TNFα-induced NF-κB signaling. These observations demonstrate a reduced responsiveness of hypoxic adipocytes towards inflammatory stimuli like TNFα, which may represent an adaptation process to maintain adipose tissue function under hypoxia and inflammatory conditions.

Fragestellung: PEDF gehört zur Familie der nicht-inhibierenden Serpine und ist ein multi-funktionales Protein mit neurotrophen und anti-angiogenen Eigenschaften. Seit kurzem ist bekannt, dass Typ 2 Diabetiker erhöhte PEDF Konzentrationen im Serum aufweisen. In Mäusen konnte gezeigt werden, dass die PEDF Expression im Fettgewebe positiv mit Adipositas und Insulinresistenz korreliert. Durch die Sekretionsanalyse von in vitro differenzierten humanen Adipozyten mittels 2D-PAGE und MALDI-MS konnten wir zeigen, dass PEDF eines der abundantesten Proteine im Sekretom der humanen Fettzelle ist. Ziel dieser Studie war es, die Regulation und autokrine Funktion von PEDF in humanen Adipozyten zu untersuchen und die parakrinen Effekte auf humane Skelettmuskelzellen (hSkMC) und glatte Muskelzellen (hSMC) zu bestimmen.

Objective: Pigment epithelium-derived factor (PEDF) is a multifunctional protein with neurotrophic and anti-angiogenic properties. More recently it became evident that PEDF is upregulated in patients with type 2 diabetes and also contributes to insulin resistance in mice. During characterization of the secretome of in vitro differentiated human adipocytes by two-dimensional polyacrylamide gel electrophoresis and matrix-assisted laser desorption/ionization-MS, we found that PEDF is one of the most abundant proteins released by adipocytes. The aim of this study was to investigate the regulation and autocrine function of PEDF in human adipocytes and to determine its paracrine effects on human skeletal muscle cells (hSkMC) and human smooth muscle cells (hSMC). Methods and results: Human primary adipocytes secrete 130 ng ml −1 PEDF over 24 h from 1 million cells, which is extremely high as compared with adiponectin, interleukin-6 (IL-6) or IL-8. This release of PEDF is significantly higher than from other primary cells, such as adipose-tissue located macrophages (50-times), hSkMC and hSMC (5-times). PEDF protein expression significantly increases during adipogenesis, which is paralleled by increased PEDF secretion. Furthermore, tumor necrosis factor-α and hypoxia significantly downregulate PEDF protein levels. PEDF secretion was significantly reduced by troglitazone and hypoxia and significantly increased by insulin. Treatment of adipocytes and hSkMC with PEDF induced insulin resistance in adipocytes, skeletal and smooth muscle cells at the level of insulin-stimulated Akt phosphorylation, which was dose dependent and more prominent in adipocytes. Furthermore, inflammatory nuclear factor-κB (NF-κB) signaling was induced by PEDF. In hSMC, PEDF induced proliferation (1.7-fold) and acutely activated proliferative and inflammatory signaling pathways (NF-κB, p38 mitogen-activated protein kinase and mammalian target of rapamycin). Conclusion: PEDF is one of the most abundant adipokines and its secretion is inversely regulated by insulin and hypoxia. PEDF induces insulin resistance in adipocytes and hSkMC and leads to inflammatory signaling in hSMC. Because of these diverse actions, PEDF is a key adipokine, which could have an important role in diabetes and obesity-related disorders.

Fragestellung: Das Auftreten von Hypoxie in expandierendem Fettgewebe als Folge einer Adipositas ist ein Phänomen, das sowohl im humanen Fettgewebe als auch im Tiermodell nachgewiesen werden konnte. Eine Hypoxie im Fettgewebe führt zu einer Aktivierung verschiedener Signalwege und einem veränderten Sekretionsprofil der Adipozyten. Ziel dieser Studie war es, den Einfluss eines verminderten Sauerstoffangebotes auf den pro-inflammatorischen NF-κB Signalweg und die hieraus resultierenden Folgen auf der Ebene sekretorischer Zielgene von NF-κB zu untersuchen.

Fragestellung: Adipositas stellt einen großen Risikofaktor für die Entwicklung eines Typ 2 Diabetes dar. Dabei ist das Fettgewebe sekretorisch aktiv und sezerniert bei zunehmender Masse vermehrt pro-inflammatorische Adipokine. Ein erst kürzlich als Adipokin beschriebenes Sekretionsprodukt der Fettzelle ist Chemerin. Dieses Chemokin ist essentiell für die Adipogenese und beeinflusst die Insulinwirkung in Adipozyten in vitro. In dieser Studie soll geklärt werden, wie die Sekretion von Chemerin im Fettgewebe reguliert wird und ob Chemerin in Skelettmuskelzellen Insulinresistenz induziert und somit einen Link zum Diabetes darstellen könnte.

Cannabinoid type 1 receptor (CB1R) antagonists such as rimonabant (Rim) represent a novel approach to treat obesity and related metabolic disorders. Recent data suggest that endocannabinoids are also produced by human adipocytes. Here we studied the potential involvement of endocannabinoids in the negative crosstalk between fat and muscle.

Fragestellung: Cannabinoid-Typ1-Rezeptoren (CB1R) werden sowohl im zentralen und periphären Nervengewebe als auch in verschiedenen periphären Zelltypen wie Adipozyten und Leberzellen exprimiert. Der Einsatz von spezifischen CB1R-Antagonisten wie Rimonabant ist ein innovativer Therapieansatz in der Behandlung von Adipositas, da hierdurch eine Reduktion der Nahrungsaufnahme bewirkt wird. Zusätzlich konnte in verschiedenen Studien gezeigt werden, dass auch Effekte in periphären Geweben an der positiven Gesamtwirkung von Rimonabant beteiligt sind. Aktuelle Daten lassen darauf schließen, dass im humanen Fettgewebe Endocannabinoide produziert werden. Wir haben in unserer Studie untersucht, ob das Endocannabinoidsystem am negativen Crosstalk zwischen Fettgewebe und Skelettmuskulatur beteiligt ist.