AS271 - Omega-3 and Omega-6 Fatty Acids as a Biomarker of Hip Fracture Risk

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Investigator Names and Contact Information

Tonya Orchard, MS, RC/LD;



Osteoporosis is a major public health problem, especially in women. Each year in the United States, there are 1.5 million fractures attributable to osteoporosis including 329,000 hip fractures, one of the most disabling consequences of aging in women (1). Estimates in European white women suggest that the one in six lifetime risk for hip fracture exceeds the one in nine lifetime risk for breast cancer (2). Worldwide, older adults are the fastest growing segment of the population; even if age-adjusted hip fracture rates remain stable, the graying of society will result in an estimated worldwide rise in incidence of hip fractures from 1.7 million in 1990 to 6.3 million in 2050 (3). In response to this growing epidemic, The Healthy People 2010 (Healthy People 2010: objectives established a national target to reduce the proportion of adults who have osteoporosis and the risk for fracture during the current decade.
Though there has been much research on the benefits of n-3 fatty acids in various diseases, there is limited research addressing the relationship of these long chain polyunsaturated fatty acids (PUFAs) to fracture risk in osteoporosis. The n-6 and n-3 PUFAs seem to have specific, and often opposing effects on biomarkers of bone turnover in animal (4-6) and cell studies (7, 8); with n-6 fatty acids associated with more negative effects and n-3 fatty acids associated with more positive effects on bone. In humans, higher consumption of n-3 fatty acids and lower n-6/n-3 ratios have been associated with higher bone mineral density (BMD) (9-11) and biomarkers favoring bone formation (12, 13). Data from the Nurses’ Health Study (14)  and the Health Professionals Follow-Up Study (15) suggest an inverse relationship between dietary intake of n-3 fatty acids and inflammatory cytokines important in regulation of bone metabolism. However, in research conducted in Spain with dietary intakes of n-3 fatty acids more than 10 times higher than typical U.S. intakes, no significant relationship between self-reported n-3 fatty acids and fractures was found, but an elevated fracture risk was reported for individuals consuming higher n-6 fatty acids (16).
Current epidemiological research has focused on the use of food frequencies to evaluate the association of long chain PUFAs to bone health (16-22). Food frequencies give imperfect estimates of food intake (18). Analysis of red blood cell (RBC) fatty acids is one approach that may increase the accuracy of assessing fat intake and provide important information on target levels to achieve therapeutic benefit. With the exception of one small study in young males (10), there is limited research regarding blood levels of n-3 fatty acids in relation to biomarkers of bone health. To our knowledge, there are no published reports on blood levels of n-3 fatty acids and fracture risk in humans.
We propose to measure red blood cell levels of n-3 and n-6 PUFAs in postmenopausal women with hip fractures in the Women’s Health Initiative (WHI) using a nested case-control design.  The large sample size and breadth of diversity of the WHI women offer the unique opportunity to evaluate BMD data as well as hip fracture risk and test for interactions of long chain PUFA intake and known clinical risk factors (23) such as physical activity, health status, height, weight, fracture history, smoking status and diabetes to further predict risk of hip fracture. The proposed study will be the first to quantify red blood cell concentrations of long chain PUFAs in relation to hip fracture in humans.
Primary Hypothesis: Women with higher red blood cell levels of n-3 fatty acids and lower red blood cell n-6/n-3 ratios will have fewer hip fractures.
Secondary Hypothesis: Women with higher red blood cell n-3 fatty acids and lower red blood cell n-6/n-3 fatty acid ratios will have higher bone mineral density.
In order to test our hypotheses, we plan to pursue the following specific aims:
Specific Aim 1: Investigate the relationship between red blood cell levels of n-3 and n-6 fatty acids in women with hip fractures versus matched controls in the WHI.
Specific Aim 2: Investigate the association of longitudinal measures of BMD in women with hip fractures and matched controls in the WHI across varying levels of red blood cell n-3 and n-6 fatty acids. 
Increasing blood levels of n-3 fatty acids offers an especially attractive intervention for osteoporosis because it is relatively inexpensive and is associated with minimal risks. Fatty fish intake of as little as two 4 oz. servings per week or the equivalent fish oil capsule supplementation of 485 mg eicosapentanoic acid (EPA) + docosahexanoic acid (DHA) per day has been shown to increase blood levels of n-3 fatty acids to ~5.5 – 6.8 % of  total fatty acids (24), and confer a cardiovascular benefit (25). A solid foundation of evidence needs to be developed to clarify the relationship of long chain PUFA levels to bone health as well. This research has the potential to address this important question in the group most highly vulnerable to osteoporotic fractures, postmenopausal women.



Some of the publications related to this ancillary study are:

Ms1292 - Orchard TS, Ing SW, Lu B, Belury MA, Johnson K, Wactawski-Wende J, Jackson RD. The association of red blood cell n-3 and n-6 fatty acids to dietary fatty acid intake, bone mineral density and hip fracture risk in the Women's Health Initiative. Miner Res. 2012 Sep 27. [Epub ahead of print]

For a complete, up-to-date list of WHI papers related to this ancillary study, please use the searchable Bibliography section of this website. To search for papers by study number, access the Simple Search, and enter the study number in the “Related Studies” field. 


1.   Braithwaite RS, Col NF, Wong JB. Estimating hip fracture morbidity, mortality and costs. Journal of the American Geriatrics Society. 2003 Mar;51:364-70.
2.   van Staa TP, Dennison EM, Leufkens HG, Cooper C. Epidemiology of fractures in England and Wales. Bone. 2001 Dec;29:517-22.
3.   Sambrook P, Cooper C. Osteoporosis. Lancet. 2006 Jun 17;367:2010-8.
4.   Watkins BA, Li Y, Seifert MF. Dietary ratio of n-6/n-3 PUFAs and docosahexaenoic acid: actions on bone mineral and serum biomarkers in ovariectomized rats. The Journal of nutritional biochemistry. 2006 Apr;17:282-9.
5.   Matsushita H, Barrios JA, Shea JE, Miller SC. Dietary fish oil results in a greater bone mass and bone formation indices in aged ovariectomized rats. Journal of Bone & Mineral Metabolism*. 2008;26:241.
6.   Shen CL, Yeh JK, Rasty J, Li Y, Watkins BA. Protective effect of dietary long-chain n-3 polyunsaturated fatty acids on bone loss in gonad-intact middle-aged male rats. Br J Nutr. 2006 Mar;95:462-8.
7.   Coetzee M, Haag M, Kruger MC. Effects of arachidonic acid, docosahexaenoic acid, prostaglandin E(2) and parathyroid hormone on osteoprotegerin and RANKL secretion by MC3T3-E1 osteoblast-like cells. The Journal of nutritional biochemistry. 2007 Jan;18:54-63.
8.   Rahman MM, Bhattacharya A, Fernandes G. Docosahexaenoic acid is more potent inhibitor of osteoclast differentiation in RAW 264.7 cells than eicosapentaenoic acid. Journal of cellular physiology. 2008 Jan;214:201-9.
9.   Weiss LA, Barrett-Connor E, von Muhlen D. Ratio of n-6 to n-3 fatty acids and bone mineral density in older adults: the Rancho Bernardo Study. Am J Clin Nutr. 2005 April 1, 2005;81:934-8.
10.  Hogstrom M, Nordstrom P, Nordstrom A. n-3 Fatty acids are positively associated with peak bone mineral density and bone accrual in healthy men: the NO2 Study. Am J Clin Nutr. 2007 March 1, 2007;85:803-7.
11.  Kruger MC, Coetzer H, de Winter R, Gericke G, van Papendorp DH. Calcium, gamma-linolenic acid and eicosapentaenoic acid supplementation in senile osteoporosis. Aging (Milano). 1998 Oct;10:385-94.
12.  Griel AE, Kris-Etherton PM, Hilpert KF, Zhao G, West SG, Corwin RL. An increase in dietary n-3 fatty acids decreases a marker of bone resorption in humans. Nutr J. 2007;6:2.
13.  Terano T. Effect of omega 3 polyunsaturated fatty acid ingestion on bone metabolism and osteoporosis. World Rev Nutr Diet. 2001;88:141-7.
14.  Lopez-Garcia E, Schulze MB, Manson JE, Meigs JB, Albert CM, Rifai N, Willett WC, Hu FB. Consumption of (n-3) Fatty Acids Is Related to Plasma Biomarkers of Inflammation and Endothelial Activation in Women. J Nutr. 2004 July 1, 2004;134:1806-11.
15.  Pischon T, Hankinson SE, Hotamisligil GS, Rifai N, Willett WC, Rimm EB. Habitual Dietary Intake of n-3 and n-6 Fatty Acids in Relation to Inflammatory Markers Among US Men and Women. Circulation. 2003 July 15, 2003;108:155-60.
16.  Martinez-Ramirez MJ, Palma S, Martinez-Gonzalez MA, Delgado-Martinez AD, de la Fuente C, Delgado-Rodriguez M. Dietary fat intake and the risk of osteoporotic fractures in the elderly. Eur J Clin Nutr. 2007;61:1114-20.
17.  Michels KB, Bingham SA, Luben R, Welch AA, Day NE. The effect of correlated measurement error in multivariate models of diet. Am J Epidemiol. 2004 Jul 1;160:59-67.
18.  Byers T. Food frequency dietary assessment: how bad is good enough? Am J Epidemiol. 2001 Dec 15;154:1087-8.
19.  Hebert JR, Patterson RE, Gorfine M, Ebbeling CB, St Jeor ST, Chlebowski RT. Differences between estimated caloric requirements and self-reported caloric intake in the women's health initiative. Ann Epidemiol. 2003 Oct;13:629-37.
20.  Neuhouser ML, Kristal AR, McLerran D, Patterson RE, Atkinson J. Validity of short food frequency questionnaires used in cancer chemoprevention trials: results from the Prostate Cancer Prevention Trial. Cancer Epidemiology, Biomarkers and Prevention. 1999;8:721-5.
21.  Kaaks R, Ferrari P. Dietary intake assessments in epidemiology: can we know what we are measuring? Ann Epidemiol. 2006 May;16:377-80.
22.  Block G. Human dietary assessment: methods and issues. Prev Med. 1989 Sep;18:653-60.
23.  Robbins J, Aragaki AK, Kooperberg C, Watts N, Wactawski-Wende J, Jackson RD, LeBoff MS, Lewis CE, Chen Z, et al. Factors Associated With 5-Year Risk of Hip Fracture in Postmenopausal Women. JAMA. 2007 November 28, 2007;298:2389-98.
24.  Harris WS, Pottala JV, Sands SA, Jones PG. Comparison of the effects of fish and fish-oil capsules on the n 3 fatty acid content of blood cells and plasma phospholipids. Am J Clin Nutr. 2007 December 1, 2007;86:1621-5.
25.  Harris WS. Omega-3 fatty acids and cardiovascular disease: A case for omega-3 index as a new risk factor. Pharmacological Research. 2007;55:217-23.