Progress 03/03/98 to 02/28/04
Outputs
Our earlier observations that dietary conjugated linoleic acid (trans-10, cis-12; CLA) causes body fat depletion in mice was extended to include evidence that this action of CLA depends upon composition of dietary fat. Mice fed a diet of 7% coconut oil were much more sensitive to CLA-induced body fat depletion than were mice fed a diet of 7% soy oil. This may be explained by either: a) an interaction of essential fatty acids with CLA; or b) a metabolic difference in mice fed saturated fats versus polyunsaturated fats. We conducted several experiments to distinguish between these competing hypotheses. In summary, although the severity of an essential fatty acid deficiency depends upon the duration of consuming essential fatty acid-deficient diets, the interaction with CLA does not. Furthermore, replenishment of individual essential fatty acids (linoleate, linolenate, or arachidonate) does not diminish the interation of coconut oil with CLA. It appears, in contrast, that
mice that are metabolizing mostly saturated fatty acids (either from dietary coconut oil or from de novo synthesis), are particularly sensitive to the body fat depleting effects of CLA, regardless of essential fatty acid intake.
Impacts
The immediate impact is that our results are interpreted to indicate the use of CLA as a human food supplement should not be endorsed unless the potential effect of CLA on insulin resistance in humans is ruled out. A longer term impact will be an improved understanding of mechanisms determining fat deposition which should ultimately enable applications for manipulation of body fatness.
Publications
- Hargrave,K.M., B.J. Meyer, C. Li, M.J. Azain, C.A. Baile, J.L. Miner. 2004. Influence of Conjugated Linoleic Acid and Fat Source on Body Fat and Apoptosis in Mice. Obes. Res. 12(9):1435-1444.
- Hargrave, K.M., D. Pomp, and J.L. Miner. 2005. Effect of dietary conjugated linoleic acid on adiposity and the adipose-transcriptome. J. Anim. Sci. (Natl. Meeting).
- Behlke, K., E. Behlke, P. Robinson, J. Takacs, R. Dumitru, S. Ragsdale, P. Newsome, and J. Miner. 2005. Inhibition of methanogenesis in free living vs. protozoa-associated ruminal methanogens. J. Anim. Sci. (Midwest Section).
- Hadenfeldt, T.J., K.M. Hargrave, and J.L. Miner. 2005. The interaction of dietary CLA and fat source on triglyceride turnover in adipose tissue of mice. J. Anim. Sci. (Midwest Section).
- Hargrave, K.M., M.J. Azain, M.G. Obukowicz, and J.L. Miner. 2005. Effect of conjugated linoleic acid and/or a specific delta6-desaturase inhibitor on body composition of mice. J. Anim. Sci. (Midwest Section).
- Hargrave, K.M., T.J. Hadenfeldt, M.J. Azain, and J.L. Miner. 2005. Coconut oil and fat free diets enhance conjugated linoleic acid-induced lipolysis and body fat loss in mice. Faseb J.
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Progress 10/01/02 to 09/30/03
Outputs
The mechanism by which dietary conjugated linoleic acid causes depletion of body fat in mice has been investigated. We have found that the trans10, cis12, conjugated linoleic acid isomer, specifically, can cause significant loss of body fat within 5 days of inclusion in the diet at .5% by weight; DNA fragmentation in adipose tissue is caused by this isomer of CLA; that the effect of dietary CLA on body fat is greatly enhanced in animals consuming coconut oil versus soy oil; and that cultured mouse 3T3 fibroblasts appear more susceptible to effects of CLA on DNA fragmentation, than are 3T3 adipocytes. We have also measured the change in blood glucose concentration following insulin injection in mice fed CLA as compared to no CLA. The mice fed CLA appear to become insulin resistant.
Impacts
The immediate impact is that our results are interpreted to indicate the use of CLA as a human food supplement should not be endorsed unless the potential effect of CLA on insulin resistance in humans is ruled out. A longer term impact will be an improved understanding of mechanisms determining fat deposition which should ultimately enable applications for manipulation of body fatness.
Publications
- Miner, J.L. 2003. The Adipocyte as an Endocrine Cell. J. Anim. Sci. (in press).
- Hargrave, K.M.,M.J. Azain, S.D. Kachman, and J.L. Miner.2003. Conjugated linoleic acid does not improve insulin tolerance in mice. Obes. Res. 11:1104-15.
- Hargrave, K.M., M.J. Azain, and J.L. Miner. 2003. Aspirin does not alter conjugated linoleic acid-induced body fat loss in mice. Faseb J. 17(4):A700.
- Hargrave, K.M. and J.L. Miner. 2003. Effect of conjugated linoleic acid on DNA fragmentation of preadipocytes in culture. J. Anim. Sci. 81(Suppl. 2): Midwestern Sect., ASAS:30, #121.
- Hargrave, K.M. and J.L. Miner. 2003. Effect of conjugated linoleic acid on DNA fragmentation in cultured adipocytes. J. Anim. Sci.(Suppl. 1):165.
- Miner, J.L. and K.M. Hargrave. 2003. The adipocyte as an endocrine cell. J. Anim. Sci. 81(Suppl. 1):7.
- Miner, J.L. 2003. Recently identified signals for feed intake regulation. J. Anim. Sci. 81(Suppl. 1):123.
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Progress 10/01/01 to 09/30/02
Outputs
Modulation of Adipose Tissue in a Mouse Model by Dietary Conjugated Linoleic Acid Consumption of a linoleic acid isomer (CLA) reduces body fat. The central hypothesis was that CLA is desaturated and elongated to a conjugated isomer of arachidonic acid that competes with arachidonic acid for cyclooxygenase, thereby influencing prostaglandin synthesis and fat cell metabolism. Specific hypotheses were that: 1) dietary linoleic acid deficiency can enhance sensitivity to CLA, and 2) CLA reduces sensitivity to insulin. The first hypothesis was tested using a 2 x 2 factorial arrangement. The factors were basal diet (linoleic acid deficient vs adequate) including CLA vs no CLA. Mice were fed for 8 wk then body fat and programmed cell death in adipose tissue were determined. By itself, linoleic acid deficiency had no effect. However, it did increase sensitivity of mice to CLA. When linoleic acid was adequate, CLA reduced body fatness by 25%. When linoleic acid was deficient,
CLA reduced body fat by 75%. This supports the model's prediction that CLA and linoleic acid metabolites compete for cyclooxygenase. The second hypothesis was tested by comparing blood glucose response to insulin injection in mice fed CLA vs no CLA. A third group of mice with restricted intake controlled for CLA effects on body fat. Mice fed the basal diet, restricted or ad libitum, exhibited a reduction in blood glucose following insulin injection. This reduction was not observed in mice fed CLA. Therefore, CLA can cause resistance to insulin and this is not simply a result of reduced body fat. CLA should not be recommended for treatment of obesity in people that exhibit diabetic tendencies. Second, this work teaches something about the fundamental mechanism by which CLA mediates its potent effect on body fat. Perhaps foods rich in linoleic acid (many plant oils) prevent beneficial effects CLA-containing animal fats (anti-cancer and anti-obesity).
Impacts
This work adds to our understanding of how fat deposition in adipose tissue is regulated, and is thereby expected to facilitate future development of therapies for manipulation of adipose tissue function.
Publications
- Hargrave,K.M., B.J. Meyer, C. Li, M.J. Azain, C.A. Baile, J.L. Miner. 2002. Influence of Conjugated Linoleic Acid and Fat Source on Body Fat and Apoptosis in Mice. Obes. Res. (submitted, ARD No. 13830).
- Hargrave, K.M., M.J. Azain, S.D. Kachman, and J.L. Miner. 2002. Conjugated linoleic acid does not improve insulin sensitivity in mice. Obes. Res. (submitted, ARD No. 13831).
- Hargrave, K.M., C.L. Li, B.J. Meyer, S.D. Kachman, D.L. Hartzell, M.A. Della-Fera, J.L. Miner, and C.A. Baile. 2002. Adipose Depletion and Apoptosis Induced by Trans-10, Cis-12 Conjugated Linoleic Acid in Mice. Obes. Res. (Accepted, in press ARD No. 13667).
- Hargrave, K.M. and J.L. Miner. 2002. Influence of linoleic acid isomer on body fat in mice. J. Anim. Sci. 80(Suppl. 2):50.
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Progress 10/01/00 to 09/30/01
Outputs
Acylation Stimulating Protein. Porcine acylation stimulating protein purified from serum was found to be capable of stimulating esterification of oleate by more than two-fold in monolayer cultures of human fibroblasts. Conjugated Linoleic Acid (CLA). Using a mouse model, we determined that all of the body fat-depleting effect of a commercial CLA mixture could be replicated with a single isomer, trans10, cis9 CLA. Feeding mice 1% CLA mixture (.44% trans10, cis9 CLA) caused body fat depletion within five days and induced apoptosis in adipose tissue. We hypothesized that CLA must be elongated and desaturated in order to cause these effects. Based on this hypothesis, and the additional assumption that linoleic acid could compete for the enzymes that would elongate and desaturate CLA, we determined the interaction of dietary essential fatty acids with dietary CLA. Mice were fed either a base diet containing soy oil or one containing coconut oil (essential fatty acid
deficient). After 6 wk, half of the mice on each of the two base diets, were supplemented with 0.5% CLA mixture for 2 wk. Percent body fat of the mice at termination for Soy oil, Coconut oil, Soy + CLA, and Coconut + CLA was 22, 21, 15, and 6%, respectively. Based on these results, we conclude that CLA induces body fat loss either by inhibiting linoleic acid metabolism, or alternately that linoleic acid prevents CLA-induced body fat depletion by inhibiting the elongation and desaturation of CLA. The second interpretation appears more correct because a base diet of fish oil, which was reported by others to deplete arachidonate stores, does not influence the response of mice to dietary CLA.
Impacts
The observation that conjugated linoleic acid causes programmed cell death of fat cells reveals dramatically a new way in which fat deposition may be regulated.
Publications
- Zhang, H., S.K. Jacobi, C.F. Toombs, G. Sarath, and J.L. Miner. 2001. Purification and characterization of acylation stimulating protein from porcine serum. Protein Expression and Purification (submitted, ARD No. 13412).
- McDaneld, T.G., M.K. Nielsen, and J.L. Miner. 2001. Uncoupling proteins (UCP) and energy expenditure in mice divergently selected for heat loss. J. Anim. Sci. (in press, ARD No. 13218).
- Miner, J.L., J.M. Evans, T.G. McDaneld, C.F. Toombs, and M.K. Nielsen. 2001. Leptin sensitivity of mice bred for high and low energy expenditure. Obes. Res. (submitted, ARD No. 13236)
- Jacobi, S.K., and J.L. Miner. 2001. Human acylation-stimulating protein and lipid biosynthesis in bovine adipose tissue explants. J. Anim. Sci. (in press, ARD No. 13194).
- Miner, J.L., K.J. Hahn, M.E. Spurlock, and N.R. Staten. 2001. Characterization of porcine adipsin. Protein Expression and Purif. 23:14-21. ARD No. 13202.
- Hargrave, K.M. and J.L. Miner. 2002. Influence of linoleic acid isomers on body fat in mice. J. Anim. Sci. 80 (in press).
- Hargrave, K.M., B.J. Meyer, and J.L. Miner. 2002. Dietary fatty acids modify CLA effect on body fat. FASEB J. (submitted for April meeting).
- Meyer, B.J., K.M. Hargrave, and J.L. Miner. 2002. Fish oil, conjugated linoleic acid, and body fat deposition. J. Anim. Sci. 80 (in press).
- McDaneld, T.G., M.K. Nielsen, and J.L. Miner. 2001. Effect of uncoupling protein 1 knockout in mice divergently selected for heat loss. J. Anim. Sci. 79 (Suppl. 2):47.
- Nollette, K.R., and J.L. Miner. 2001. Conjugated linoleic acid and body fat reduction in mice. J. Anim. Sci. 79(Suppl. 2):109.
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Progress 10/01/99 to 09/30/00
Outputs
Acylation Stimulating Protein (ASP). ASP may be an important adipose signalling molecule. We previously found that human ASP could enhance incorporation of either fatty acid or acetate into triacylglycerol in bovine adipose explants. Human ASP also stimulated esterification of fatty acid in porcine adipose explants. Now, we have obtained rabbit antisera to an ASP peptide and used this to develop a method for purification of ASP. We have obtained ASP from porcine serum. Porcine ASP is currently being evaluated for activity in porcine adipocytes. Apoptosis in Adipocytes. We found that feeding conjugated linoleic acid (CLA) isomers to mice causes loss of body fat and apoptosis of adipocytes within 5 d. Cis10,trans12/trans10,cis12 CLA causes a similar depletion of body fat as the mixture whereas 9,12 isomers do not.
Impacts
The purification of ASP from porcine serum: 1) verifies its presence in a farm species, and 2) provides a reagent to allow determination of its activity in swine. The observation that CLA can induce apoptosis in adipocytes is novel. It may represent a novel mechanism of CLA effects on body composition, and another means of regulating adiposity.
Publications
- Conjugated Linoleic Acid (CLA), Body Fat, and Apoptosis. 2000. Miner, J.L., C.A. Cederberg, M.K. Nielsen, X. Chen, and C.A. Baile. Obesity Res. (In press).
- Jones, J.G. and J.L. Miner. 2000. Behavior of mice selected for high and low heat loss during light and dark photoperiods. J. Anim. Sci. 78(Suppl. 2):50.
- McDaneld, T.G. and J.L. Miner. 2000. Increased uncoupling protein-1 (UCP-1) mRNA in mice selected for low heat loss versus high heat loss. J. Anim. Sci. 78(Suppl. 2):45.
- Miner, J.L., C.A. Cederberg, M.K. Nielsen, X. Chen, and C.A. Baile. 2000. Conjugated linoleic acid (CLA), body fat, and apoptosis in mice. Faseb J. 14(4):A479.
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Progress 10/01/98 to 09/30/99
Outputs
One emphasis of this project is to determine if acylation stimulating protein (ASP) regulates triglyceride storage in adipocytes. We have found that human ASP can enhance esterification of fatty acids in both bovine and porcine adipose tissue in culture. We also have cloned portions of the complement C3 cDNA which include the regions coding for ASP. The porcine sequence was used to direct synthesis of a peptide. We have generated antisera to this peptide with the hope that we can use the antisera to assay ASP in pig serum and during purification of ASP from pig serum. Our initial bleed was used in Western blotting experiments and recognizes human ASP, and two bands in porcine serum but nothing in bovine serum. Concerning our work with the mouse model of maintenance energy, we have found that a line selected for high heat production (MH) does not have elevated uncoupling protein one mRNA in brown adipose tissue compared to a line selected for low heat production (ML).
We are currently developing ribonuclease protection assays for uncoupling proteins 2 and 3. We have backcrossed an uncoupling protein 1 knockout allele into the MH and ML lines and should have uncoupling protein one homozygous congenic lines by March.
Impacts
The impact of our results thus far is support of a hypothesis that ASP is important for regulation of fat storage and development of reagents for study of this molecule in livestock species is warranted.
Publications
- Jacobi, S. K., K. Cianflone, and J.L. Miner. 1999. Feeding status and the responsiveness of bovine adipose tissue to human acylation stimulating protein (hASP). J. Anim. Sci. (Abstr.).
- Knobbe, M.G., M. K. Nielsen, and J.L. Miner. 1999. Effect of triiodothyronine (T3) on heat production of MH and ML mice. J. Anim. Sci. (Abstr.).
- Ramsel, C.L., S.K. Jacobi, K. Cianflone, and J.L. Miner. 1999. Acylation stimulating protein and fatty acid esterification in the pig. J. Anim. Sci. (Abstr.).
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Progress 10/01/97 to 09/30/98
Outputs
Acylation Stimulating Protein (ASP) and Lipid Synthesis. Subcutaneous adipose tissue from cattle and from pigs (inner layer) was cultured as 10- to 20-mg explants for 1 to 6 h. Addition of human ASP increased the quantity of extractable lipid synthesized from 3H-oleate (porcine and bovine) and from 14C-acetate (bovine). A region of bovine and porcine complement C3 mRNA which codes for ASP was cloned by reverse transcriptase-polymerase chain reaction and sequenced. Angiotensin II and Lipid Synthesis. Addition of angiotensin II to bovine adipose explant cultures did not influence synthesis of lipid from 14C-acetate though it did increase lipid synthesis in mouse explant cultures. Energy Expenditure. Mice selected divergently for heat loss into two lines which differ by about 45% were found to have a similar heat production response to stimulation with the beta-3 adrenergic agonist, CL316,243. These two lines also have similar serum concentration of triiodothyronine,
thyroxine, and free triiodothyronine, and increase heat production similarly in response to injection of triiodothyronine. To test the hypothesis that the difference in thermogenesis between the two mouse lines is dependent upon uncoupling protein-1, two congenic lines are being produced. These congenic lines will have the uncoupling protein-1 gene disrupted by homologous recombination.
Impacts
(N/A)
Publications
- Jacobi, S.K. and J.L. Miner. 1998. Effect of human acylation stimulating protein (ASP) and adenosine in bovine adipose explants. J. Anim. Sci. 76 (Abstr., Suppl.2):44.
- Strickland, J.L., J.L. Miner, and M.K. Nielsen. 1998. Feed intake, thyroid hormones, and adrenergic stimulation in high and low heat loss mice. J. Anim. Sci. 76 (Abstr., Suppl. 2):45.
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