7-Oxysterols constitute the major component (40%) of oxidized low-density lipoprotein (oxLDL). They arise in the body via auto-oxidation of cholesterol and are known to induce endothelial dysfunction, oxidative stress and apoptosis in the vascular wall, prior to development of atherosclerosis. A novel pathway has been described for hepatic inter-conversion of 7-ketocholesterol (7-KC) and 7β -hydroxycholesterol (7β OHC) by the enzyme 11β-hydroxysteroid dehydrogenase type-1 (11β HSD1), better known for metabolizing glucocorticoids. Inhibition of 11βHSD1 is atheroprotective and the potential underlying mechanism for this may involve altered metabolism and actions of glucocorticoids. However, alterations in the metabolism of 7-oxysterols may also play an important role in this atheroprotective effect. The work described here addresses the hypotheses that (i) 7-oxysterols are substrates for murine 11βHSD1; (ii) inhibition of 11β HSD1 may abolish cellular metabolism of 7-oxysterols; (iii) this route of metabolism may modulate the actions of 7-oxysterols and glucocorticoids on murine vascular physiology. Murine 11β HSD1 inter-converted 7-oxysterols (Km=327.6±98ìM, Vmax=0.01±0.001pmol/ìg/min) but the regulation of reaction direction is different from that for glucocorticoids. Predominant dehydrogenation of 7β OHC to 7-KC was quantified in several models (recombinant protein, cultured cells stably transfected with 11β HSD1), in which predominant reduction of glucocorticoids was measured. Furthermore, in murine hepatic microsomes, dehydrogenation of 7β OHC occurred exclusively. In aortic rings in culture, however, both reduction and dehydrogenation of 7-oxysterols were evident. 7-Oxysterols and glucocorticoid substrates competed for metabolism by 11β HSD1, with 7β OHC inhibiting dehydrogenation of glucocorticoids (Ki=908±53nM). The circulating concentrations of 7-oxysterols in the plasma of C57Bl6 and 11β HSD1-/- mice were in the ìM range (0.02 – 0.13ìM). The disruption of 11β HSD1 has resulted in increased ratios of 7-KC and 7β OHC over total plasma cholesterol levels (*p<0.05). This finding suggested that 11β HSD1 is involved in metabolizing and determining the plasma levels of 7-KC and 7β OHC. To assess the consequences of these alterations for vascular function, studies were undertaken in aortic rings. Prolonged incubation with 7-oxysterols (20-25 ìM) showed a tendency to attenuate noradrenaline-mediated contractions of C57Bl6 aortae, but had no effect on contractions in response to 5-hydroxytryptamine or KCl. Similarly, endothelium-dependent and -independent relaxations of murine aortae were unaltered after exposure to 7-oxysterols. Thus in the mouse, 11β HSD1 may influence the balance of circulating and cellular 7-oxysterols which may have consequential effects on glucocorticoid action. Although this work suggests that concentrations present in murine tissues are unlikely to cause vascular dysfunction, they may influence further cellular events as yet undescribed. Under pathological conditions where high concentrations of 7-oxysterols occur, 11β HSD1 may influence the extracellular-transport and delivery of 7-KC and 7β OHC to the plaque. This work therefore proposes that inhibition of metabolism of 7-oxysterols by 11β HSD1 inhibitors, may contribute to the atheroprotective effects of these drugs.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:562893 |
| Date | January 2010 |
| Creators | Mitić, Tijana |
| Contributors | Andrew, Ruth. : Hadoke, Patrick W. F. : Walker, Brian R. |
| Publisher | University of Edinburgh |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/1842/4416 |
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