Dietary Intakes of Arachidonic Acid and α-Linolenic Acid Are Associated with Reduced Risk of Hip Fracture in Older Adults

Abstract

PUFA are hypothesized to influence bone health, but longitudinal studies on hip fracture risk are lacking. We examined associations between intakes of PUFA and fish, and hip fracture risk among older adults (n = 904) in the Framingham Osteoporosis Study. Participants (mean age ~75 y at baseline) were followed for incident hip fracture from the time they completed the baseline exam (1988–1989) until December 31, 2005. HR and 95% CI were estimated for energy-adjusted dietary fatty acid exposure variables [(n-3) fatty acids: α-linolenic acid (ALA), EPA, DHA, EPA+DHA; (n-6) fatty acids: linoleic acid, arachidonic acid (AA); and the (n-6):(n-3) ratio] and fish intake categories, adjusting for potential confounders and covariates. Protective associations were observed between intakes of ALA (P-trend = 0.02) and hip fracture risk in a combined sample of women and men and between intakes of AA (P-trend = 0.05) and hip fracture risk in men only. Participants in the highest quartile of ALA intake had a 54% lower risk of hip fracture than those in the lowest quartile (Q4 vs. Q1: HR = 0.46; 95% CI = 0.26–0.83). Men in the highest quartile of AA intake had an 80% lower risk of hip fracture than those in the lowest quartile (Q4 vs. Q1: HR = 0.20; 95% CI = 0.04–0.96). No significant associations were observed among intakes of EPA, DHA, EPA+DHA, or fish. These findings suggest dietary ALA may reduce hip fracture risk in women and men and dietary AA may reduce hip fracture risk in men.

Introduction

PUFA may affect bone health via multiple mechanisms, which have been described in several reviews (1–6). In addition to modulating inflammatory cytokine production (7–11) and prostaglandin E₂ production (12), (n-3) fatty acids have been shown to favorably affect intestinal calcium transport (13) and calcium excretion (14). Both (n-3) and (n-6) fatty acids may be hypothesized to exert effects on bone by serving as ligands for PPARα and PPARγ (15–18) and by functioning as pro-resolving lipid mediators (19–21).

Beneficial effects of (n-3) fatty acids or a lower (n-6):(n-3) ratio on bone health outcomes have been demonstrated in numerous animal studies (22–32). In adult human studies, protective effects have been observed for (n-3) fatty acid intake, as well as a lower (n-6):(n-3) fatty acid ratio, in relation to hip bone mineral density (BMD)⁷ (33, 34) and for α-linolenic acid (ALA) in relation to bone resorption (35). Protective effects of supplementation with (n-3) fatty acids or fish oil in relation to bone formation markers (36), bone resorption markers (37), and bone mass (38) have also been observed. Additionally, fish intake was recently shown to be associated with less postflight BMD loss in astronauts (39). In young men (<18 y), serum DHA concentration was positively associated with total body and spine BMD (40).

Investigations on the relationship between (n-3) and (n-6) fatty acids and fracture risk are sparse. A case-control study of age- and sex-matched hospitalized elderly patients found a significantly increased risk of low-energy fracture among participants in the highest quartile of (n-6) fatty acid intake relative to those in the lowest quartile of intake, but no relationship between (n-3) fatty acids and fracture (41). In another study, no association was observed between either fish or EPA+DHA intakes and hip fracture incidence in over 5000 older adult men and women (42). Recently, total (n-6) fatty acid intake was inversely associated with total fracture risk, and EPA+DHA intake was positively associated with total fracture risk among over 135,000 postmenopausal women enrolled in the Women’s Health Initiative (WHI) (43).

To our knowledge, the association among individual types of short- and long-chain (n-3) and (n-6) fatty acids and fish intakes with hip fracture risk has not been examined in a cohort of both men and women in a longitudinal observation study. Thus, we evaluated the relationship between intakes of PUFA and fish and hip fracture risk over 17-y of follow-up among older adult men and women in the Framingham Osteoporosis Study.

Materials and Methods

Participants.

Participants were drawn from the Framingham Heart Study (FHS), which began in 1948 with the goal of identifying risk factors for cardiovascular disease. At its inception, 5209 women and men aged 28–62 y were recruited from a random sample of two-thirds of the households in Framingham, MA to participate in the initial examination and were examined biennially thereafter for >50 y (44). In 1988, the Framingham Osteoporosis Study was initiated as an ancillary study to the FHS and 1164 of the 1402 surviving members of the original cohort had BMD measurements taken at the 20th biennial examination (1988–1989). This study utilizes data from participants of the original cohort at exam 20 (baseline), who were followed for incident hip fracture from the date they completed a FFQ during their 20th biennial examination until December 31, 2005. We excluded participants with missing or invalid FFQ (based on the criteria of >12 food items left blank or energy intakes < 2512 or > 16,747 kJ/d) at exam 20 (n = 334 for missing FFQ and n = 92 for invalid FFQ). We further excluded participants with hip fracture prior to baseline (n = 30), hip fracture caused by trauma (n = 1), and missing covariate information for BMI (n = 18), smoking status (n = 3), or physical activity (n = 20). The final sample size (n = 904) comprised 552 women and 352 men. This study was approved by the Institutional Review Boards for Human Research at Boston University, Hebrew Rehabilitation Center, Institute for Aging Research, and Tufts University.

Assessment of PUFA and fish intake.

Usual dietary intake during the previous 12 mo was assessed with a self-administered, 126-item, semiquantitative Willett FFQ that was previously validated in other adult populations of men and women for several nutrients, including total PUFA and linolenic acid (LA) (45–48) and recently for EPA, DHA, and fish intakes in elderly participants via an interviewer-administered questionnaire (49). Additionally, a previous investigation using participants from the FHS cohort used in our study observed that both mean dietary DHA and fish intakes were significantly associated with plasma DHA concentrations (50). Participants were mailed the FFQ prior to their exam visit and asked to report their frequency of consumption of each food item before returning the completed FFQ at the exam visit. The FFQ identifies nutrient intakes from both food and vitamin and mineral supplements. (n-3) Fatty acid supplement use was identified by the FFQ, but few participants reported supplement use (n = 2 for women and n = 2 for men). The (n-6):(n-3) fatty acid ratio was derived by dividing total (n-6) fatty acid intake by total (n-3) fatty acid intake. Total fish intake (1 serving = 85–142 g or 3–5 oz) was defined as the sum of canned tuna, dark fish (mackerel, salmon, sardines, bluefish, and swordfish), white fish, and shellfish as reported by participants on the FFQ.

Assessment of hip fracture.

Hip fractures were reported by participants at each biennial exam upon interview, beginning at exam 18 in 1984. For those unable to attend examinations, hip fractures were reported by telephone interview. Occurrence of hip fractures was further identified through systematic review of medical records of hospitalizations and deaths and were confirmed by reviewing medical records and radiographic and operative reports, as previously described (51). Incident hip fracture was defined as first-time fracture of proximal femur that occurred following the date participants completed the FFQ at baseline through the follow-up period until December 31, 2005.

Confounding variables and covariates.

Covariates included factors known to affect BMD or fracture risk: age (y), BMI (kg/m²), height at exam 1 (m), dietary and supplemental intakes of calcium (mg/d) and vitamin D (μg/d), energy intake (kJ/d), alcohol intake, physical activity, smoking status, and estrogen use for women (52). Protein intake (g/d) was considered as a confounder, because it has been identified as a possible nutritional risk factor for osteoporosis (52) and because dietary sources of (n-3) fatty acids (namely fish and seafood) and the (n-6) fatty acid, arachidonic acid (AA), are also rich sources of protein. We also considered vitamin K (μg/d) and fruits and vegetables (servings/d) as confounders for (n-6) fatty acid exposures, because both vitamin K (53) and fruit and vegetable intakes (54) have been found to be associated with BMD and because vitamin K is a fat-soluble vitamin found in leafy green vegetables usually consumed with salad dressings, which are a rich source of the (n-6) fatty acid, LA. One serving of fruits and vegetables was considered the equivalent of 1 cup raw leafy vegetables (30 g), 1/2 cup cut-up raw or cooked vegetable (91 g), 1/2 cup fruit or vegetable juice (124 g), 1 medium whole fruit (138 g), 1/4 cup dried fruit (36 g), or 1/2 cup fresh, frozen, or canned fruit (115 g). Usual dietary intakes of calcium, vitamin D, alcohol, energy, protein, and vitamin K were previously assessed with the FFQ described. Intakes of calcium, vitamin D, and vitamin K were calculated as intakes from both food and supplement sources. For alcohol intake, participants were classified as nondrinkers, moderate drinkers (<13.2 g/d for women and <26.4 g/d for men), or heavy drinkers (≥13.2 g/d for women and ≥26.4 g/d) based on the Dietary Guidelines for Americans, which recommend a maximum 1 drink/d for women and 2 drinks/d for men (55). Physical activity was measured using the Framingham physical activity index (PAI), which represents the sum of the number of hours per day spent engaged in activity, weighted according to level of activity as follows: sleep (1.0), sedentary (1.1), slight (1.5), moderate (2.4), and heavy (5.0) (56). PAI from exam 19 (1986–1987) was used for participants with missing PAI at exam 20. Smoking status was determined based on questionnaire responses and was defined as nonsmoker, past smoker, or current smoker (smoked regularly in the past year). BMI (kg/m²) was calculated from weight measurements taken at exam 20 and height measurements taken at exam 1 (adult height achieved before height loss may have occurred as a result of osteoporosis). Because BMI is a measure of relative adiposity designed to be independent of height, we further adjusted for height at exam 1 to account for overall adult body size (57), which may be related to dietary intake and BMD. For women, estrogen use was defined as current use (currently using estrogen for >1 y) compared with never or past use. Never or past users were categorized together, because it has been shown that beneficial effects on bone are not retained with past estrogen use (58). Femoral neck-BMD (FN-BMD) (g/cm²) from exam 20 was also included in statistical models in a subset of the sample that had FN-BMD measurements available (n = 831) to evaluate whether associations observed among essential fatty acid and fish intakes with fracture risk are independent of the exposure’s effect on BMD. BMD of the right proximal hip was measured at baseline using a dual-photon absorptiometer (DP3, Lunar Radiation) (59).

Statistical analysis.

SAS statistical software (version 9.1; SAS Institute) was used to perform statistical analyses. P < 0.05 (2-sided) was considered significant for all analyses. Analyses were performed in both a combined sample and separately for women and men. We also tested for effect modification by sex by first including an interaction term in analyses on the combined sample. For final analyses on the combined sample, sex and estrogen use were adjusted by creating a categorical variable (0, 1, 2), such that: referent group = men, 1 = never or former estrogen-using women, and 2 = current estrogen-using women.

Cox proportional-hazards regression was used to estimate HR and 95% CI continuously and categorically by quartile of energy-adjusted dietary fatty acid exposure variables and fish intake categories. Fish intakes were categorized as low (<1 serving/wk), moderate (≥1 serving/wk, but <3 servings/wk), and high (≥3 servings/wk) to ensure that the AHA recommendation of consuming 2 servings of fish/wk would be included in the moderate fish intake category. Models were adjusted for age, sex (in the combined sample), estrogen use for women, BMI, height at exam 1, dietary and supplemental intakes of calcium and vitamin D, physical activity, smoking status, alcohol intake, and energy intake. For fish exposures, models were first adjusted for supplemental vitamin D intakes, followed by dietary vitamin D, to assess confounding by dietary vitamin D intake. We observed no attenuation of effect estimates or significance values after adjustment for dietary vitamin D; thus, final models for fish intake included dietary and supplemental intakes of vitamin D.

In addition to including total energy intake as a covariate in statistical models, we used the residual energy adjustment method in which the energy-yielding nutrients (fatty acid intakes and protein intake) were regressed on total energy intake to obtain nutrient intake residuals. Prior to performing regression models, these residuals were used to create quartiles of fatty acid intakes for categorical analyses. Using this method of energy adjustment provides a measure of nutrient intake that is not associated with energy intake (60).

Other possible confounding variables were also considered, including: protein intake, total fat intake, vitamin K intake, and fruit and vegetable intake [for the (n-6) fatty acid exposures only], and baseline FN-BMD in a subset of participants. We adjusted for total fat intake (residually energy adjusted and as an absolute amount in g/d + energy intake, separately in the model) to ensure that relationships observed between essential fatty acid intakes and bone were independent of total fat intake. Saturated fat intake has been inversely associated with hip BMD (61), whereas monounsaturated fat intake was found to be positively associated with BMD (62). We tested for effect modification by EPA+DHA intake for the (n-6) fatty acid exposures to examine whether (n-6) fatty acid intake would be associated with beneficial effects on fracture risk, when long-chain (n-3) fatty acid intake is sufficiently high to blunt detrimental effects of (n-6) fatty acids that may be mediated through inflammatory pathways (63). The EPA+DHA variable was categorized (0,1) as less than or equal to or greater than the median value (0.145 g/d for men and 0.14 g/d for women) to test for this interaction in addition to being analyzed as a continuous variable.

Dietary essential fatty acid exposure variables and calcium and vitamin K intakes were positively skewed. A natural logarithmic transformation was applied to intakes of ALA, EPA, DHA, EPA+DHA, calcium, and vitamin K, and a square root transformation was applied to intakes of LA and AA prior to performing all statistical analyses to achieve normality of the nutrient intakes and energy-adjusted nutrient intake residuals. Transforming skewed nutrient intake exposures has been recommended to improve heteroscedasticity that occurs as a result of greater variation in nutrient intakes at higher energy intakes (64). A test of linear trend across the quartiles of essential fatty acid intakes was also performed by creating a continuous variable set equal to the median intake for each quartile.

Results

Participant characteristics.

At baseline, women comprised 61% of the study population (Table 1). Proportionately more women (~39%) than men (~17%) reported (n-3) ALA intakes that meet or exceed the current Adequate Intake values of 1.1 g/d for women and 1.6 g/d for men according to DRI (55). The percentages of women and men reporting (n-6) LA intakes above the upper limit of the Acceptable Macronutrient Distribution Range of 10 g/d were ~42 and ~48%, respectively. The Acceptable Macronutrient Distribution Range is a range of intake that, when consumed in excess of the upper limit, is associated with increasing risk of chronic diseases (55). Mean fish intakes were consistent with current recommendations of the AHA to eat fish 2 times/wk (65). The type of fish was derived from mainly from white and other fish (~38% of total fish intake for both women and men) and tuna (~37% for women and ~32% for men), and dark fish accounted for ~12% (women) and ~17% (men). The median follow-up periods for women and men were 12.7 and 10.4 y, respectively (Table 2). Approximately 14% of women (78 of 552) and 6% of men (20 of 352) sustained hip fractures during the follow-up period. These figures are slightly lower for women, but similar for men, than estimations that 17 and 6% of Caucasian women and men, respectively, aged 50 y and older will experience a hip fracture in their lifetime (66). The person-time incidence rate was higher in women (12.06/1000 person-y), than in men (5.68/1000 person-y).

TABLE 1.

Participant characteristics at baseline exam 20 of the Framingham Study¹

	Women, n = 5522	Men, n = 352
Descriptive variables
Age, y	75.2 ± 4.8	75. 2 ± 4.9
BMI, 2	26.3 ± 5.0	27.0 ± 4.0
PAI	33.3 ± 4.9	33.8 ± 6.2
Smoking status, %
Past smoker	43.8	64.5
Current smoker	12.1	9.4
Estrogen use, %
Past or never	95.3	NA
Current	4.7	NA
FN-BMD, 2	0.724 ± 0.114	0.879 ± 0.146
Dietary variables
ALA intake, g/d	1.03 ± 0.48	1.05 ± 0.43
EPA intake, g/d	0.06 ± 0.07	0.06 ± 0.07
DHA intake, g/d	0.14 ± 0.12	0.14 ± 0.13
LA intake, g/d	9.92 ± 4.95	10.8 ± 5.33
AA intake, g/d	0.10 ± 0.05	0.11 ± 0.05
(n-6):(n-3) Fatty acid ratio	8.19 ± 2.91	8.95 ± 3.99
Energy intake, kJ/d	6990 ± 2,330	7870 ± 2,620
Protein intake, g/d	67.0 ± 24.2	69.4 ± 23.7
Alcohol use, %
Moderate	35.5	45.2
Heavy	16.9	19.9
Calcium intake,3mg/d	818 ± 437	765 ± 391
Vitamin D intake,3μg/d	8.3 ± 6.5	8.1 ± 6.9
Vitamin K intake,3μg/d	165 ± 112	141 ± 96.7
Fruit and vegetables,4servings/d	5.5 ± 2.8	4.8 ± 2.4
Fish variables,5servings/wk
Tuna	0.84 ± 1.01	0.67 ± 0.87
Dark fish	0.27 ± 0.62	0.35 ± 0.77
White and other fish	0.88 ± 1.02	0.79 ± 0.98
Shellfish	0.30 ± 0.55	0.28 ± 0.41
Total fish	2.29 ± 2.02	2.09 ± 1.84

¹Values are mean ± SD or percent.

²Based on participants with complete diet and covariate information followed for hip fracture until the end of 2005. For FN-BMD (g/cm²), = 831 for the subset of participants with measurements available.

³Intake from diet plus supplements.

⁴One serving of fruit and vegetables equivalent to 1 cup raw leafy vegetables (30 g), 1/2 cup cut-up raw or cooked vegetable (91 g), 1/2 cup fruit or vegetable juice (124 g), 1 medium whole fruit (138 g), 1/4 cup dried fruit (36 g), or 1/2 cup fresh, frozen, or canned fruit (115 g).

⁵One serving of fish equivalent to 85–142 g (3–5 oz).

TABLE 2.

Descriptive characteristics of hip fracture occurrence and follow-up time

	Women, n = 552	Men, n = 352
Follow-up, y
Mean ± SD	11.7 ± 5.07	10.0 ± 5.46
Median	12.7	10.4
Person-y, n	6467	3522
Hip fractures, n	78	20
Crude incidence rate/1000 person-y1	12.1 (5.25, 18.9)	5.68 (1.01,10.4)

¹Crude incidence rate calculated as total number of hip fractures divided by total number of person-y/1000 person-y, 95% CI in parentheses.

Associations between fatty acid intakes and hip fracture.

Associations between PUFA intakes and fracture risk were similar in analyses conducted separately in men and women in both continuous and categorical (quartile) models, with the exception that significant protective associations were observed between intakes of AA and hip fracture risk in men, but not women. The interaction between sex and AA intake bordered significance (P = 0.05). No significant interactions with sex were observed for any of the other essential fatty acid intake exposures (P > 0.05). Results of analyses in the combined sample are therefore presented for intakes of ALA, EPA, DHA, EPA+DHA, LA, and the (n-6):(n-3) ratio, and results of analyses conducted separately in women and men are presented for AA intake. Because results of the continuous and categorical analyses were similar, we present only results of categorical analyses. Additionally, no significant interactions with EPA+DHA intakes, expressed categorically (less than or equal to or greater than the median) or continuously, were observed for the (n-6) fatty acid exposures (P > 0.05).

No significant associations were observed between intakes of EPA, DHA, EPA+DHA, LA, or the (n-6):(n-3) fatty acid ratio and hip fracture risk in the combined sample of men and women (Table 3). Higher ALA intake was associated with lower hip fracture risk (P-trend = 0.02). Participants in the highest quartile of ALA intake had a 54% lower risk of hip fracture than those in the lowest quartile (Q4 vs. Q1: HR = 0.46, 95% CI = 0.26–0.83) and participants in quartile 2 had a 50% lower risk of hip fracture than those in the lowest quartile (Q2 vs. Q1: HR = 0.50, 95% CI = 0.29–0.88). When adjusted for baseline BMD, the HR was not substantially changed compared with models in the subsample of participants with baseline BMD measures available. Additional adjustment for intakes of protein, total fat, and fruit and vegetable or vitamin K [for the (n-6) fatty acids] did not substantially change the HR or significance levels.

TABLE 3.

HR (95% CI) for hip fracture according to quartile of PUFA intakes among women and men combined in the FHS^1–3

	Q2	Q3	Q4	P-trend
ALA	0.50 (0.29, 0.88)*	0.70 (0.42, 1.16)	0.46 (0.26, 0.83)**	0.02
EPA	1.01 (0.58, 1.78)	1.11 (0.64, 1.93)	0.80 (0.44, 1.47)	0.60
DHA	0.62 (0.34, 1.10)	0.94 (0.55, 1.59)	0.74 (0.42, 1.31)	0.49
EPA+DHA	0.83 (0.48, 1.43)	0.71 (0.40, 1.27)	0.83 (0.47, 1.46)	0.41
LA	0.75 (0.43, 1.32)	0.82 (0.47, 1.43)	0.73 (0.41, 1.29)	0.33
(n-6):(n-3) Fatty acid ratio	0.80 (0.44, 1.46)	1.09 (0.63, 1.86)	0.95 (0.53, 1.69)	0.93

¹Adjusted for age, sex, and estrogen use in women, BMI, height at exam 1, dietary and supplemental intakes of calcium and vitamin D, physical activity, smoking status, alcohol intake, and energy intake. Energy adjustment performed using the residual method ( = 904). *P < 0.05 relative to Q1; **P < 0.01 relative to Q1.

²Prior to energy adjustment, natural log transformation applied to ALA, EPA, DHA, EPA+DHA, and (n-6):(n-3) fatty acid ratio; square root transformation applied to LA.

³Q1 is reference group (HR = 1.00).

Higher AA intake was associated with lower hip fracture risk in men (P-trend = 0.05) but not women (Fig. 1). Men in the highest quartile of AA intake had an 80% lower risk of hip fracture than those in the lowest quartile (Q4 vs. Q1: HR = 0.20, 95% CI = 0.04–0.96). Adjustment for protein intake attenuated this association (Q4 vs. Q1: HR = 0.49, 95% CI = 0.07–3.59), but adjustment for total fat intake or baseline BMD (in the subsample with available BMD measures) did not substantially attenuate HR or significance levels.

FIGURE 1.

HR for the risk of hip fracture according to quartile of AA intake in the FHS for women (n = 552) (A) and men (n = 352) (B). Adjusted for age, BMI, height at exam 1, dietary and supplemental intakes of calcium and vitamin D, physical activity, smoking status, alcohol intake, energy intake, and estrogen use for women. Energy adjustment performed using the residual method. Square root transformation applied to AA prior to energy adjustment. Q1 is reference group. Bars are 95% CI. *Different from Q1, P < 0.05.

Associations between fish intake and hip fracture.

No significant associations were observed among intakes of total fish, dark fish, tuna, or dark fish + tuna and hip fracture in either simple or multivariable-adjusted models in the combined sample of men and women (Table 4). Associations were similar in analyses conducted separately in women and men and no significant interactions with sex were observed for any of the fish types (P > 0.05). Additional adjustment for protein intake, total fat intake, baseline BMD (in the subsample with available BMD measures), or remaining fish intake (for dark fish, tuna, and dark fish + tuna exposures) did not substantially change the HR or significance levels.

TABLE 4.

HR (95% CI) for hip fracture according to level of fish intakes among women and men combined in the FHS¹²

	Moderate	High	P-trend
Total fish	0.84 (0.53, 1.34)	0.66 (0.38, 1.17)	0.28
Dark fish	0.90 (0.43, 1.88)	1.23 (0.41, 3.67)	0.75
Tuna	1.00 (0.62, 1.62)	0.78 (0.38, 1.59)	0.63
Dark fish + tuna	1.03 (0.64, 1.62)	0.99 (0.53, 1.85)	0.82

¹Multivariable adjusted for age, sex, and estrogen use in women, BMI, height at exam 1, dietary and supplemental intakes of calcium and vitamin D, physical activity, smoking status, alcohol intake, and energy intake ( = 904).

²Low fish intake is reference group (HR = 1.00); low = < 1 serving/wk, moderate = ≥1/<3 servings/wk, and high = ≥3 servings/wk.

Discussion

In this longitudinal observational study, ALA intake was significantly associated with reduced risk of hip fracture in a combined sample of women and men, and AA intake was significantly associated with reduced risk of hip fracture in men but not women. Fish consumption, intakes of the (n-3) fatty acids derived from marine sources (EPA, DHA, and EPA+DHA), LA, and the (n-6):(n-3) fatty acid ratio were not associated with hip fracture risk.

Few data are available that describe the effects of ALA on bone health. Dietary ALA intervention has been shown to reduce the bone resorption marker N-telopeptide in humans (35). A diet of flaxseed, rich in ALA, combined with low doses of estrogen was shown to preserve BMD and biomechanical strength of vertebrae in ovariectomized rats (67). Our findings suggest that the protective effect of ALA on hip fracture risk in humans may be independent of BMD, because adjustment for baseline FN-BMD did not substantially attenuate this association. A previous investigation by our group that found no association between ALA intake and either baseline hip BMD or 4-y loss of hip BMD in the Framingham Study supports this notion (E.K. Farina, D.P. Kiel, R. Roubenoff, E.J. Schaefer, L.A. Cupples, and K.L. Tucker, unpublished data). It is not likely that the protective effect of ALA that we observed was due to conversion of ALA to EPA and DHA, because conversion is generally regarded as limited (68). Thus, ALA may reduce hip fracture via alternative mechanisms, such as protective effects on bone quality not measured by BMD. These effects may include a reduction in bone turnover, as suggested by the reduction in N-telopeptide following ALA intervention observed by Griel et al. (35).

The strongest protective effects of ALA were observed among participants in the highest quartile of energy-adjusted ALA intake, which corresponded to a mean ALA intake of 1.39 g/d. This level of intake can be easily achieved, because ALA is found in a variety of foods, including leafy green vegetables, nuts, canola oil, and particularly in flaxseed and flaxseed oil. One tablespoon (1 tablespoon = 15 mL) of canola oil, e.g., contains ~1.3 g of ALA, 1 tablespoon of flaxseed oil contains ~7 g, and 1 tablespoon of ground flaxseed contains ~1.6 g (69). Our findings are in contrast to those among postmenopausal women enrolled in the WHI in which no association between ALA intake and hip fracture was observed (43). However, average ALA intake in the highest quartile was reported to be 0.84 g/d in this study, which is considerably lower than the average of 1.39 g/d in the highest quartile (Q4) of ALA intake in our study.

AA intake was not associated with hip fracture risk in the combined sample of women and men, but in multivariable-adjusted models among men only, those in the highest quartile of AA intake had an 80% reduced risk of hip fracture relative to those in the lowest quartile of AA intake. This protective effect of AA may be due in part to protein intake, because adjustment for protein attenuated this association. However, we may not be able to separate the effects of AA and protein intake, because AA is found in protein-rich animal food sources, including eggs and meats. Indeed, energy-adjusted intakes of AA and protein were correlated among the men in this study (r = 0.62). In addition, we previously observed that participants within the highest quintile of plasma phosphatidylcholine AA had a significantly lower risk of hip fracture than participants within the lowest quintile (E.K. Farina, D.P. Kiel, R. Roubenoff, E.J. Schaefer, L.A. Cupples, and K.L. Tucker, unpublished data). Taken together, this may also suggest that protective effects of protein intake on bone health may be partly due to AA, but we are not able to definitely elucidate these mechanisms in this study.

AA itself may protect bone by suppressing NF-κB activation (15, 16, 70, 71, 72) or by generating lipoxins involved in resolving inflammation. A derivative of AA, 12d-PGJ2, has also been shown to stimulate collagen synthesis in human osteoblast cells (72, 73, 74). Dietary supplementation with AA has been shown to elevate bone mass in piglets (74, 75, 76) and maternal umbilical cord AA has been related to bone mass in healthy full-term infants in humans (76).

We found no evidence of an association between either long-chain (n-3) fatty acid intakes or fish intakes and hip fracture risk. These findings are consistent with other studies (41, 42) and with the WHI investigation in relation to hip fracture (43), although EPA+DHA intake was associated with a small increased risk of total fracture in this cohort (43). One explanation for a lack of association in our study is that the fish intakes estimated in our study may not have been representative of long-term intake. Public health messages on the health benefits of eating fish are relatively recent, because large-scale prospective epidemiological studies investigating relationships between fish intake and coronary heart disease did not emerge until the mid 1980s (65). Recent changes to fish intake in the short term may be more likely to affect associations between fish intake and BMD than between fish intake and fracture risk.

It is also possible that the amount of EPA and DHA consumed in the diet by study participants and contributed by fish intake (even high fish intakes of ≥3 servings/wk) was not enough to reduce bone loss sufficiently to translate to a reduction in hip fracture risk. Average EPA+DHA intakes among women and men in high-fish intake categories were, respectively, 0.41 and 0.42 g/d for total seafood, 0.86 and 0.94 g/d for dark fish, 0.53 and 0.52 g/d for tuna, and 0.56 and 0.61 g/d for dark fish and tuna. Clinical trials investigating cardiovascular disease have been noted to use doses of EPA+DHA of 1 g/d in supplement form. Only 3 men and 2 women in our study achieved an EPA+DHA intake ≥ 1 g/d and all of these participants consumed ≥3 servings/wk of dark fish.

Our study has some limitations. The dietary exposures we examined were obtained from a FFQ, which obtains dietary information in a semiquantitative manner. This instrument is useful for ranking participants to distinguish one another according to intake rather than estimating precise, absolute intakes. However, validation of this FFQ with subcutaneous fat as the comparison measure showed reasonable correlations of 0.50 for total PUFA, 0.47 for EPA, and 0.48 for LA when these nutrients were expressed as percentage of total fat (48). A recent validation of this FFQ to estimate EPA, DHA, and fish intakes against plasma phospholipid fatty acid profiles also showed modest correlations of 0.37 for EPA, 0.48 for DHA, and 0.48 for EPA+DHA, and correlations ranging from 0.30 to 0.36 for dark fish, dark fish and tuna, and total fish (49). Additionally, we were unable to account for changes in diet that may have occurred over the follow-up period, including the use of (n-3) fatty acid supplements. For example, the prevalence of (n-3) fatty acid supplement use in a longitudinal study of adult men and women aged 43–86 y at baseline (n = 4926) was reported to be 0.3% at baseline (1988–1990), and at each subsequent follow-up survey, conducted at 5-y intervals, was 0.3% (1993–1995), 1.0% (1998–2000), and 5.8% (2003–2005) (77). The prevalence of (n-3) fatty acid supplement use reported in our study of 0.4% at baseline (1988–1989) is consistent with the prevalence of 0.3% at baseline in the aforementioned study. However, it is not known whether participants in our study similarly increased (n-3) fatty acid supplement use over the course of follow-up or whether this increase in exposure would affect fracture risk estimates. Future longitudinal studies examining relationships between PUFA and effects on fracture risk would be strengthened by ensuring that information on both dietary and supplemental sources of fatty acids is collected throughout the follow-up period.

Nevertheless, to our knowledge, this is the first longitudinal study to examine the association among individual types of short- and long-chain PUFA with hip fracture risk in men and women. Our findings suggest that ALA intake may reduce the risk of hip fracture in women and men and that AA intake may reduce the risk of hip fracture in men. Further population-based research is needed to confirm our results and determine whether longer exposure periods to higher fish intakes and EPA and DHA intakes that account for (n-3) fatty acid supplementation may be related to hip fracture risk.