Higher Omega-3, but Especially DHA, Preserve Bone in Ovariectomized Rats

Abstract

Dietary ratio of n-6/n-3 PUFAs and docosahexaenoic acid: actions on bone mineral and serum biomarkers in ovariectomized rats.

Hypoestrogenic states escalate bone loss in animals and humans. This study evaluated the effects of the amount and ratio of dietary n-6 and n-3 polyunsaturated fatty acids (PUFAs) on bone mineral in 3-month-old sexually mature ovariectomized (OVX) Sprague-Dawley rats. For 12 weeks, the rats were fed either a high-PUFA (HP) or a low-PUFA (LP) diet with a ratio of n-6/n-3 PUFAs of 5:1 (HP5 and LP5) or 10:1 (HP10 and LP10). All diets (modified AIN-93G) provided 110.4 g/kg of fat from safflower oil and/or high-oleate safflower oil blended with n-3 PUFAs (DHASCO oil) as a source of docosahexaenoic acid (DHA). Fatty acid analyses confirmed that the dietary ratio of 5:1 significantly elevated the amount of DHA in the periosteum, marrow and cortical and trabecular bones of the femur. Dual-energy X-ray absorptiometry measurements for femur and tibia bone mineral content (BMC) and bone mineral density showed that the DHA-rich diets (HP5 and LP5) resulted in a significantly lower bone loss among the OVX rats at 12 weeks. Rats fed the LP diets displayed the lowest overall serum concentrations of the bone resorption biomarkers pyridinoline (Pyd) and deoxypyridinoline, whereas the bone formation marker osteocalcin was lowest in the HP groups. Regardless of the dietary PUFA content, DHA in the 5:1 diets (HP5 and LP5) preserved rat femur BMC in the absence of estrogen. This study indicates that the dietary ratio of n-6/n-3 PUFAs (LP5 and HP5) and bone tissue concentration of total long-chain n-3 PUFAs (DHA) minimize femur bone loss as evidenced by a higher BMC in OVX rats. These findings show that dietary DHA lowers the ratio of 18:2n-6 (linoleic acid)/n-3 in bone compartments and that this ratio in tissue correlates with reduced Pyd but higher bone alkaline phosphatase activity and BMC values that favor bone conservation in OVX rats.

Watkins BA, Li Y, Seifert MF
J. Nutr. Biochem. Apr 2006
PMID: 16102959

Fish Oil, Especially DHA, Increases Bone Density in Rats

Abstract

Is docosahexaenoic acid more effective than eicosapentaenoic acid for increasing calcium bioavailability?

Experimental animal and human studies have indicated that long chain polyunsaturated fatty acids (LCPUFA) may enhance calcium absorption, reduce urinary calcium excretion, and increase bone calcium content. In the present study, the effect of LCPUFA, as provided in evening primrose oil, fish and tuna oils, on calcium bioavailability was investigated. Growing male rats were fed a semi-synthetic diet for 6 weeks, after which calcium absorption, bone mineral density (ex vivo), bone calcium content, and bone biomechanics were measured. Calcium absorption, ex vivo bone mineral density, and bone calcium content were significantly higher in the animals fed tuna oil compared with those of a control group fed corn oil. Significant correlations were found between the docosahexaenoic acid (DHA) (22:6n-3) content of the red cell membranes and bone density and bone calcium content. DHA increased accretion of calcium in bone significantly more so than eicosapentaenoic acid (EPA) (20:5n-3).

Kruger MC, Schollum LM
Prostaglandins Leukot. Essent. Fatty Acids Nov 2005
PMID: 16154334

Fish Oil Reduces Resorption in Rats with Periodontal Disease

Abstract

Omega-3 fatty acid effect on alveolar bone loss in rats.

Gingival inflammation and alveolar bone resorption are hallmarks of adult periodontitis, elicited in response to oral micro-organisms such as Porphyromonas gingivalis. We hypothesized that omega (omega)-3 fatty acids (FA) dietary supplementation would modulate inflammatory reactions leading to periodontal disease in infected rats. Rats were fed fish oil (omega-3 FA) or corn oil (n-6 FA) diets for 22 weeks and were infected with P. gingivalis. Rats on the omega-3 FA diet exhibited elevated serum levels of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), documenting diet-induced changes. PCR analyses demonstrated that rats were orally colonized by P. gingivalis; increased IgG antibody levels substantiated this infection. P. gingivalis-infected rats treated with omega-3 FA had significantly less alveolar bone resorption. These results demonstrated the effectiveness of an omega-3 FA-supplemented diet in modulating alveolar bone resorption following P. gingivalis infection, and supported that omega-3 FA may be a useful adjunct in the treatment of periodontal disease.

Kesavalu L, Vasudevan B, Raghu B, Browning E…
J. Dent. Res. Jul 2006
PMID: 16798867

Statins and Fish Oil Improve Lipid in Bones

Abstract

Statins and dietary fish oils improve lipid composition in bone marrow and joints.

There have been numerous efforts to alter the lipid content of cardiovascular tissues. Although equally important, only limited information is available about musculoskeletal tissues. I characterized joint and bone marrow lipids in patients having joint replacement surgery and explored the effects of fish oils and statins on lipid composition in bone marrow and joint fluid. Joint drainage catheters were used to collect marrow lipids from 84 patients having 128 hip and knee replacements for osteoarthritis, osteonecrosis, and femoral neck fractures (osteoporosis). Statins reduced the amount of lipid by 22% in patients with osteoporosis, 26% in patients with osteoarthritis, and 41% in patients with osteonecrosis compared with pretreatment lipid levels in the same patients. Taking fish oils reduced the amount of lipid in bone marrow by 20%. Lipid profiles of disturbed marrow and joint fluid from patients who took statins or dietary fish oil showed an increase in the proportion of unsaturated fatty acids and longer-chain fatty acids relative to pretreatment profiles. The ability to change the amount and character of bone and joint lipids may have major importance for strengthening bone, reducing the severity or preventing osteonecrosis, and enhancing joint lubrication.

Pritchett JW
Clin. Orthop. Relat. Res. Mar 2007
PMID: 17496750

DHA More Potent Inhibitor of Osteoclasts than EPA

Abstract

Docosahexaenoic acid is more potent inhibitor of osteoclast differentiation in RAW 264.7 cells than eicosapentaenoic acid.

Fish oil rich in n-3 polyunsaturated fatty acids, especially eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) protects inflammation induced bone loss in chronic inflammatory diseases like rheumatoid arthritis, periodontitis, and osteoporosis. EPA and DHA differentially regulate functional parameters and gene expression in different cell types. One of the risk factors for bone loss in inflammatory bone diseases is the elevation of bone-resorbing osteoclasts and a very few studies so far have indicated that attenuation of osteoclastogenesis might be one of the mechanisms by which n-3 PUFA exert its effect on bone loss protection. However, the precise mechanism underlying this process remains unclear. Receptor activator of NF-kappaB ligand (RANKL) is known to be the most critical mediator of osteoclastogenesis. Therefore, in this study, we examined the differential effect of EPA and DHA on RANKL-stimulated osteoclastogenesis and RANKL signaling using a murine monocytic cell line RAW 264.7. DHA was found to inhibit osteoclast differentiation, activation and function more potently than EPA. The differential potential also closely correlated with the inhibition of osteoclast-specific genes like tartrate resistant acid phosphatase, cathepsin K, calcitonin receptor, matrix metalloproteinase-9 expression and osteoclast-specific transcription factor, c-Fos, as well as osteotropic proinflammatory cytokine, TNF-alpha to a greater extent with DHA than EPA. Further, pretreatment of RAW 264.7 cells with DHA also showed significantly reduced activation of NF-kappaB and p38MAPK than EPA. Our findings suggest that DHA may be much more effective than EPA in alleviating RANKL induced proinflammatory cytokine production, intracellular signaling activation, thereby decreasing osteoclast activation and bone resorption.

Rahman MM, Bhattacharya A, Fernandes G
J. Cell. Physiol. Jan 2008
PMID: 17929247

DHA prevents, and GLA Exacerbates, Bone Loss in Ovariectomized Rats

Abstract

Specific effects of gamma-linolenic, eicosapentaenoic, and docosahexaenoic ethyl esters on bone post-ovariectomy in rats.

Long chain polyunsaturated fatty acids (LCPUFAs) are involved in the regulation of bone metabolism. Increased dietary consumption of n-3, and possibly some n-6, LCPUFAs may limit postmenopausal bone loss. The aim of this study was to determine the effects on bone of specific fatty acids within the n-3 and n-6 LCPUFA families in ovariectomized (OVX) rats. Rats were OVX or sham-operated and fed either a control diet (OVX and sham) or a diet supplemented with 0.5 g/kg body weight/day of gamma-linolenic (GLA), eicosapentaenoic (EPA), docosahexaenoic (DHA) ethyl esters or a mixture of all three (MIX) for 16 weeks. Bone mineral content (BMC), area, and density and plasma concentrations of insulin-like growth factor-I, vitamin D, selected biochemical markers of bone metabolism, and parathyroid hormone (PTH) were determined. The OVX-induced decrease in lumbar spine BMC was significantly attenuated by DHA but not by EPA or GLA supplementation or supplementation with a mixture of all three LCPUFAs. Endosteal circumferences of tibiae were significantly greater in DHA and EPA compared to OVX. Plasma C-terminal telopeptide of type I collagen and osteocalcin concentrations were not significantly different in the DHA group compared to OVX. Femur BMC decreased by a significantly greater amount in GLA than OVX, and final plasma PTH concentrations were significantly higher in GLA compared to all other groups. In conclusion, DHA ameliorated OVX-induced bone mineral loss. GLA exacerbated post-OVX bone mineral loss, possibly as a result of PTH-induced bone catabolism.

Poulsen RC, Firth EC, Rogers CW, Moughan PJ…
Calcif. Tissue Int. Dec 2007
PMID: 18008020

Japanese Apricot Shows Potential to Prevent Osteoporosis

Abstract

A Prunus mume extract stimulated the proliferation and differentiation of osteoblastic MC3T3-E1 cells.

Osteoporosis is a serious disease caused by decreased bone mass. There is constant matrix remodeling in bones, by which bone formation is performed by osteoblastic cells, whereas bone resorption is accomplished by osteoclast cells. We investigated the effect of a Japanese apricot (Prunus mume SIBE. et ZUCC.) extract on the proliferation and osteoblastic differentiation in pre-osteoblastic MC3T3-E1 cells. An alkaline phosphatase (ALP) activity assay, cell proliferation assay, alizarin red staining and expression analysis of osteoblastic genes were carried out to assess the proliferation and osteoblastic differentiation. The water-soluble fraction of Prunus mume (PWF) increased the ALP activity, cell proliferation and mineralization. The gene expression of osteopontin and bone morphogenetic protein-2, which are markers in the early period of osteoblastic differentiation, were significantly enhanced by the PWF treatment. PWF therefore stimulated the proliferation and osteoblastic differentiation of cells and may have potential to prevent osteoporosis.

Kono R, Okuno Y, Inada K, Tokuda A…
Biosci. Biotechnol. Biochem. 2011
PMID: 21979066 | Free Full Text

High Cortisol is an Independent Predictor of Fractures in Older People

Abstract

The relation between cortisol excretion and fractures in healthy older people: results from the MacArthur studies-Mac.

In persons with depression, higher urinary cortisol is associated with lower bone mineral density.
To examine the relation between urinary free cortisol (UFC) and fractures.
Community-based samples from Durham, NC, East Boston, MA, and New Haven, CT.
684 men and women, aged 70 to 79 at baseline, who were part of the MacArthur Study of Successful Aging.
Cohort study. Participants with previous history of fractures at baseline were excluded.
The primary exposure variable was overnight (8:00 p.m. to 8:00 a.m.) UFC (microg/g creatinine) at baseline (1988). Outcomes were self-reported hip, arm, spine, wrist, or other fracture during the follow-up period (1988-1995). Covariates were baseline age, gender, race, body mass index, current physical activity, lower extremity strength, depression subscale of the Hopkins Symptom Checklist, and current use of cigarettes and alcohol.
Logistic regression was used to predict the occurrence of incident fractures (1988-1995) as a function of quartiles of baseline UFC. Models were adjusted for age, gender, and race and were also multiply adjusted for the remaining covariates listed above. Gender-stratified models and models that excluded corticosteroid users were also run.
In multiply adjusted models, higher baseline levels of UFC were significantly associated with incident fractures. Odds of fracture (95% Confidence Intervals) for increasing quartiles of baseline UFC, multiply adjusted, were: 2.28 (.91, 5.77); 3.40 (1.33, 8.69); 5.38 (1.68, 17.21). Results were not materially influenced by exclusion of persons using corticosteroids.
Higher baseline UFC is an independent predictor of future fracture.

Greendale GA, Unger JB, Rowe JW, Seeman TE
J Am Geriatr Soc Jul 1999
PMID: 10404922

NAC Stimulates Osteoblastogenesis in Rats

Abstract

N-acetyl cysteine as an osteogenesis-enhancing molecule for bone regeneration.

Bone regeneration often requires cues from osteogenesis-inducing factors for successful outcome. N-acetyl cysteine (NAC), an anti-oxidant small molecule, possibly modulates osteoblastic differentiation. This study investigated the potential of NAC as an osteogenesis-enhancing molecule in vitro and in vivo. Various concentrations of NAC (0, 2.5, 5.0, and 10 mM) were added to rat bone marrow stromal cell or osteoblastic cell culture in media with or without dexamethasone. The results showed marked enhancement of alkaline phosphatase activity and mineralized matrix formation together with consistent upregulation of bone-related gene markers such as collagen I, osteopontin, and osteocalcin in the osteoblastic culture with addition of 2.5 or 5.0 mM NAC regardless of the presence of dexamethasone. Micro-CT-based analysis and histological observation revealed that addition of NAC to a collagenous sponge implanted in a critical size cortical bone defect (3.0 mm × 5.0 mm) in rat femur yielded acceleration and completion of defect closure, with thick, compact, and contiguous bone after 6 weeks of healing. In contrast, with sponge alone, only sparse and incomplete bone regeneration was observed during the matching healing period. These results indicate that NAC can function as an osteogenesis-enhancing molecule to accelerate bone regeneration by activating differentiation of osteogenic lineages.

Yamada M, Tsukimura N, Ikeda T, Sugita Y…
Biomaterials Aug 2013
PMID: 23711675

NAC Prevents Skeletal Aging in Transgenic Mice

Abstract

Swedish mutant APP suppresses osteoblast differentiation and causes osteoporotic deficit, which are ameliorated by N-acetyl-L-cysteine.

Reduced bone mineral density and hip fracture are frequently observed in patients with Alzheimer’s disease (AD). However, mechanisms underlying their association remain poorly understood. Amyloid precursor protein (APP) is a transmembrane protein that is ubiquitously expressed in bone marrow stromal cells (BMSCs), osteoblasts (OBs), macrophages (BMMs), and osteoclasts (OCs). Mutations in the APP gene identified in early-onset AD patients are believed to cause AD. But little is known about APP’s role in bone remodeling. Here, we present evidence for Swedish mutant APP (APPswe) in suppression of OB differentiation and function in culture and in mouse. APP expression in BMSCs increases during aging. Ubiquitous expression of APPswe in young adult Tg2576 transgenic mice (under the control of a prion promoter) recaptured skeletal “aging-like” deficits, including decreased OB genesis and bone formation, increased adipogenesis and bone marrow fat, and enhanced OC genesis and bone resorption. Remarkably, selective expression of APPswe in mature OB-lineage cells in TgAPPswe-Ocn mice (under the control of osteocalcin [Ocn] promoter-driven Cre) also decreased OB genesis and increased OC formation, resulting in a trabecular bone loss. These results thus suggest a cell-autonomous role for APPswe in suppressing OB formation and function, but a nonautonomous effect on OC genesis. Notably, increased adipogenesis and elevated bone marrow fat were detected in young adult Tg2576 mice, but not in TgAPPswe-Ocn mice, implying that APPswe in BMSCs and/or multicell types in bone marrow promotes bone marrow adipogenesis. Intriguingly, the skeletal aging-like deficits in young adult Tg2576 mice were prevented by treatment with N-acetyl-L-cysteine (NAC), an antioxidant, suggesting that reactive oxygen species (ROS) may underlie APPswe-induced osteoporotic deficits. Taken together, these results demonstrate a role for APPswe in suppressing OB differentiation and bone formation, implicate APPswe as a detrimental factor for AD-associated osteoporotic deficit, and reveal a potential clinical value of NAC in the treatment of osteoporotic deficits.

Xia WF, Jung JU, Shun C, Xiong S…
J. Bone Miner. Res. Oct 2013
PMID: 23649480