To the Editor:
Oxysterols are oxygenated metabolites of cholesterol. On the basis of in vitro experiments, oxysterols have been ascribed many regulatory roles in connection with inflammation, neurodegeneration and atherosclerosis1. They are present in trace amounts in biological systems, and their analysis represents a challenging analytical problem.
The 15-oxygenated cholesterol species 5α-cholest-8(14)ene-3β,15α-diol and 3β-hydroxy-5α-cholest-8(14)-en-15-one have been chemically synthesized from 7-dehydrocholesterol. Although commercially available, these have not been identified before in humans, and an enzymatic pathway for their synthesis has not been defined. Farez et al. have reported in Nature Immunology that these two sterols occur in the human circulation at concentrations considerably higher than those of any other endogenous oxysterol2. They report higher concentrations in patients with multiple sclerosis in the progressive phase. On the basis of their findings, the authors did extensive in vitro experiments in which they exposed cultured cells to the same high concentrations of 5α-cholest-8(14)ene-3β,15α-diol as they found in vivo (1,000 ng/ml). They concluded that 15-oxygenated sterols may be important factors in pathogenic mechanisms that underlie multiple sclerosis.
We represent three independent research groups with decades of experience in the analysis of oxysterols. We have been unable to confirm the findings reported by Farez et al. in terms of the presence of 15-oxygenated sterols in human plasma or serum.
One of our groups (I.B. and U.D.) has used combined gas chromatography–mass spectrometry to analyze plasma from 40 healthy Swedish volunteers without finding substantial amounts of the two 15-oxygenated sterols noted above3. In collaboration with a clinical group (T.O.), they analyzed plasma from 33 Swedish patients with multiple sclerosis at various stages of disease, again without detecting substantial concentrations of the sterols noted above. Successful recovery experiments excluded the possibility of loss of 15-oxygenated sterols during extraction and workup procedures3.
The second group (D.W.R. and J.G.M.) has used combined liquid chromatography–mass spectrometry (a modification of a published method4) to analyze more than 1,200 subjects in Dallas (USA) who represent a broad cross-section of ethnicities, social strata and health status. Using these methods, these researchers found that none of the subjects had substantial concentrations of 15-oxygenated sterols in the circulation (D.W.R. and J.G.M., data not shown).
The third group (W.J.G. and Y.W.) has used derivatization combined with liquid chromatography–high-resolution mass spectrometry and multistage fragmentation to analyze more than 40 plasma samples from healthy and sick infants and adults5. They did not find substantial concentrations of 15-oxygenated steroids in any of the plasma samples (W.J.G. and Y.W., data not shown).
We suggest that 15-oxygenated steroids should not be used as biomarkers in connection with multiple sclerosis and that a pathogenic role for these sterols must be reconsidered.
Björkhem, I. & Diczfalusy, U. Arterioscler. Thromb. Vasc. Biol. 22, 734–742 (2002).
Farez, M.F. et al. Nat. Immunol. 10, 958–964 (2009).
Björkhem, I. et al. J. Lipid Res. 52, 170–174 (2011).
McDonald, J.G., Thompson, B.T., McCrum, E.C. & Russell, D.W. Methods Enzymol. 432, 145–170 (2007).
Griffiths, W.J. et al. J. Proteome Res. 7, 3602–3612 (2008).
The authors declare no competing financial interests.
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Björkhem, I., Diczfalusy, U., Olsson, T. et al. Detecting oxysterols in the human circulation. Nat Immunol 12, 577 (2011). https://doi.org/10.1038/ni0711-577a
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