Effect of omega-3 polyunsaturated fatty acids on the endocannabinoid system in osteoblast-like cells and muscle
Endogenous cannabinoids, anandamide (AEA) and 2-arachidonylglycerol (2-AG) are formed from the n-6 polyunsaturated fatty acid (PUFA), arachidonic acid (AA) and activate cannabinoid receptors CB1 and CB2. Dietary intake or cell treatments of n-3 PUFA have been shown to reduce AA, AEA and 2-AG levels in peripheral and central tissues and cells. The impact of n-3 PUFA on EC system mRNA levels in bone and skeletal muscle is unknown. Therefore, we hypothesized that n-3 PUFA decreases AA concentrations and changes mRNA levels of EC system receptors and enzymes that play a role in bone and muscle functions. To test the hypothesis we used an osteoblast-like cell model and an in vivo model of muscle disuse.^ The study findings revealed that eicosapentaenoic acid (EPA) enrichment of osteoblast-like cells promoted lower CB2 and n-acyl phosphatidylethanolamine selective phospholipase D (NAPE-PLD) mRNA levels. Osteoblast-like cells were treated with a CB2 antagonist/inverse agonist (AM630) prior to AA or EPA. The AM630 pre-treatment resulted in lower levels of docosahexaenoic acid (DHA) in the osteoblast compared to AA, EPA or vehicle control. CB2 antagonism followed by AEA treatment resulted in lower levels of all PUFA with 20 or 22C chain length and 4 or more double bonds when compared to AEA alone, a CB2 specific agonist (HU-308) or vehicle control.^ The mouse model showed in increases in EC system mRNA (n-acyl phosphatidylethanolamine selective phospholipase D [NAPE-PLD], and diacylglycerol lipase [DAGL]) for the synthesis enzymes and cannabinoid receptors in quadriceps muscle as a result of high n-3 PUFA in the diet. AA levels were lower as a result of n-3 PUFA intake as was the ratio of n-6/n-3 PUFA in the quadriceps muscle. The quadriceps muscle atrophied from hind limb suspension compared to the weight bearing mice and this tissue was used to evaluate the effect of muscle disuse atrophy on mRNA levels of the EC system. Hind limb suspension resulted in greater mRNA levels for AEA and 2-AG synthesis enzymes and CB2 levels compared to weight bearing mice. The high n-3 PUFA diet also influenced EC system mRNA levels. Quadriceps muscle from the high n-3 PUFA fed mice had greater levels of mRNA for EC synthesis enzymes and both CB1 and CB2 receptors compared to quadriceps muscle from mice in the control fed group. EC substrate availability and AA levels were decreased with n-3 PUFA intake. The increase in EC synthesis enzymes and cannabinoid receptors that occurred in response to hind limb suspension and the high n-3 PUFA appear to be involved in the metabolic and physiological effects of hind limb suspension.^ The two models used in this research, osteoblast-like cell cultures and mouse hind limb suspension, demonstrate that the EC system in bone and muscle is responsive to dietary and physiological changes. These new findings justify the importance of diet and the endocannabinoid system in musculoskeletal health. ^
Bruce A. Watkins, Purdue University.
Health Sciences, Nutrition
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