Abstract
Objective: Heme oxygenase (HO-1) plays a critical role in adipogenesis and it is important to understand its function in obesity. Many studies have shown that upregulation of HO-1 can affect the biologic parameters in obesity-mediated diabetes, hypertension and vascular endothelial cell function. Thus, we aimed to explore the hypothesis that upregulation of HO-1, using a pharmacologic approach as well as gene targeting, would improve both adiposity and endothelial cell dysfunction by direct targeting of endothelial cells. Our second aim was to compare the short-term effect of a HO-1 inducer, cobalt-protoporphrin IX (CoPP), with the long-term effects of gene targeted therapy on vascular and adipocyte stem cells in obese mice. Method: We examined the effect of CoPP on fat pre-adipocytes and mesenchymal stem cells (MSC) in mice fed a high-fat diet (HFD). We also used a lentiviral construct that expressed heme oxygenase (HO-1) that was under the control of an endothelium specific promoter, vascular endothelium cadherin (VECAD) heme oxygenase (VECAD-HO-1). We targeted endothelial cells using vascular endothelium cadherin/green fluorescent protein fusion construct (VECAD-GFP) as the control. Conditioned media (CM) from endothelial cells (EC) was added to fat derived adipocytes. Additionally, we treated renal interlobar arteries with phenylephrine and dosed cumulative increments of acetylcholine both with and without exposure to CoPP. We did the same vascular reactivity experiments with VECAD-HO-1 lentiviral construct compared to the control. Results: CoPP improved vascular reactivity and decreased adipogenesis compared to the control. MSCs exposed to CM from EC transfected with VECAD-HO-1 showed decreased adipogenesis, smaller lipid droplet size and decreased PPAR-γ, C/EBP and increased Wnt 10b compared to the control. HO-1 upregulation had a direct effect on reducing adipogenesis. This effect was blocked by tin mesoporphrin (SnMP). EC treated with VECAD-HO-1 expressed lower levels of ICAM and VCAM compared to the control, suggesting improved EC function. This also improved ACH induced vascular reactivity. These effects were also reversed by SnMP. The effect of viral transfection was much more specific and sustained than the effects of pharmacologic therapy, CoPP. Conclusion: This study demonstrates that a pharmacological inducer of HO-1 such as CoPP improves endothelial cell function while dampening adipogenesis, but long-term HO-1 expression by direct targeting of endothelial cells by gene transfer therapy may offer a more specific and ideal solution. This was evidenced by smaller healthier adipocytes that had improved insulin sensitivity, suggesting increased adiponectin levels. HO-1 upregulation reestablished the “crosstalk” between perivascular adipose tissue and the vascular system that was lost in the chronic inflammatory state of obesity. This study demonstrates that gene targeting of EC may well be the future direction in treating obesity induced EC dysfunction, with the finding that targeting the vasculature had a direct and sustained effect on adipogenesis.
Highlights
Obesity remains a crisis of epidemic proportions in this country with a third of our population classified as overweight and another third as obese [1]
We have shown that obese females with metabolic syndrome and obesity is associated with inflammation that leads to increased levels of reactive oxygen species (ROS)
We have shown that oxidized HDL (Ox-HDL) increases adipogenesis in mesenchymal stem cells (MSC) derived adipocytes, which was further increased with the addition of either 20-HETE or Ang II
Summary
Obesity remains a crisis of epidemic proportions in this country with a third of our population classified as overweight and another third as obese [1]. We have shown that obese females with metabolic syndrome and obesity is associated with inflammation that leads to increased levels of reactive oxygen species (ROS) This oxidant attack results in increased levels of oxidized HDL (Ox-HDL) and isoprostane [5]. CoPP is a potent inducer of HO-1 upregulation, which decreases ROS, increases mitochondrial function and has been shown to reduce body weight [9,10] It is not specific or long lasting in effect. The “crosstalk” between adipose tissue and endothelial cell function is lost in the chronic inflammatory state of obesity We determined if this is restored by HO-1 upregulation by measuring vascular reactivity [20,21,22]. It includes parameters We evaluated the mechanism by testing CM from EC transfected by HO-1-VECAD and its effect on adipogenesis
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