researchnews.osu.edu - Omega-3 Supplements May Slow a Biological Effect of AgingBy Emily Caldwell. October 1, 2012
Questions:1. Why do you think that they didn't exclude those volunteers who were taking an aspirin a day? I only can think of a possible explanation.
2. Did they compare change in telomerase activity vs. change in telomere length? If so, why didn't they show those results?
Comment:I have the suspicion that the reason why the telomeres of the peripheral T-cells are more prone to lengthened with omega-3 supplementation is because with the modulation of the pro-inflammatory signal induced by omega-3, the T-cell precursors proliferate less, and as it's mentioned in the same paper that presents the results of this study,
"Inflammation triggers T-cell proliferation, one known cause of telomere shortening."_________________ Excerpt from Ohio State University press release:<<The study showed that most overweight but healthy middle-aged and older adults who took omega-3 supplements for four months altered a ratio of their fatty acid consumption in a way that helped preserve tiny segments of DNA in their white blood cells.>>
<<Omega-3 supplementation also reduced oxidative stress, caused by excessive free radicals in the blood, by about 15 percent compared to effects seen in the placebo group.>>
<<In another recent publication from this study, Kiecolt-Glaser and colleagues reported that omega-3 fatty acid supplements lowered inflammation in this same group of adults.>>
<<Study participants took either 2.5 grams or 1.25 grams of active omega-3 polyunsaturated fatty acids>>
<<Participants on the placebo took pills containing a mix of oils representing a typical American’s daily intake.>>
<<Participants received either the placebo or one of the two different doses of omega-3 fatty acids. The supplements were calibrated to contain a ratio of the two cold-water fish oil fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), of seven to one. Previous research has suggested that EPA has more anti-inflammatory properties than DHA.>>
<<Both groups of participants who took omega-3 supplements showed, on average, lengthening of telomeres compared to overall telomere effects in the placebo group, but the relationship could have been attributed to chance. However, when the researchers analyzed the participants’ omega-6 to omega-3 ratio in relationship to telomere lengthening, a lower ratio was clearly associated with lengthened telomeres.>>
<<The researchers also measured levels of compounds called F2-isoprostanes to determine levels of oxidative stress, which is linked to a number of conditions that include heart disease and neurodegenerative disorders. Both omega-3 groups together showed an average overall 15 percent reduction in oxidative stress compared to effects seen in the placebo group.
When the scientists revisited their earlier inflammation findings, they also found that decreases in an inflammatory marker in the blood called interleukin-6 (IL-6) were associated with telomere lengthening. In their earlier paper on omega-3s and inflammation, they reported that omega-3 supplements lowered IL-6 by 10 to 12 percent, depending on the dose. By comparison, those taking a placebo saw an overall 36 percent increase in IL-6 by the end of the study.
“This finding strongly suggests that inflammation is what’s driving the changes in the telomeres,” Kiecolt-Glaser said.>>
<<study co-author Ron Glaser, professor of molecular virology, immunology and medical genetics and director of the Institute for Behavioral Medicine Research (IBMR) at Ohio State.>>
<<this population was disease-free and reported very little stress. The study included 106 adults, average age 51 years, who were either overweight or obese and lived sedentary lives. The researchers excluded people taking medications to control mood, cholesterol and blood pressure as well as vegetarians, patients with diabetes, smokers, those routinely taking fish oil, people who got more than two hours of vigorous exercise each week and those whose body mass index was either below 22.5 or above 40.>>
<<Co-authors of the study include Elissa Epel, Jue Lin and Elizabeth Blackburn of the University of California, San Francisco; Rebecca Andridge and Beom Seuk Hwang of Ohio State’s College of Public Health; and William Malarkey of the IBMR.
This work was supported in part by grants from the National Institutes of Health.
OmegaBrite, a company based in Waltham, Mass., supplied the supplements as an unrestricted gift but did not participate in the study design, results or publication. Study co-authors Blackburn, Epel and Lin are co-founders of Telome Health Inc., a telomere measurement company>>
Source:- Caldwell, Emily.
Omega-3 Supplements May Slow A Biological Effect of Aging. The Ohio State University - Research and Innovation Communications. October 1, 2012researchnews.osu.edu/archive/omega3aging.htm _______________________ Reference paper:
- Kiecolt-Glaser
et al. Omega-3 fatty acids, oxidative stress, and leukocyte telomere length: A randomized controlled trial. Brain Behav Immun (2012) [uncorrected proof, article in press]
ncbi.nlm.nih.gov/pubmed/23010452 linkinghub.elsevier.com/retrieve/pii/S0889-1591(12)00431-X sciencedirect.com/science/article/pii/S088915911200431X Excerpts of interest:
1.1. Telomeres, telomerase, inflammation, and oxidative stressTelomeres, the caps found at the ends of chromosomes, are essential for chromosomal stability and replication; the enzyme telomerase is important for telomere formation, maintenance, and restoration (Blackburn, 2005; Epel et al., 2004). A growing literature has linked shorter telomeres with health behaviors, age-related diseases, and earlier mortality (Brouilette et al., 2003; Epel et al., 2009; Kimura et al., 2008; Valdes et al., 2005).
Telomeres can be maintained or lengthened by telomerase, an intra-cellular enzyme that adds telomeric DNA to shortened telomeres (Chan and Blackburn, 2003).
Telomere length is also linked to, and likely regulated by, exposure to proinflammatory cytokines and oxidative stress (Aviv, 2006; Carrero et al., 2008; Damjanovic et al., 2007).
Inflammation triggers T-cell proliferation, one known cause of telomere shortening (Aviv, 2004; Carrero et al., 2008; Gardner et al., 2005).
Oxidative stress promotes telomere erosion during cellular replication in vitro and also stimulates the synthesis of proinflammatory cytokines (Aviv, 2006; Lipcsey et al., 2008).
1.2. Telomeres, telomerase, and omega-3 PUFAsAlthough
telomeres typically shorten with aging, shortening is not inevitable, and telomeres can also lengthen (Aviv et al., 2009; Ehrlenbach et al., 2009; Epel et al., 2009; Farzaneh-Far et al., 2010a; Nordfjall et al., 2009). It is important to identify malleable factors that might promote telomere stability over time. Based on theoretical and empirical reasons, it is possible that blood levels of polyunsaturated fatty acids (PUFAs) may be one of the factors that can prevent telomere shortening over time. The omega-3 (n-3) PUFAs can reduce inflammation and decrease oxidative stress (Calder, 2005; Kiecolt-Glaser et al., 2011; Mori et al., 1999; Nalsen et al., 2006), described below, and thus could buffer telomeres from their damaging effects.
In the Heart and Soul Study, which followed 608 people with stable coronary heart disease over 5 years, average telomere length increased in 23% of the individuals, shortened in 45%, and remained unchanged in 32% (Farzaneh-Far et al., 2010a).
Slower telomere attrition was predicted by higher baseline levels of the two key n-3 PUFAs, eicosapentaenoic acid (EPA) and docosahexanoic acid (DHA), which were
the only significant predictors out of 16 clinical and behavioral factors examined (Farzaneh-Far et al., 2010b).
Each standard deviation increase in the DHA + EPA total was associated with a 32% reduction in the odds of telomere attrition. In a different pilot study, an intensive
three-month lifestyle change program that included n-3 PUFA supplementation significantly increased telomerase activity (Ornish et al., 2008).
1.3. The present studyIn our recent four-month randomized controlled trial (RCT), serum interleukin 6 (IL-6) decreased by 10% and 12% in our low (1.25 g/day) and high (2.5 g/day) dose n-3 PUFA groups, respectively, compared to a 36% increase in the placebo group (Kiecolt-Glaser et al., 2012). Similarly, low and high dose n-3 PUFA groups showed modest 0.2% and -2.3% changes in serum TNF-α, in contrast to the 12% increase in the control group. Depressive symptoms, the other primary trial outcome, were low at baseline and did not change. This study assessed the impact of n-3 PUFA supplementation, and consequent changes in the n-6:n-3 PUFA ratio, on secondary outcomes in our RCT: leukocyte telomere length, telomerase, and oxidative stress.
2.2. Design and study components 164Data collection for this double-blind placebo-controlled four 165 month RCT began in September, 2006 and ended in February, 2011. At baseline and at 4 months we assessed telomere length, telomerase, and oxidative stress. Blood samples were collected between 7:00 and 9:00 AM to control for diurnal variation.
2.2.1. Supplement and Placebo[ . . . ]
Both the placebo and the* n-3 PUFA pills included 1 IU of vitamin E. In order to ensure integrity of the oil supplements, dietary oils were analyzed every 6–8 months* by gas chromatography of methylated fatty acids prepared in the Belury lab.
2.6. Telomere lengthPeripheral blood lymphocytes (PBL) were purified from whole blood by density-gradient centrifugation in Lymphocyte Separation Medium (Mediatech, Inc.).
[ . . . ]
The telomere length measurement assay is adapted from the published original method by Cawthon (Cawthon, 2002; Lin et al., 2010a). The telomere thermal cycling profile consists of: Cycling for T(telomic) PCR: denature at 96 °C for 1 s, anneal/extend at 54 °C for 60 s, with fluorescence data collection, 30 cycles. Cycling for S (single copy gene) PCR: denature at 95 °C for 15 s, anneal at 58 °C for 1 s, extend at 72 °C for 20 s, 8 cycles; followed by denature at 96°C for 1s, anneal at 58°C for 1s, extend at 72 °C for 20 s, hold at 83 °C for 5 s with data collection, 35 cycles. The primers for the telomere PCR are
tel1b [5'-CGGTTT(GTTTGG)5GTT-3'], used at a final concentration of 100 nM, and _tel2b_ [5'-GGCTTG(CCTTAC)5CCT-3'], used at a final concentration 282 of 900 nM. The primers for the single-copy gene (human beta-globin) PCR are
hbg1 [5'- GCTTCTGACACAACTGTGTTCACTAGC-3'], used at a final concentration of 300 nM, and
hbg2 [5'-CACCAACTTCATCCACGTTCACC-3'], used at a final concentration of 700 nM. The final reaction mix contains 20 mM Tris–HCl, pH 8.4; 50 mM KCl; 200 lM each dNTP; 1% DMSO; 0.4x Syber Green I; 22 ng
Escherichia coli DNA per reaction; 0.4 U of Platinum Taq DNA polymer- ase (Invitrogen Inc.) per 11 ll reaction; 0.5–10 ng of genomic DNA. Tubes containing 26, 8.75, 2.9, 0.97, 0.324 and 0.108 ng of a reference DNA (from Hela cancer cells) are included in each PCR run so that the quantity of targeted templates in each sample can be determined relative to the reference DNA sample by the standard curve method. Each concentration of the reference DNA is run as quadruplets and samples are run as triplicates. To control for inter-assay variability, 8 control DNA samples from cancer cell lines (including 293T, H1299, UMUC3, and UMUC3 cells infected with a lentiviral construct containing the telomerase RNA gene to extent telomeres, harvested at various population doublings after infection) are included in each run. In each batch, the T/S ratio of each control DNA is divided by the average T/S for the same DNA from 10 runs to get a normalizing factor. This is done for all eight samples and the average normalizing factor for all eight samples is used to correct the participant DNA samples to get the final T/S ratio. The T/S ratio for each sample is measured twice, each time in triplicate wells. When the duplicate T/S value and the initial value vary by more than 7%, the sample is run the third time and the two closest values will be reported. The formula to convert the T/S ratio to base pairs is base pairs = 3274 + 2413 * (T/S). The inter-assay coefficient of variation for telomere length measurement was 4.3% for this study.
2.7. Telomerase activityPBLs were purified from whole blood as above. Cells were lysed with 1x CHAPS buffer (10 mM Tris–HCl, pH 7.5, 1 mM MgCl2, 315 1 mM EGTA, 0.1 mM benzamidine, 5 mM β-mercaptoethanol, 0.5% CHAPS, 10% glycerol) on ice for 30 min and spun at 4 °C at 14 k rpm for 20 min to generate an extract corresponding to 10,000 cells/ll. Extracts were stored at -80° for batch analysis of telomerase activity. Telomerase activity was measured by the TRAPeze Telomerase detection kit (Millipore, Cat# S7700) using a modified protocol developed by the Blackburn lab (Lin et al., 2010a). Three concentrations (2000, 5000 and 10,000 cells) were used for TRAP reactions to ensure that the assay was in the linear range. Details of the method are published elsewhere (Lin et al., 2010a). The inter-assay coefficient of variation (CV) was 6.8%.
2.8. Oxidative stressF2-isoprostanes provide the most reliable index of in vivo oxidative stress when compared to other well-known biomarkers (Milne et al., 2007). Plasma samples were analyzed by Vanderbilt’s Eicosanoid Core Laboratory, following their published protocol (Milne 331 et al., 2007).
2.9. Statistical methods[ . . . ]
Analysis of covariance was used to separately model change in telomerase (N = 94), telomere length (N = 106), and F2-isoprostanes (N = 97) from baseline to 4 months, adjusting for baseline levels, using all subjects with available four-month follow-up data. In order to control Type I error, the Tukey–Kramer method was used for between-group comparisons. Although supplementation group was the main predictor of interest, secondary analyses used change in continuous n-6:n-3 PUFA ratio in place of group because individuals differ in absorption and metabolism of n-3 PUFA supplements, as well as in adherence. Despite the relative balance between the groups due to the randomization, analyses were repeated controlling for age, gender, and sagittal abdominal diameter (SAD). Alpha was set to 0.05, and two-sided tests were conducted. All analyses were carried out in SAS version 9.3 (SAS Institute, Cary, NC).
3.1. Study population, baseline dataTable 1 shows baseline characteristics of the analysis sample (N = 106), with
31 subjects in the placebo arm, 40 in the low dose fish oil arm, and 35 in the high dose fish oil arm. Randomization produced groups that did not differ on age, baseline FFQ dietary variables, sleep quality, depressive symptoms, and history of major depressive disorder, p > 0.19 for all tests. There were no baseline group differences on SAD or BMI (p > 0.60 for both). Using BMI cut points of 25 and 30 kg/m2, 100 participants (94%) were overweight, and 54 (51%) were obese. Groups were similar on telomere length, telomerase, and F2-isoprostanes at baseline (p > 0.26 for all tests).
3.3. Changes in F2-isoprostanesTable 4 shows the significant group differences in changes in log-F2-isoprostanes after supplementation. The estimated mean *change in log-F2-isoprostanes was 0.073 for the placebo group, corresponding to an 8% increase in geometric mean.* In contrast, the estimated mean
change in log-F2-isoprostanes was -0.094 for the low dose group and -0.086 for the high dose group, corresponding to decreases in the geometric mean of 9% and 8%, respectively. For both doses these changes were significantly different than the placebo group (Tukey–Kramer adjusted p = 0.02; p = 0.04, respectively), resulting in the intervention groups having a 15% lower geometric mean F2-isoprostanes at 4 months compared to control.
There was not a significant difference between the two supplemented groups (p = 0.99). These results remained the same in analyses additionally controlling for age, gender and SAD.
3.4. Changes in telomere length and telomeraseThe adjusted mean
change in telomere length, expressed in base pairs (bp), was an
increase of 21 bp for the low dose group 406 and an increase of 50 bp in the high dose group compared to a decrease of 43 bp for placebo (Table 4); however;
differences between the groups were not significant. Telomere lengthening (defined as a positive change) was observed
in 54% (n = 19) of the 2.5 g/d n-3 PUFA group and 53% (n = 21) of the 1.25 g/d n-3 PUFA group, but only 39% (n = 12) of the placebo group, though these
differences were not significant (p = 0.39). There were
no differences among the groups in change in telomerase activity at four months (Table 4). Models additionally
controlling for age, gender, and SAD produced similar results.3.5. Changes in telomere length based on n-6:n-3 PUFA plasma ratiosSecondary analyses explored the effect of changes in plasma n-6:n-3 PUFA ratios on changes in telomere length, since individuals differ in absorption and metabolism of n-3 PUFA supplements. Table 5 shows the resulting linear regression analysis, controlling for baseline telomere length and baseline n-6:n-3 PUFA ratio. A one unit decrease in n-6:n-3 PUFA ratio was associated with an estimated 20 bp increase in telomere length (p = 0.02).
The analysis was repeated using the change in AA:(EPA + DHA) ratio in place of n-6:n-3 PUFA ratio; the AA:(EPA + DHA) ratio is favored by some researchers because of a more direct tie to eicosanoid metabolism. Since the change in AA:(EPA + DHA) ratio was highly correlated with the change in n-6:n-3 PUFA ratio (r = 0.90, p < 0.001), results were similar, with a one unit decrease in AA:(EPA + DHA) ratio associated with a 35 bp increase in telomere length (p = 0.08). Similarly, when the change in the sum of EPA, DHA, and DPA was used in place of either ratio, results were again comparable, with a one unit increase in EPA + DHA + DPA associated with a 22 bp increase in telomere length (p = 0.07). All results were similar after adjusting for age, gender, and SAD.
3.6. Changes in telomere length related to IL-6Since supplementation reduced serum IL-6 in both low and high dose groups in the parent study (Kiecolt-Glaser et al., 2012), as a secondary analysis we investigated the association between *change in telomere length and change in IL-6* for the 101 subjects in the present study who had IL-6 data available at baseline and four months. There was a
significant negative correlation between change in telomere length and change in IL-6 (Spearman r = -0.20, p = 0.05). Of the 51 subjects who experienced telomere lengthening, 61% (n = 31) had lowered IL-6, compared to 34% (n = 17) of the 50 who did not experience telomere lengthening (p = 0.007).
4.1. Intervention-related changes[ . . . ]
Telomerase activity level did not change in our sample,
in contrast to the changes observed following an intensive three-month lifestyle change program in a different study group – men with early prostate cancer – that included 3 g/day of fish oil (Ornish et al., 2008).
That intervention also included dietary change (low-fat and high plant-based), aerobic exercise, and stress management. Further, as that previous study had no non-intervention control group, those data must be interpreted cautiously as regards involvement of n-3 PUFAs.
Depressive symptoms were quite low at baseline in this sample, and did not change (Kiecolt-Glaser et al., 2012). However, prior studies that have linked lower n-3 PUFA plasma levels and depression suggest a potential benefit for more distressed groups (Appleton et al., 2010; Hibbeln, 1998). Depression and chronic stress have been associated with shorter telomeres (Damjanovic et al., 2007; Epel et al., 2004; Wolkowitz et al., 2010). Depression and chronic stress boost inflammation (Glaser and Kiecolt-Glaser, 2005; Kiecolt-Glaser et al., 2003) as well as oxidative stress (Epel et al., 2004; Wolkowitz et al., 2010), and could speed telomere erosion through these pathways. Accordingly, n-3 PUFA supplementation might also slow telomere attrition by enhancing mood in more depressed samples.
Although age-related reductions in telomeres are the average situation, recent studies showed that telomeres can both shorten and elongate in vivo, and leukocyte telomere length can change 491 within a period of months (Shlush et al., 2011; Svenson et al., 2011). Alterations in oxidative stress were highlighted as a potential mechanism in two recent reports (Shlush et al., 2011; Svenson 494 et al., 2011). Our data suggest that a dietary intervention that reduces the joint burden of oxidative stress and inflammation may in turn have positive consequences for telomere length. During aging there is a shift from naïve to memory T-cells, and the latter have shorter telomeres (Svenson et al., 2011). We do not know if the reductions in inflammation and oxidative stress in our n-3 PUFA supplemented participants reflected shifts in leukocyte subpopulations that contributed to the telomere changes observed, one limitation of the present study.
[ . . . ]
It would have been desirable to examine n-6 and n-3 PUFAs in red blood cells (RBCs) in addition to plasma levels. Circulating PUFA levels reflect the interplay among dietary intake, absorption, and metabolism and are not always strongly correlated with dietary intake of fatty acids (Fusconi et al., 2003; Seierstad et al., 2005). RBC PUFA levels reflect longer-term PUFA consumption as the turn-over is slow and more reliable (Harris, 2008, 2009). For example, DHA levels in RBCs are thought to indicate dietary fat intake for the past four months, while levels in plasma may only mirror intake from the last few days (Arab, 2003; Sun et al., 2007). However, our intervention spanned four months and serum and plasma proinflammatory cytokines can change in hours (Glaser and Kiecolt-Glaser, 2005); for example, infusion of a fish oil-based lipid emulsion substantially reduced monocyte production of IL-6, TNF-α, IL-1, and IL-8 in response to endotoxin (Mayer et al., 2003). For these reasons, plasma PUFA data were essential to assess recent dietary influences on inflammatory markers in this study. As described earlier, the n-6 and n-3 PUFAs compete for key enzymatic pathways, and thus the relative balance is of interest (Simopoulos, 2008). ATTICA, a large health and nutrition survey of healthy Greek adults, showed that higher n-6:n-3 PUFA plasma ratios were associated with higher TNF-α and IL-6 (Kalogeropoulos et al., 2010).
4.2. Health implicationsWe found that telomere length increased with decreasing n-6:n-3 PUFA ratios. These data suggest that rather than just considering the absolute amount of n-3 PUFA, the background levels of both the n-6 and the n-6:n-3 PUFAs should also be taken into account for clinical studies or for evaluation of nutritional interventions. For example, the n-6:n-3 PUFA ratio can be altered by increasing n-3 PUFA supplementation, but also by decreasing n-6 intake.
Several large studies have linked higher n-3 PUFA levels with lower all-cause mortality (Lee et al., 2009; Pottala et al., 2010) including a large 3.5 year trial (Marchioli et al., 2002). The n-3 PUFA’s anti-inflammatory and antioxidant properties provide one obvious pathway for these reductions in mortality, consistent with the finding that decreases in IL-6 were associated with telomere lengthening in this study.
Our data suggest that the n-3 PUFAs can impact cell aging in addition to inflammation and oxidative stress. This translational research broadens our understanding of the n-3 PUFA’s potential therapeutic effects.
Short telomeres predict early disease, and
slowing immune cell aging could have broad
effects by slowing the onset of age-related diseases. Recent work has demonstrated the causal effect of telomerase deficiency and telomere shortening on cellular health and premature aging and mortality in rodents (Bernardes de Jesus et al., 2012; Jaskelioff et al., 2011; Sahin et al., 2011). In summary, the current study provides compelling initial evidence that lower n-6:n-3 PUFA ratios may be beneficial for slowing biological aging.
Conflict of interest statementDrs. Blackburn, Epel, and Lin are co-founders in Telome Health, Inc., a telomere measurement company.
_______________________ Previous paper with other results from the same study:
- Kiecolt-Glaser JK
et al. Omega-3 supplementation lowers inflammation in healthy middle-aged and older adults: a randomized controlled trial. Brain Behav Immun (2012) vol. 26 (6) pp. 988-95ncbi.nlm.nih.gov/pubmed/22640930 linkinghub.elsevier.com/retrieve/pii/S0889-1591(12)00118-3 sciencedirect.com/science/article/pii/S0889159112001183 Excerpts of interest:
A number of epidemiological and observational studies have demonstrated that lower n-3 PUFA levels are associated with higher serum IL-6, TNF-a, and CRP (Farzaneh-Far et al., 2009; Ferrucci et al., 2006; Kalogeropoulos et al., 2010; Kiecolt-Glaser et al., 2007). In contrast, comparisons of supplemented and placebo groups in n-3 PUFA randomized controlled trials (RCTs), the gold standard for demonstrating causality, have not produced reliable serum cytokine differences (Calder et al., 2009; Fritsche, 2006; Kiecolt-Glaser et al., 2011; Sijben and Calder, 2007). Problematic methodological issues that muddy interpretation have included severely underpowered small treatment groups (e.g., 8–10 per group), low n–3 PUFA supplementation doses, insensitive cytokine assays, use of young and healthy subjects and/or highly-trained athletes, and very low levels of baseline inflammation. For example, serum cytokines did not differ significantly among 58 monks who received 0, 1.06, 2.13 or 3.19 g/d of n-3 PUFAs for a year (Blok et al., 1997); however, basal cytokine data did not differ between vegetarians and non-vegetarians even before supplementation, suggesting that the monks’ extremely healthy lifestyle limited the ability to see meaningful downward change.
The strongest RCT support for the n-3 PUFA’s anti-inflammatory properties has come from studies with older, hypertriglyceridemic or diabetic individuals with elevated inflammatory markers (Fritsche, 2006; Sijben and Calder, 2007; Wu, 2004; Yusof et al., 2008). Consequently, it has been suggested that cytokine produc- tion in healthy people is relatively insensitive to long-chain n-3 PUFAs (Sijben and Calder, 2007; Wu, 2004).
2.1. ParticipantsThe 138 participants, 45 men and 93 women, ranged in age from 40 to 85 (Table 1). Campus and community print and web-based announcements were used for recruitment. The institutional review board approved this study, and each participant provided informed consent.
The online screening form assessed health history, medications, and health behaviors.
Exclusions included psychoactive drugs or mood altering medications, lipid-altering drugs, beta blockers, steroids,
regular use of non-steroidal anti-inflammatory drugs other than an aspirin a day, ACE-inhibitors, prostaglandin inhibitors, heparin, warfarin, and alcohol/drug abuse (Buckley et al., 2004; Ferrucci et al., 2006). We also excluded pregnant or nursing women, vegetarians, diabetics, people who routinely took fish oil or flaxseed supplements or ate more than two portions of oily fish per week, smokers, and individuals with recurrent digestive problems, convulsive disorders, and autoimmune and/or inflammatory diseases. Individuals who typically engaged in 2 or more hours of vigorous physical activity per week, as well as individuals with a body mass index (BMI) less than 22.5 or greater than 40 were excluded (Fernandez-Real et al., 2003).
In addition, we used participants’ ability to follow the regimen as a criterion for study entry. Participants received a 7-day supply of placebo capsules (single blind) at the subsequent in-person screening session, and those who had taken less than 80% of the capsules a week later were dropped before randomization. We also verified height and weight at the screening visit.
2.2.1. Supplement and placeboThis three-arm parallel group RCT compared responses to (A) 2.496 g/d n-3, (B) 1.25 g/d n-3, and placebo, or (C) placebo. All participants took 6 pills (3 g oil) per day. For the two omega-3 groups, each 500 mg gel capsule contained 347.5 mg eicosapentaenoic acid (EPA) and 58 mg docosahexaenoic acid (DHA). Thus, for the high dose group the full daily supplement would equal 2085 mg/d of EPA and 348 mg/d of DHA. We chose the 7:1 EPA/DHA balance because of evidence that EPA has relatively stronger anti-inflammatory and antidepressant effects than DHA (Lin et al., 2010; Sijben and Calder, 2007). The placebo was a mixture of palm, olive, soy, canola, and coco butter oils that approximated the saturated:monounsaturated:polyunsaturated (SMP) ratio consumed by US adults, 37:42: 21 (USDA Continuing Survey of Food Intake by Individuals, 1994–1996). OmegaBrite (Waltham, MA) supplied both the n-3 and the matching placebo; all pills were coated with a fuchsia coloring. OmegaBrite
added a mild fish flavor to the placebo to help disguise any differences between the n-3 PUFA pills and the placebo, and we told participants about the fish flavoring to promote blindness (Stoll et al., 2001).
2.3. Health-related behaviorsParticipants’ height and weight were assessed at the screening visit, and participants were weighed during each subsequent visit. At each study visit, participants were evaluated for changes in fatty acid composition of plasma and PBMCs, mood, and proinflammatory cytokines.
Adipose tissue in the abdomen may secrete up to three times as much IL-6 as other subcutaneous fat tissues (Browning, 2003). Sagittal abdominal diameter measurements provided data on abdominal fat.
2.7. Sample sizeSample size was based on detection of conservative effect sizes for the lower dose versus placebo comparisons for the primary outcome of cytokine levels.
The literature suggested that the higher dose would have a greater effect, thus we expected the higher dose versus placebo contrast to have more power than low versus pla- cebo. All power analyses were based on contrasts within mixed effect linear models with two-sided alpha = 0.05 and assumed a 10% attrition rate. Our conservative estimated effect size was a decrease in IL-6 of 0.45 pg/mL in the low dose group, extrapolated from results from Ferrucci et al. (2006). Pilot data from our lab provided an estimate of standard deviation of 0.88 pg/mL, thus to achieve 85% power a sample size of 46 in each group was required.
3.1. Study population, diet, and health behaviorsRandomized groups were equivalent on key dimensions (Table 1). Randomization produced groups that did not differ on age, baseline FFQ dietary variables, depression, and sleep quality, p > 0.2 for all tests. Using BMI cut points of 25 and 30 kg/m2, 125 (91%) were overweight, and 65 (47%) were obese, respectively. There were small differences between groups on weight at baseline (p=0.002), with the placebo group having the lowest average weight, however there were not significant differences in either sagittal abdominal diameter or BMI (p > 0.05 for both).
Analyses of FFQ data at the last visit revealed no differences among the groups in reported changes in intake of calories, fiber, total fat, protein, saturated fat, monounsaturated fat, polyunsaturated fat, omega-3 fatty acids, or linoleic acid during the study period, P > .11 for all tests. Similarly, sleep and exercise did not show differential group changes, P > .26 for both.
Both the lower and higher n-3 groups had modest but statistically significant increases in weight across the trial (average pounds gained: 2.1 lbs and 2.5 lbs in the two groups, respectively), compared to no change in the placebo group (average pounds gained: 0.39 lbs). However, the increased weight would have theoretically fueled inflammation in this overweight sample which was not observed.
Few participants reported taking any medication during the study, and the numbers did not differ among groups. The most common medications were multivitamins (n = 42), NSAIDs (n = 29;
15 aspirin, 8 ibuprofen, 5 naproxen, 1 meloxicam), antihistamines (n = 13), estrogen with or without progesterone (n = 10), and levothyroxine (n = 10).
3.4. Changes in fatty acidsBaseline levels of plasma fatty acids as well as changes over time for the three groups are summarized in Table 3. As expected, randomization produced groups that did not differ on EPA (p = 0.73), DHA (p = 0.38), or total n-3 (p = 0.41) at baseline. By the end of the study period plasma levels of
EPA were approximately 3.5-fold higher in the 1.25 g/d n-3 group and 6-fold higher in the 2.5 g/d n-3 group (P < 0.0001 for both), and plasma
DHA levels were approximately 1.4-fold higher in the 1.25 g/d n-3 group and 1.5-fold higher in the 2.5 g/d n-3 group (P < 0.0001 for both). The n-6:n-3 ratio was significantly decreased after supplementa- tion for both low and high dose groups (P < 0.0001 for both).
3.5. Primary OutcomesResults for inflammatory outcomes and depression symptoms are summarized in Table 5. After adjusting for gender and sagittal abdominal diameter, there were significant supplementation effects on cytokines as evidenced by significant group by visit inter- actions for both TNF-α (p = 0.0002) and IL-6 (p = 0.0003). The estimated mean change in log-TNF-α from visit 1 to visit 5 was 0.11 units for the
placebo group, corresponding to a
12% increase in the geometric mean of TNF-α. In comparison, the estimated
mean change in log-TNF-α was *0.0002 units for the 1.25 g/d and -0.024 for the 2.5 g/d group, corresponding to
changes of 0.2% and -2.3%, respectively. After Bonferroni-adjustment, these group differences were
significant for the comparison of placebo to 1.25 g/d (p = 0.03) and placebo to 2.5 g/d (p = 0.004);
no significant difference was noted between the two supplementation doses (p = 1.0).
A similar pattern was observed for IL-6. The estimated mean change in log-IL-6 from visit 1 to visit 5 was 0.31 units for the
placebo group (36% increase), -0.106 for the 1.25 g/d group (10% decrease), and -0.123 for the 2.5g/d group (12% decrease). Significant differences were observed
between placebo and 1.25 g/d (p = 0.0003)
and placebo and 2.5 g/d (p = 0.0002), but
not between the two doses of fish oil (p = 1.0). To ensure that results were not driven by a small number of highly influential data points, residual plots were examined and one subject in the placebo group who appeared to be an outlier was removed and analyses were rerun. Resulting conclusions were the same and are not shown.
There did not appear to be group effects on depression (p = 0.86), adjusting for gender. There was a trend toward larger decreases in depression from visit 1 to visit 5 for the two fish oil groups than the placebo, but no differences were statistically significant.
4.1. Intervention-related reductions in inflammationOmega-3 supplementation significantly altered production of serum cytokines. IL-6 decreased by 10% and 12% in our low and high dose n-3 groups, respectively, compared to a 36% increase in the placebo group. Similarly, low and high dose n-3 groups showed modest 0.2% and -2.3% changes in TNF-α, compared to a 12% increase in the control group. This is the first well-powered trial to show significant changes in serum cytokines in healthy middle-aged and older adults.
4.2. Randomized PUFA trialsThe largely negative serum cytokine data from prior n-3 PUFA trials have led to the suggestion that cytokine production is relatively insensitive to the n-3 PUFAs among healthy individuals, i.e., people who do not have chronic inflammatory diseases such as inflammatory bowel disease, rheumatoid arthritis, chronic obstructive pulmonary disease, or diabetes (Sijben and Calder, 2007; Wu, 2004). However, there are several notable differences between our study and prior RCTs. We carefully assessed
variables known to influence inflammation including
smoking, medication use, physical activity, and abdominal adiposity. We had minimal attrition; only 5 of our 138 subjects failed to complete the full trial. Our rigorous exclusion criteria produced a
group of overweight sedentary adults who were more likely to have an inflammatory profile and who were otherwise
healthy aside from their weight.[ . . . ]
In
prior work from our laboratory, 68 medical students received either 2.5 g/d n-3 or a placebo for 12 weeks (Kiecolt-Glaser et al., 2011). Compared to controls, those students who received n-3 showed a 14% decrease in lipopolysaccharide (LPS) stimulated IL- 6 production. Planned secondary analyses that used the plasma n-6:n-3 ratio in place of
treatment group showed that decreasing n-6:n-3 ratios led to
reductions in stimulated IL-6 and TNF-α production, as well as
marginal differences in serum TNF-α. The
absence of significant serum inflammatory changes was likely related to the
very low baseline levels of serum cytokines
in the healthy, young, and relatively thin population.
4.3. Dosage and risksNeither our IL-6 nor our TNF-α data showed significant differences between our 1.25 and 2.5 g/d n-3 dose, although both clearly differed from the placebo. One review concluded that while the effects were inconsistent, it appeared that significant changes in cytokine production by lymphocytes only occurred with P2.0 g/d of EPA + DHA (Sijben and Calder, 2007). In addition, variables such as typical dietary intake influence responses (Yee et al., 2010), and our sample had a higher than expected average n-6:n-3 ratio at baseline, as described earlier. The FDA has concluded that intakes of
up to 3 g/d of marine n-3 PUFAs are
‘‘Generally Recognized As Safe’’ (Kris-Etherton et al., 2002); our higher dose,
2.5 g/d, fell within that range and would appear to be
a good choice for future studies.Side effects were infrequent and did not differ between groups. These data are in accord with the low incidence reported in large n-3 PUFA studies (Leaf et al., 1994; Valagussa et al., 1999).
4.4. Health implicationsSeveral large studies have linked higher n-3 PUFA levels with lower all-cause mortality (Lee et al., 2009; Pottala et al., 2010), including a large 3.5 year trial (Marchioli et al., 2002). The n-3 PUFA’s anti-inflammatory properties provide one obvious pathway for these reductions in mortality. Inflammation is a robust and reli- able predictor of all-cause mortality in older adults (Pedersen and Febbraio, 2008). Chronic inflammation has been linked to a spec- trum of health problems including depression, cardiovascular disease, osteoporosis, cancer, and arthritis (Pedersen and Febbraio, 2008). In fact, more globally, chronic inflammation has been suggested as one key biological mechanism that may fuel declines in physical function leading to frailty, disability, and, ultimately, death.
_______________________ Related paper of a previous study by the same group:
- Kiecolt-Glaser JK
et al. Omega-3 supplementation lowers inflammation and anxiety in medical students: a randomized controlled trial. Brain Behav Immun (2011) vol. 25 (8) pp. 1725-34ncbi.nlm.nih.gov/pubmed/21784145 linkinghub.elsevier.com/retrieve/pii/S0889-1591(11)00468-5 sciencedirect.com/science/article/pii/S0889159111004685 _______________________ Related information available at MedLine Plus:
nlm.nih.gov - Fish OilRetrieved on October 3, 2012
nlm.nih.gov/medlineplus/druginfo/natural/993.html Excerpt of interest:
How effective is it?Natural Medicines Comprehensive Database rates effectiveness based on scientific evidence according to the following scale: Effective, Likely Effective, Possibly Effective, Possibly Ineffective, Likely Ineffective, Ineffective, and Insufficient Evidence to Rate.
The effectiveness ratings for FISH OIL are as follows:
Effective for...-
High triglycerides. High triglycerides are associated with heart disease and untreated diabetes. To reduce the risk of heart disease, doctors believe it is important to keep triglycerides below a certain level. Doctors usually recommend increasing physical activity and restricting dietary fat to lower triglycerides. Sometimes they also prescribe drugs such as gemfibrozil (Lopid) for use in addition to these lifestyle changes. Now researchers believe that fish oil, though not as effective as gemfibrozil, can reduce triglyceride levels by 20% to 50%. One particular fish oil supplement called Lovaza has been approved by the FDA to lower triglycerides. Lovaza contains 465 milligrams of EP and 375 milligrams of DHA in 1-gram capsules.
Likely effective for...-
Heart disease. Research suggests that consuming fish oil by eating fish can be effective for keeping people with healthy hearts free of heart disease. People who already have heart disease might also be able to lower their risk of dying from heart disease by eating fish or taking a fish oil supplement. However, for people who already take heart medications such as a “statin,” adding on fish oil might not offer any additional benefit.
Possibly effective for...-
High blood pressure. Fish oil seems to produce modest reductions in blood pressure in people with high blood pressure. The omega-3 fatty acids in fish oil seem to be able to expand blood vessels, and this brings blood pressure down.
-
Rheumatoid arthritis. Fish oil alone, or in combination with the drug naproxen (Naprosyn), seems to help people with rheumatoid arthritis get over morning stiffness faster. People who take fish oil can sometimes reduce their use of pain medications such as nonsteroidal anti-inflammatory drugs (NSAIDs).
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Menstrual pain (dysmenorrhea). Taking fish oil alone or in combination with vitamin B12 seems to improve painful periods and reduce the need for pain medications such as nonsteroidal anti-inflammatory drugs (NSAIDs).
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Attention deficit-hyperactivity disorder (ADHD) in children. Taking fish oil seems to improve thinking skills and behavior in 8 to 12 year-old children with ADHD.
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Raynaud’s syndrome. There’s some evidence that taking fish oil can improve cold tolerance in some people with the usual form of Raynaud’s syndrome. But people with Raynaud’s syndrome caused by a condition called progressive systemic sclerosis don’t seem to benefit from fish oil supplements.
-
Stroke. Moderate fish consumption (once or twice a week) seems to lower the risk of having a stroke by as much as 27%. However, eating fish doesn’t lower stroke risk in people who are already taking aspirin for prevention. On the other hand, very high fish consumption (more than 46 grams of fish per day) seems to increase stroke risk, perhaps even double it.
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Weak bones (osteoporosis). Taking fish oil alone or in combination with calcium and evening primrose oil seems to slow bone loss rate and increase bone density at the thigh bone (femur) and spine in elderly people with osteoporosis.
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Hardening of the arteries (atherosclerosis). Fish oil seems to slow or slightly reverse the progress of atherosclerosis in the arteries serving the heart (coronary arteries), but not in the arteries that bring blood up the neck to the head (carotid arteries).
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Kidney problems. Long-term use (two years) of fish oil 4-8 grams daily can slow the loss of kidney function in high-risk patients with a kidney disease called IgA nephropathy. Fish oil also seems to reduce the amount of protein in the urine of people who have kidney disease as a result of diabetes.
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Bipolar disorder. Taking fish oil with the usual treatments for bipolar disorder seems to improve symptoms of depression and increase the length of time between episodes of depression. But fish oil doesn’t seem to improve manic symptoms in people with bipolar disorder.
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Psychosis. Taking a fish oil supplement might help prevent full psychotic illness from developing in people with mild symptoms. This has only been tested in teenagers and adults up to age 25.
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Weight loss. Some evidence shows that eating fish improves weight loss and decreases blood sugar in overweight people and people with high blood pressure. Preliminary research also shows that taking a specific fish oil supplement 6 grams daily (Hi-DHA, NuMega), providing 260 mg DHA/gram and 60 mg EPA/gram, significantly decreases body fat when combined with exercise.
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Endometrial cancer. There is some evidence that women who regularly eat about two servings of fatty fish per week have a reduced risk of developing endometrial cancer.
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Age-related eye disease (age-related macular degeneration, AMD). There is some evidence that people who eat fish more than once per week have a lower risk of developing age-related macular degeneration.
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Reducing the risk of blood vessel re-blockage after heart bypass surgery or “balloon” catheterization (balloon angioplasty). Fish oil appears to decrease the rate of re-blockage up to 26% when given for one month before the procedure and continued for one month thereafter. Apparently, taking fish oil before surgery is important. When taken for less than one month before angioplasty, fish oil doesn’t help protect the blood vessel against closing down.
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Recurrent miscarriage in pregnant women with antiphospholipid syndrome. Taking fish oil seems to prevent miscarriage and increase live birth rate in pregnant women with a condition called antiphospholipid syndrome.
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High blood pressure and kidney problems after heart transplant. Taking fish oil seems to preserve kidney function and reduce the long-term continuous rise in blood pressure after heart transplantation.
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Damage to the kidneys and high blood pressure caused by taking a drug called cyclosporine. Cyclosporine is a medication that reduces the chance of organ rejection after an organ transplant. Fish oil might help reduce some of the unwanted side effects of treatment with this drug.
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Movement disorder in children (dyspraxia). Taking fish oil orally, in combination with evening primrose oil, thyme oil, and vitamin E (Efalex, Efamol Ltd), seems to improve movement disorders in children with dyspraxia.
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Developmental coordination disorder. A combination of fish oil (80%) and evening primrose oil (20%) seems to improve reading, spelling, and behavior when given to children age 5-12 years with developmental coordination disorder. However, it doesn’t seem to improve motor skills.
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Preventing blockage of grafts used in kidney dialysis. Taking fish oil orally seems to help prevent clot formation in hemodialysis grafts.
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Psoriasis. There is some evidence that administering fish oil intravenously (by IV) can decrease severe psoriasis symptoms. But taking fish oil by mouth doesn’t seem to have any effect on psoriasis.
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High cholesterol. There is interest in using fish oil in combination with “statin” drugs for some people with high cholesterol. Doctors were worried at first that taking fish oil might interfere with statin treatment, but early studies show this is not a problem, at least with the statin called simvastatin. Scientists think fish oil may lower cholesterol by keeping it from being absorbed in the intestine. There is some evidence that using vitamin B12 along with fish oil might boost their ability to lower cholesterol.
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Coronary artery bypass surgery. Taking fish oil seems to prevent coronary artery bypass grafts from re-closing following coronary artery bypass surgery.
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Cancer-related weight loss. Taking a high dose (7.5 grams per day) of fish oil seems to slow weight loss in some cancer patients. Some researchers believe these patients eat more because the fish oil is fighting depression and improving their mood.
-
Asthma. Some research suggests fish oil may lower the occurrence of asthma in infants and children when taken by women late in pregnancy. Furthermore, fish oil seems to improve airflow, reduce cough, and lower the need for medications in some children with asthma. However, fish oil treatment doesn’t seem to provide the same benefit for adults.
Possibly ineffective for...-
Chest pain (angina).-
Gum infection (gingivitis).-
Liver disease.-
Leg pain due to blood flow problems (claudication).-
Preventing migraine headaches.-
Preventing muscle soreness caused by physical exercise.-
Breast pain.-
Skin rashes caused by allergic reactions.-
Stomach ulcers.Likely ineffective for...-
Type 2 diabetes. Taking fish oil doesn’t seem to lower blood sugar in people with type 2 diabetes. However, fish oil can provide some other benefits for people with diabetes, such as lowering blood fats called triglycerides.
[ . . . ]
nlm.nih.gov/medlineplus/druginfo/natural/993.html#Effectiveness _______________________ Links to other science news outlets:
sciencedaily.com/releases/2012/10/121001140957.htm medicalxpress.com/news/2012-10-omega-supplements-biological-effect-aging.html esciencenews.com/articles/2012/10/01/omega.3.supplements.may.slow.a.biological.effect.aging scienceblog.com/56909/omega-3-supplements-may-slow-a-biological-effect-of-aging _______________________ URL related G+ posts:
plus.google.com/102370347732140106252/posts/C9p4AJG2xtY plus.google.com/105903603302602842440/posts/fuEu9ixJx7S plus.google.com/117029437254252483108/posts/7BvUQpWDxx9 _______________________