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In General


Cell membranes are phospholipid-bilayers. Apolar fatty acids face each other, and polar groups are on the outer surface. The composition of the cell membrane impacts on cell function, and even on interaction between cells. Before the introduction of the HS-Omega-3 Index, there was no standardized fatty acid analysis. Each fatty acid laboratory analyzed fatty acids with its own method and in some fatty acid compartment: whole blood, plasma, serum, plasma phospholipids, plasma triglycerides, plasma cholesterolesters, erythrocytes, platelets, leukocytes, samples from all sorts of tissue, and many more. Each laboratory used and still uses often its own method, hardly ever tested for reproducibility, analytical variability and other features of analytical quality. This approach made sure that results of studies from different groups were, and frequently still are, not comparable. This also made sure that fatty acid analyses would never become a part of clinical routine.

In 2002, WS Harris and C von Schacky jointly had the idea to standardize fatty acid analysis of one compartment. One of the motives was generating uniform data, thus making it possible for fatty acid analysis to become part of clinical routine. Based on previously generated data, WS Harris and C von Schacky quickly agreed on erythrocytes as viable compartment, since the fatty acid composition of erythrocytes had already demonstrated slow incorporation and excorporation kinetics, making a low biological variability a distinct possibility (von Schacky et al, 1985). Moreover, it had already been demonstrated in a randomized controlled trial that an increase of omega-3 fatty acids in erythrocytes mitigated the “natural” history of coronary atherosclerosis (von Schacky et al, 1999). Shaping the idea, building up the standardized analytical methodology, and first experiments took another two years. In 2004, the Omega-3 Index was defined (eicosapentaenoic acid, EPA, plus docosahexaenoic acid, DHA, in erythrocytes), and suggested as a new risk factor for death from coronary artery disease (Harris & von Schacky, 2004).

Initial publications demonstrated that EPA and DHA in erythrocytes was representative for EPA and DHA in cardiac tissue, which was found to be true also for other tissues in the experimental animal (Harris et al, 2004, Arnold et al, 2010). Soon, the low biologic variability of fatty acids in erythrocytes and the low analytical variability of the analytical method were formally demonstrated (Harris & Thomas, 2010). Early review articles put the Omega-3 Index close to clinical routine (Harris, 2007, von Schacky, 2007), with the consequence that various laboratories call their respective fatty acid analysis “Omega-3 Index” as well, even if serum or plasma were (and still are) analyzed. In order to minimize confusion, we had to rename the Omega-3 Index “HS-Omega-3 Index”, and trademark it in many countries, so everybody can recognize the standard. “HS-Omega-3 Index” is the unique name for a strictly standardized analytical method of erythrocyte fatty acid composition.

The availability of a standardized method for fatty acid analyses paved the way for innumerable co-operations with premier groups (e.g. Framingham or KORA), universities (e.g. Harvard or Charité), companies or other organizations (e.g. Cooper Clinic or the German Army). More than 150 publications, based on the HS-Omega-3 Index, originated from these co-operations (see list of publications). More than 50 research projects are currently ongoing. Current projects investigate topics like cardiovascular disease, congestive heart failure, cognitive capabilities, bioavailability, and many more. The results of this research are discussed in the respective chapters of this website, and also in current review articles (e.g. von Schacky, 2014).

Pre-analytics

The HS-Omega-3 Index uses erythrocytes, separated from plasma by centrifugation of EDTA-blood. Citrate-blood is possible, too. A separate application is a kit for a self-test. Substantial test series have shown that the fatty acid composition of erythrocytes in EDTA-blood is stable at ambient temperature for seven days, making it possible to ship samples by regular mail in most cases. Already after a few days of storage at -20°C, the fatty acid composition of erythrocytes can no longer be measured. If stored at -80°C, samples are stable for decades. Please contact us for more information.

Quality assurance

At Omegametrix, each analysis is subject to checks for plausibility, constancy, asf. As a rule, Omegametrix applies all measures of quality assurance customary for clinical chemistry laboratories. This was put on a formal level by Omegametrix working along DIN ISO 15 189, as typical for clinical chemistry laboratories; the accreditation has been applied for. Moreover, on a regular basis, Omegametrix performs proficiency tests with both sister laboratories, Omegaquant in the USA, and Omegaquant in South-Korea. These proficiency tests cannot be performed by other laboratories. The HS-Omega-3 Index is subject to current measures of quality assurance, meaning that the HS-Omega-3 Index is the only method for fatty acid analysis that can be used in clinical routine.

Safety and tolerability

The HS-Omega-3 Index is determined in erythrocytes from a small amount of EDTA-blood, as obtained from venipuncture. The remains of a blood count can be used. Determining the HS-Omega-3 Index is as safe as a blood count.

Tolerability of Omega-3 fatty acids was comparable to placebo in large intervention trials with clinical endpoints (e.g. Gissi-Investigators, 2008, ORIGIN, 2012). In these trials, clinically relevant events (like bleeding), were as frequent under Verum, as under Placebo (von Schacky, 2014). The world’s most important regulatory authorities also think that EPA and DHA are safe: The European Food Safety Authority (EFSA) thinks that up to 5 g / day are safe, whereas the American Food and Drug Adminstration (FDA) thinks that up to 3 g / day are safe (EFSA 2012, FDA).

According to data based on the HS-Omega-3 Index, approximately 80% of the populations in Korea and in Japan have an HS-Omega-3 Index above 8% (Harris et al, 2013). According to cardiovascular data based on the HS-Omega-3 Index, no further benefit can be detected at levels above 11% (Harris et al, 2013). A similar picture emerges for complex brain functions. Therefore, a target range for the HS-Omega-3 Index between 8 and 11% has been suggested.

This target range might be too low for chronic-inflammatory diseases, but this remains to be substantiated. According to our anecdotal experience, patients with chronic-inflammatory diseases aim for a HS-Omega-3 Index around 15%. These patients did not report an increased bleeding tendency, nor did individuals with a HS-Omega-3 Index >16%, whom we contacted by phone. Apparently, such a high HS-Omega-3 Index does not seem to be associated with a bleeding tendency either, more formal data need to be obtained. However, it has been formally investigated that during treatment of acute myocardial infarctions, which calls for a number of drugs maximizing a bleeding tendency, no bleeding tendency can be found in correlation with the HS-Omega-3 Index (Salisbury, 2012).

Mammals seem to have a mechanism prohibiting a HS-Omega-3 Index above 20%. In dolphins, consuming approximately 100 times as much EPA and DHA as humans on a Western diet, the mean HS-Omega-3 Index was 19.9+1.42% (Harris & Schmitt, 2014). In none of the populations investigated so far, a HS-Omega-3 Index way above 20% was observed. Thus, there seems to be a metabolic safeguard against too much EPA and DHA.

Study Design in the Past and in the Future

For intervention trials with omega-3 fatty acids, participants used to be recruited irrespective of their baseline omega-3 fatty acids. In every population investigated so far, the HS-Omega-3 Index had a statistically normal distribution. Therefore, for intervention trials, individuals with high baseline levels were among the ones recruited. No effect on an intervention with omega-3 fatty acids is can be expected in these study participants, if randomized to verum, while no events can be expected, if randomized to placebo.

In most intervention trials with cardiovascular endpoints, participants were recommended to ingest their fish oil capsules (or placebo) with breakfast (von Schacky, 2014). In many countries, breakfast is a low-fat meal. Uptake of fat depends on fat digestion with activation of pancreas and bile. This does not occur in response to small quantities of fat, e.g. 1 g in a capsule. In the intervention trials mentioned, EPA plus DHA were used as ethyl-ester or triglyceride in a capsule (von Schacky, 2014). If EPA plus DHA are ingested as ethyl-ester with a low-fat meal, bioavailability is substantially worse than if ingested with a high-fat meal; in one study by a factor of 13 (Davidson et al, 2012). Due to the mechanisms involved, this is probably also true for EPA plus DHA as a triglyceride. To improve bioavailability, EPA plus DHA ethyl-ester could have been emulsified, which improved bioavailability up to a factor of 21 in one study (Hussey et al, 2012). Another possibility would be to ask trial participants to take their capsules with their main meal. Taken together, unintentionally, timepoint and form (capsule) of EPA plus DHA were chosen in a way minimizing bioavailability.

To make the topic bioavailability even more complicated, it turned out that there are large inter-individual differences in bioavailability of EPA plus DHA. We found a 13-fold inter-individual difference in absorption in a 8-week trial with 0.5 g / day EPA plus DHA in an emulsion (Köhler et al, 2010). Similar data have been reported from other trials (e.g. Flock et al, 2013). In a recent trial, a single dose of EPA plus DHA was given, and taken under observation – with identical results (Köhler et al, 2015). The latter rules out issues of compliance. Bioavailability of fat has turned into a research topic in its own right.

The large inter-individual variability in bioavailability, together with recruiting trials participants irrespective of baseline levels will lead to small mean differences in levels of EPA plus DHA between Verum and Placebo/Control groups during a trial, and to substantial overlap between the two groups (von Schacky, 2014). Previously, this has not been looked at frequently. However, if investigated, a small difference in mean levels, and a large overlap between Verum and Placebo groups has been found (Muhlhausler et al, 2014). If there is only a small difference between Verum and Placebo groups with respect to the study intervention, it will become almost impossible to detect an effect of the study intervention, even if the study’s intervention is effective.

Ignoring issues of bioavailability and not measuring levels at baseline and during the trials, has led to many neutral results of intervention trials, especially in the cardiovascular field.

Nevertheless, some trials, especially in the field of neurology and psychiatry, had positive results. How can this be explained?

As a rule, we find a low HS-Omega-3 Index in populations, in which trials with the study design just discussed had positive results. According to our data, the disease investigated, like depression, ADHD, cognitive impairment, or congestive heart failure, comes with a low HS-Omega-3 Index (von Schacky, 2014). Thus, the disease selects for a homogeneous trial population, making it easier to detect an effect of increased intake of EPA plus DHA. If the discussed issues were to be incorporated into future trial designs, larger effects are liked to be detected in the future.

Literatur


Arnold C, Markovic M, Blossey K, Wallukat G, Fischer R, Dechend R, Konkel A, von Schacky C, Luft FC, Muller DN, Rothe M, Schunck WH. Arachidonic acid-metabolizing cytochrome P-450 enzymes are targets of omega-3 fatty acids. J Biol Chem, 2010;285:32720-33
Baghai TC, Varallo-Bedarida G, Born C, Häfner S, Schüle C, Eser D, Rupprecht R, Bondy B, von Schacky C. Major depression is associated with cardiovascular risk factors and low Omega-3 Index. J Clin Psychiat 2011;72:1242-7
Davidson MH, Johnson J, Rooney MW, Kyle ML, Kling DF. A novel omega-3 free fatty acid formulation has dramatically improved bioavailability during a low-fat diet compared with omega-3-acid ethyl esters: the ECLIPSE (Epanova(®) compared to Lovaza(®) in a pharmacokinetic single-dose evaluation) study. J. Clin. Lipidol 2012;6:573-84.
EFSA Journal 2012;10(7):2815
FDA Docket No. 91N-0103
Gissi-HF Investigators, L. Tavazzi, A.P. Maggioni, R. Marchioli, S. Barlera, M.G. Franzosi, R. Latini et al. Effect of n-3 polyunsaturated fatty acids in patients with chronic heart failure (the GISSI-HF trial): a randomised, double-blind, placebo-controlled trial. Lancet 2008;372:1223-1230
Harris WS. Omega-3 Fatty Acids and Cardiovascular Disease. A case for the Omega-3 Index as a New Risk Factor. Pharmacol Res 2007;55:217-23
Harris WS and von Schacky C. The Omega-3 Index: A New Risk Factor for Death from CHD? Preventive Medicine 2004;39:212-20.
Harris WS, Sands SA, Windsor SL, Ali HA, Stevens TL, Magalski A, Porter CB, Borkon AM. Omega-3 Fatty Acid Levels in Transplanted Human Hearts: Effect of Supplementation and Comparison with Erythrocytes. Circulation 2004;110;1645-9.
Harris WS, Thomas RM. Biological variability of blood omega-3 biomarkers. Clin Biochem 2010;43:338-40
Harris WS, von Schacky C, Park Y. Standardizing Methods for Assessing Omega-3 Fatty Acid Biostatus. In The Omega-3 Fatty Acid Deficiency Syndrome; McNamara RK ed., Nova Science Publishers 2013
Harris WS, Schmitt TL. Unexpected similarity in RBC DHA and AA levels between bottlenose dolphins and humans. Prostaglandins Leukot Essent Fatty Acids. 2014;90:55-9
Hussey EK, Portelli S, Fossler MJ, Gao F, Harris WS, Blum RA, Lates CD, Gould E, Abu-Baker O, Johnson S, Reddy KK. Relative bioavailability of an Emulsion Formulation for Omega-3-Acid Ethyl Esters Compared to the Commercially Available Formulation : A Randomized, Parallel-Group, Single-Dose Study Followed by Repeat dosing in healthy volunteers. Clin Pharm Drug Develop 2012;1:14-23
Flock MR, Skulas-Ray AC, Harris WS, Etherton TD, Fleming JA, Kris-Etherton PM. Determinants of Erythrocyte Omega-3 Fatty Acid Content in Response to Fish Oil Supplementation: A Dose-Response Randomized Controlled Trial. J Am Heart Assoc. 2013 Nov 19;2(6):e000513.
Köhler A, Bittner D, Löw A, von Schacky C. Effects of a convenience drink fortified with n-3 fatty acids on the n-3 index. Br J Nutr 2010; 104:729-36.
Muhlhausler BS, Gibson RA, Yelland LN, Makrides M. Heterogeneity in cord blood DHA concentration: towards an explanation. Prostaglandins Leukot Essent Fatty Acids. 2014;91:135-40
ORIGIN Trial Investigators, J. Bosch J, H.C. Gerstein, G.R. Dagenais, R. Díaz R, L. Dyal, H. Jung, et al. N-3 fatty acids and cardiovascular outcomes in patients with dysglycemia. N. Engl. J. Med. 2011;367:319-328
Salisbury AC, Harris WS, Amin AP, Reid KJ, O'Keefe Jr JH, Spertus JA. Relation Between Red Blood Cell Omega-3 Fatty Acid Index and Bleeding During Acute Myocardial Infarction. Am J Cardiol. 2012;109:13-8
von Schacky C. Omega-3 Fettsäuren in der Kardiologie – neueste Entwicklungen. Münch Med Wochenschr 2007;149:97-101
von Schacky C. Der HS-Omega 3 Index®: klinische Wertigkeit standardisierter Fettsäureanalytik. J Lab Med 2014;38:167-78
von Schacky C. Omega-3 Fatty Acids in Cardiovascular Disease - an Uphill Battle. PLEFA 2015;92:41-7
von Schacky C, Angerer P, Kothny W, Theisen K, Mudra H. A randomized, double-blind, placebo-controlled trial. Ann Intern Med. 1999;130:554-62.
von Schacky C, Fischer S, Weber PC. Long-term effects of dietary marine omega-3 fatty acids upon plasma and cellular lipids, platelet function, and eicosanoid formation in humans. J Clin Invest. 1985;76:1626-31.
Widenhorn-Müller K, Schwanda S, Scholz E, Spitzer M, Bode H. Effect of supplementation with long-chain ω-3 polyunsaturated fatty acids on behavior and cognition in children with attention deficit/hyperactivity disorder (ADHD): A randomized placebo-controlled intervention trial. Prostaglandins Leukot Essent Fatty Acids. 2014;91:49-60