Older American Women May Especially Benefit from Distributing and Consuming Protein for Decreasing Odds of Functional Limitations.

OBJECTIVES: Protein is a key macronutrient for preserving physical function, but the role of protein intake on functional status may differ in men and women. We sought to examine the associations of daily protein intake and distribution on functional limitations in older American men and women. DESIGN: Cross-sectional. SETTING: Population-based survey. PARTICIPANTS: The analytic sample included 3,976 men and 4,081 women aged ≥60-years from the 2007-2016 waves of the National Health and Nutrition Examination Survey. MEASUREMENTS: Participants reported their ability to perform basic activities of daily living, instrumental activities of daily living, leisure and social activities, lower extremity mobility activities, and general physical tasks. Those reporting difficulty or an inability in completing such functional tasks were considered as having a functional limitation. Protein intake was determined with dietary recalls and participants revealed functional limitations. Protein recommendations of ≥0.80, ≥1.00, and ≥1.50 g/kg/day were used. Based on these cut-points, we also investigated distribution of protein across 4 eating occasions at ≥0.20, ≥0.25, and ≥0.38 g/kg/meal, respectively. RESULTS: Older women meeting each recommendation had decreased odds for functional limitations: 0.55 (95% confidence interval (CI): 0.40-0.75) for ≥0.80 g/kg/day, 0.75 (CI: 0.58-0.97) for ≥1.00 g/kg/day, and 0.72 (CI: 0.55-0.94) for ≥1.5 g/kg/day. No significant associations were observed in older men. Further, older women with protein consumption ≥0.20 g/kg/meal had decreased odds for functional limitations: 0.24 (CI: 0.10-0.61) for 1 occasion, 0.20 (CI: 0.08-0.49) for 2 occasions, 0.16 (CI: 0.07-0.40) for 3 occasions, and 0.12 (CI: 0.04-0.32) for 4 occasions. A similar trend was observed for intake ≥0.25 g/kg/meal: 0.31 (CI: 0.16-0.62) for 2 occasions, 0.30 (CI: 0.14-0.61) for 3 occasions, and 0.31 (CI: 0.12-0.78) for 4 occasions. Women with 1 and 2 eating occasions at ≥0.38 g/kg/meal of protein had 0.66 (CI: 0.48-0.91) and 0.54 (CI: 0.37-0.79) decreased odds for functional limitations, respectively. CONCLUSION: Trials that are powered to detect the effects of protein on functional status in women will help to establish causality.

Depression, daily stressors and inflammatory responses to high-fat meals: when stress overrides healthier food choices.

Depression, stress and diet can all alter inflammation. This double-blind, randomized crossover study addressed the impact of daily stressors and a history of major depressive disorder (MDD) on inflammatory responses to high-fat meals. During two separate 9.5 h admissions, 58 healthy women (38 breast cancer survivors and 20 demographically similar controls), mean age 53.1 years, received either a high saturated fat meal or a high oleic sunflower oil meal. The Daily Inventory of Stressful Events assessed prior day stressors and the Structured Clinical Interview for DSM-IV evaluated MDD. As expected, for a woman with no prior day stressors, C-reactive protein (CRP), serum amyloid A (SAA), intercellular adhesion molecule-1 (sICAM-1) and vascular cell adhesion molecule-1 (sVCAM-1) were higher following the saturated fat meal than the high oleic sunflower oil meal after controlling for pre-meal measures, age, trunk fat and physical activity. But if a woman had prior day stressors, these meal-related differences disappeared-because the stressors heightened CRP, SAA, sICAM-1 and sVCAM-1 responses to the sunflower oil meal, making it look more like the responses to the saturated fat meal. In addition, women with an MDD history had higher post-meal blood pressure responses than those without a similar history. These data show how recent stressors and an MDD history can reverberate through metabolic alterations, promoting inflammatory and atherogenic responses.

Dietary protein and beef consumption predict for markers of muscle mass and nutrition status in older adults.

OBJECTIVE: To determine the relationship of beef and protein intake to nutrition status, body composition, strength, and biochemical measures of vitamin and mineral status, inflammation and blood lipids in older adults. DESIGN: Cross-sectional observational study. SETTING: State of Ohio, U.S.A. PARTICIPANTS: 142 adults ages 60-88. MEASUREMENTS: Subjects completed a Diet History Questionnaire, and questionnaires related to nutrition status and activity. Subjects also underwent measurements of body composition and strength, and a subset took part in a blood draw for biochemical measurements. RESULTS: Beef intake (g/d) was positively correlated to muscle mass measured by mid-arm muscle area (R=0.128, p=0.030). From multiple linear regression analysis, a 1oz/d (~28g/d) increase in beef consumption predicts for a 2.3cm(2) increase in mid-arm muscle area. Beef intake was negatively correlated to total (R=-0.179, p=0.035) and HDL (R=-0.247, p=0.004) cholesterol, and there was no association between beef and LDL-cholesterol, triglycerides, liver enzymes, or inflammatory markers. Protein intake (% of total energy) was positively correlated to nutrition status measured by the Mini Nutrition Assessment (R=0.196, p=0.020), and calf circumference (R=0.190, p=0.024), and these correlations remained when potential confounders were accounted for in multiple linear regression models. Protein intake was also positively correlated with BMI when analyzed with multiple linear regression. CONCLUSIONS: Beef intake was positively associated with mid-arm muscle area, and protein intake was positively associated with nutrition status, calf circumference, and BMI in older adults. Consuming lean cuts of beef in moderation may be a healthy way in which older adults can increase protein intake, preserve muscle mass and improve nutrition status.

Regulation of adipose triglyceride lipase by rosiglitazone.

AIM: To elucidate the mechanism by which rosiglitazone regulates adipose triglyceride lipase (ATGL). METHODS: Male C57Bl/6 mice were treated with rosiglitazone daily (10 mg/kg body weight), and adipose tissues were weighed and preserved for mRNA and protein analysis of ATGL. In parallel, preadipocyte (3T3-L1) cells were differentiated with insulin/dexamethasone/3-isobutyl-1-methlxanthine cocktail or rosiglitazone, and ATGL levels were measured with real-time PCR, western blotting and immunohistochemistry. RESULTS: Rosiglitazone concomitantly promoted differentiation of pre-adipocytes to functional adipocytes and induced mRNA levels of ATGL. The peroxisome proliferator-activated receptor-gamma (PPARgamma) antagonist bisphenol A diglycidyl ether significantly abrogated the induction of mRNA, but not protein levels of ATGL by rosiglitazone in differentiated 3T3-L1 adipocytes. In the presence of epinephrine rosiglitazone stimulated free fatty acid release and increased diacylglycerol acyltransferase-1 (DGAT-1) mRNA suggest that ATGL and DGAT-1 may be cooperatively involved in rosiglitazone-stimulated triglyceride hydrolysis and fatty acid re-esterification in 3T3-L1 adipocytes. Treatment of 3T3-L1 adipocytes with rosiglitazone or insulin did not appear to alter localization of ATGL staining surrounding lipid droplets. Finally, we found that rosiglitazone increased ATGL mRNA levels in 3T3-L1 adipocytes in the presence of cycloheximide, an inhibitor of protein synthesis, suggesting that rosiglitazone regulation of ATGL occurs at the transcriptional level. CONCLUSIONS: Rosiglitazone directly regulates transcription of ATGL, likely through a PPARgamma-mediated mechanism.

Evaluation of the effects of dietary fat, conjugated linoleic acid, and ractopamine on growth performance, pork quality, and fatty acid profiles in genetically lean gilts.

An 8-wk study of the effects of CLA, rendered animal fats, and ractopamine, and their interactive effects on growth, fatty acid composition, and carcass quality of genetically lean pigs was conducted. Gilts (n = 228; initial BW of 59.1 kg) were assigned to a 2 x 2 x 3 factorial arrangement consisting of CLA, ractopamine, and fat treatments. The CLA treatment consisted of 1% CLA oil (CLA-60) or 1% soybean oil. Ractopamine levels were either 0 or 10 ppm. Fat treatments consisted of 0% added fat, 5% choice white grease (CWG), or 5% beef tallow (BT). The CLA and fat treatments were initiated at 59.1 kg of BW, 4 wk before the ractopamine treatments. The ractopamine treatments were imposed when the gilts reached a BW of 85.7 kg and lasted for the duration of the final 4 wk until carcass data were collected. Lipids from the belly, outer and inner layers of backfat, and LM were extracted and analyzed for fatty acid composition from 6 pigs per treatment at wk 4 and 8. Feeding CLA increased (P < 0.02) G:F during the final 4 wk. Pigs fed added fat as either CWG or BT exhibited decreased (P < 0.05) ADFI and increased (P < 0.01) G:F. Adding ractopamine to the diet increased (P < 0.01) ADG, G:F, and final BW. The predicted carcass lean percentage was increased (P < 0.05) in pigs fed CLA or ractopamine. Feeding either 5% fat or ractopamine increased (P < 0.05) carcass weight. Adding fat to the diets increased (P < 0.05) the 10th rib backfat depth but did not affect predicted percent lean. Bellies of gilts fed CLA were subjectively and objectively firmer (P < 0.01). Dietary CLA increased (P < 0.01) the concentration of saturated fatty acids and decreased (P < 0.01) the concentration of unsaturated fatty acids of the belly fat, both layers of backfat, and LM. Ractopamine decreased (P < 0.01) the i.m. fat content of the LM but had relatively little effect on the fatty acid profiles of the tissues compared with CLA. These results indicate that CLA, added fat, and ractopamine work mainly in an additive fashion to enhance pig growth and carcass quality. Furthermore, these results indicate that CLA results in more saturated fat throughout the carcass.

Differentiation-dependent regulation of the cyclooxygenase cascade during adipogenesis suggests a complex role for prostaglandins.

AIM: A thorough understanding of the mechanisms of adipocyte differentiation and metabolism is important for the prevention and/or treatment of obesity and its complications, including type 2 diabetes mellitus. A complex role for prostaglandins (PGs) in adipogenesis is suggested. We examined the expression and cellular localization of enzymes in the cyclooxygenase (COX) cascade that synthesize PGs as well as the PG profile as a function of differentiation status in 3T3-L1 cells. METHODS: Murine 3T3-L1 preadipocytes were used as a model for studies of adipocyte differentiation induced by a hormone cocktail and compared with the parental fibroblastic line NIH 3T3. Both cell lines were incubated in maintenance medium or differentiation medium. Nine days after differentiation, the expression of enzymes in the COX cascade was evaluated by immunoblot analysis, reverse transcriptase-polymerase chain reaction (RT-PCR) and immunocytochemistry, and PG formation was examined using enzyme immunoassay. RESULTS: A differentiation-dependent diminution of COX-1 and COX-2 mRNA and cognate proteins in 3T3-L1 cells was observed. PG release, including PGE(2), 6-keto PGF(1alpha), PGD(2) and 15d-PGJ(2), significantly decreased following differentiation in 3T3-L1 cells (anova/Tukey, p < 0.05). However, microsomal PGE synthase (mPGES) and lipocalin-type PGD synthase (L-PGDS) were selectively upregulated. Immunocytochemistry revealed that COX-1 and COX-2 became intracellularly more diffuse upon differentiation, whereas mPGES was redistributed to the nuclear compartment. CONCLUSIONS: Regulation of PG formation and COX-2 expression in 3T3-L1 cells is differentiation-dependent and involves changes in the levels of gene expression of the individual isoforms as well as redistribution of the enzymes within cellular compartments.

Effects of conjugated linoleic acid on the belly firmness and fatty acid composition of genetically lean pigs.

A study of the effects of conjugated linoleic acid (CLA) on the belly firmness and fatty acid composition of genetically lean pigs was conducted. From 75 to 120 kg live weight, 30 gilts were allowed ad libitum access to a corn-soybean meal diet supplemented with either 1% CLA oil (CLA-60) or 1% sunflower oil (SFO) or were fed the sunflower oil-supplemented diet restricted to the amount consumed by pigs fed the CLA-60 diet (RSFO). Conjugated linoleic acid oil consists of 60% positional and geometric isomers of CLA. Pigs fed SFO exhibited higher average daily gains (0.98 vs 0.80 kg/d, P < 0.01) than RSFO-fed pigs, but there were no effects of dietary treatment on feed intake or feed efficiency. Dietary treatment did not affect (P > 0.05) backfat thickness or longissimus muscle area. Bellies of gilts fed CLA-60 were subjectively evaluated to be firmer (2.91 vs 2.43 or 2.07 +/- 0.13, P < 0.01) than those of SFO- or RSFO-fed gilts, respectively. The longissimus muscle of gilts fed CLA-60 contained more saturated fatty acids (39.77 vs. 36.04 or 36.73 +/- 0.74%, P < 0.001) and less unsaturated fatty acids (60.23 vs 63.96 or 63.27 +/- 0.74%, P < 0.001) than that of gilts fed SFO or RSFO, respectively. The belly fat of gilts fed CLA-60 contained more saturated fatty acids (44.45 vs. 37.50 or 36.60 +/- 0.46%, P < 0.001) and less unsaturated fatty acids (54.78 vs. 61.75 or 62.47 +/- 0.46%, P < 0.001), resulting in lower iodine values (57.69 vs 66.37 or 65.62 +/- 0.91, P < 0.001) than that of gilts fed SFO or RSFO, respectively. Gilts fed CLA-60 accumulated more CLA in the longissimus muscle (0.55 vs 0.09 or 0.09 +/- 0.03%, P < 0.01) and belly fat (1.56 vs. 0.13 or 0.13 +/- 0.15%, P < 0.001) than did gilts fed SFO or RSFO, respectively. Dietary treatment did not affect (P > 0.05) 24-h pH, drip loss or subjective quality evaluations of the longissimus muscle. The effect of supplemental CLA to improve belly firmness is of practical significance and may provide a nutritional solution to carcass fat and belly firmness problems, thereby enhancing the overall value of extremely lean carcasses.

Activators of peroxisome proliferator-activated receptor-alpha partially inhibit mouse skin tumor promotion.

Several recent reports have suggested that peroxisome proliferator-activated receptors (PPARs) may be involved in the development of neoplasias in different tissue types. The present study was undertaken to determine whether PPARs play a role in skin physiology and tumorigenesis. In an initiation-promotion study, SENCAR mice treated topically with the PPARalpha ligands conjugated linoleic acid and 4-chloro-6-(2,3-xylidino)-2-pyrimidinylthioacetic acid (Wy-14643) exhibited an approximately 30% lower skin tumor yield compared with mice treated with vehicle. The PPARgamma and PPARdelta activators troglitazone and bezafibrate, respectively, exerted little, if any, inhibitory activity. PPARalpha was detected in normal and hyperplastic skin and in papillomas and carcinomas by immunohistochemistry. In addition, PPARalpha, PPARdelta/PPARbeta, and PPARgamma protein levels were analyzed by immunoblotting in normal epidermis and papillomas. Surprisingly, the levels of all three isoforms were increased significantly in tumors as opposed to normal epidermis. In primary keratinocyte cultures, protein levels of PPARalpha and, to a lesser extent, PPARgamma were markedly increased when the cells were induced to differentiate with high-calcium (0.12 mM) conditions. In addition, we observed that Wy-14643 enhanced transcriptional activity of a peroxisome proliferator-response element-driven promoter in a mouse keratinocyte cell line. These results demonstrate that keratinocytes express functional PPARalpha, that PPARalpha may play a role in differentiation, and that ligands for PPARalpha are moderately protective against skin tumor promotion. We conclude that selective PPARalpha ligands may exert their protective role against skin tumor promotion by ligand activation of PPARalpha.

Conjugated linoleic acid is a potent naturally occurring ligand and activator of PPARalpha.

We have previously shown that a mixture of dietary conjugated derivatives of linoleic acid (conjugated linoleic acid, CLA) induces peroxisome proliferator-responsive enzymes and modulates hepatic lipid metabolism in vivo. The present studies demonstrate that CLA is a high affinity ligand and activator of peroxisome proliferator-activated receptor alpha (PPARalpha) and induces accumulation of PPAR-responsive mRNAs in a rat hepatoma cell line. Using a scintillation proximity assay (SPA), CLA isomers were shown to be ligands for human PPARalpha with a rank order of potency of (9Z,11E)>(10E,12Z)>(9E,11E)> furan-CLA (IC(50) values from 140 nm to 400 nm). Levels of acyl-CoA oxidase (ACO), liver fatty acid-binding protein (L-FABP), and cytochrome P450IVA1 (CYP4A1) mRNA were induced by CLA in FaO hepatoma cells. Even though linoleate and CLA were incorporated into lipids of hepatoma cells to the same extent, linoleate had little or no effect on ACO, CYP4A1, or L-FABP mRNA. In agreement with its binding potency, (9Z,11E)-CLA was the most efficacious PPARalpha activator in the mouse PPARalpha-GAL4(UAS)(5)-CAT reporter system. These data indicate that CLA is a ligand and activator of PPARalpha and its effects on lipid metabolism may be attributed to transcriptional events associated with this nuclear receptor. Also, (9Z,11E)-CLA is one of the most avid fatty acids yet described as a PPARalpha ligand.

Inositol hexaphosphate reduces 12-O-tetradecanoylphorbol-13-acetate-induced ornithine decarboxylase independent of protein kinase C isoform expression in keratinocytes.

High-fiber diets have been shown to have beneficial effects on preventing tumorigenesis. Inositol hexaphosphate (InsP6 or phytic acid) which is a fiber-associated component of cereals and legumes has been demonstrated to inhibit cell proliferation and enhance cell differentiation, indicating its potential for chemopreventive roles. In this study, we investigated the effect of InsP6 on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced ornithine decarboxylase (ODC) activity, an essential event in tumor promotion in HEL-30 cells, a murine keratinocyte cell line and SENCAR mouse skin. ODC activity was significantly reduced by 0.5 mM InsP6 in keratinocytes (P < 0.01). Furthermore, when mouse skin was treated with 10 mM InsP6, ODC induction was significantly inhibited (P < 0.05). In addition, the expression of TPA-induced c-myc mRNA was significantly inhibited by the same InsP6 treatments in HEL-30 cells and CD-1 mouse skin (P < 0.01). No changes in protein kinase C (PKC) isoform expression and phorbol dibutyrate binding due to InsP6 treatment were found in HEL-30 cells. These results indicate that InsP6 reduces TPA-induced ODC activity independent of PKC isoform expression.

Effect of dietary conjugated linoleic acid on phorbol ester-induced PGE2 production and hyperplasia in mouse epidermis.

Conjugated linoleic acid (CLA) is a chemoprotective fatty acid that inhibits phorbol ester-induced skin tumor promotion in mice. The goal of the present study was to determine potential chemoprotective mechanisms through which CLA may be acting. Mice were fed diets containing 0.0%, 0.5%, 1.0%, or 1.5% CLA (by wt) for six weeks. The epidermis was evaluated for fatty acid composition, vascular permeability, prostaglandin E2 (PGE2) production, hyperplasia, ornithine decarboxylase activity, and c-myc mRNA accumulation. Fatty acid analysis of mouse epidermis demonstrated a dose-dependent increase of CLA incorporation into phospholipids and neutral lipids. In mice topically treated with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA), dietary CLA (1.5%) significantly (p < 0.05) reduced PGE2 synthesis (2-fold). Additionally, CLA lowered accumulation of c-myc mRNA, a gene commonly associated with regulating cell cycle components involved in cellular proliferation, although this trend was not significant. Vascular permeability was unaffected by dietary CLA. These data suggest that dietary CLA modulates TPA-induced tumor promotion through a mechanism involving PGE2 production; however, dietary CLA had a moderate effect on c-myc mRNA levels and little effect on TPA-induced hyperplasia and ornithine decarboxylase activity.

Conjugated linoleic acid activates peroxisome proliferator-activated receptor alpha and beta subtypes but does not induce hepatic peroxisome proliferation in Sprague-Dawley rats.

Since conjugated linoleic acid (CLA) has structural and physiological characteristics similar to peroxisome proliferators, we hypothesized that CLA would activate peroxisome proliferator-activated receptor (PPAR). We compared the effects of dietary CLA (0.0, 0.5, 1.0 and 1.5% by weight) with a peroxisome proliferator (0.01% Wy-14,643) in female and male Sprague-Dawley (SD) rats. Dietary CLA had little effect on body weight, liver weight, and hepatic peroxisome proliferation, compared to male rats fed Wy-14,643 diet. Lipid content in livers from rats fed 1.5% CLA and Wy-14,643 diets was increased (P < 0.01) when compared to rats fed control diets regardless of gender. Hepatic acyl-CoA oxidase (ACO) mRNA levels were increased 3-fold in male rats fed 1.5% CLA diet compared to rats fed control diets while Wy-14,643 supported approximately 30-fold ACO mRNA accumulation. A similar response was observed for liver fatty acid-binding protein (L-FABP) mRNA. The effect of dietary treatments on hepatic PPAR-responsive genes in female rats was weaker than in male rats. The (9Z,11E)-CLA isomer activated PPAR alpha in transfected cells to a similar extent as Wy-14,643, whereas the furan-CLA metabolite was comparable to bezafibrate on activating PPAR beta. These data suggest that while CLA was able to activate PPARs it is not a peroxisome proliferator in SD rats.

Species differences in the metabolism and regulation of gene expression by conjugated linoleic acid.

Conjugated linoleic acid (CLA) inhibits carcinogenesis and atherosclerotic plaque formation and delays the onset of diabetes in experimental animals. Whereas a plethora of data has demonstrated beneficial effects in rodent models, little work has been done to determine the role of dietary CLA in human health. The ability of CLA to modulate lipid metabolism appears to be a pivotal mechanism of CLA's beneficial effects in mice and rats. In particular, dietary CLA induces the expression of genes dependent in part on the transcription factor, peroxisome proliferator-activated receptor (PPAR). Furthermore, several CLA isomers are high-affinity ligands and activators for PPAR alpha. Within various rodent species and strains, dietary CLA exerts varying potencies; therefore, the differences in species' sensitivities are of great importance when trying to extrapolate the rodent data to be relevant in humans. This review presents the latest findings of the ability of CLA to alter lipid metabolism and gene expression in several different strains of mice and rats and speculates on the implications of these findings for human health.

Comparison of dose-response relationships for induction of lipid metabolizing and growth regulatory genes by peroxisome proliferators in rat liver.

The regulation of gene expression via the peroxisome proliferator-activated receptor (PPAR) is believed to be critical in the effects of peroxisome proliferators on lipid metabolism and possibly in hepatocarcinogenesis. The involvement of PPAR in the peroxisome proliferator-mediated induction of fatty acid metabolizing genes such as acyl-CoA oxidase (ACO), fatty acid-binding protein (FABP), and cytochrome P450IVA1 (CYP4A1) has been clearly demonstrated. However, the induction by peroxisome proliferators of important growth regulatory genes such as c-myc has not been investigated extensively. In these studies we examined the dose-response relationships for the induction of mRNA for the PPAR-regulated and lipid metabolizing genes ACO, FABP, and CYP4A1 and compared them to the immediate early gene c-myc. Liver mRNA from rats fed various amounts of the peroxisome proliferator Wy14,643 for 13 weeks was utilized. The lipid metabolism and growth regulatory genes were induced by subchronic administration of Wy14,643 but to varying degrees and with different sensitivities. The lowest dose that resulted in a significant change in ACO and FABP expression was 10 ppm. The mRNA for CYP4A1 and c-myc was significantly affected at the lowest dose examined (5 ppm). Also, the maximal induction ranged from 10(5)-fold (CYP4A1) to less than 10-fold (FABP) relative to vehicle-treated animals. The accumulation of mRNA for ACO, FABP, and CYP4A1, but not c-myc, showed typical receptor-mediated dose-response relationships. The effects on gene expression were compared to rates of hepatic cell proliferation, a pertinent marker of tumor promotion and hepatocarcinogenesis. Surprisingly, ACO mRNA showed an excellent correlation (r2 = 0.9) while c-myc mRNA exhibited a poor correlation (r2 = 0.3) with cell proliferation in rat liver. Although the differences between the dose-response relationships of ACO and c-myc mRNA accumulation may suggest immediate early genes are not controlled by PPAR, evidence from PPARalpha null mice support this receptor in both lipid metabolism and growth regulatory genes. This study shows the complexity of responses mediated by peroxisome proliferators, with ACO being a good marker of PPAR-mediated events as well as cell proliferation, while c-myc, a known growth regulatory gene, was induced by Wy14,643 partially via PPAR but did not correlate well with cell proliferation.

Conjugated linoleic acid reduces arachidonic acid content and PGE2 synthesis in murine keratinocytes.

Dietary conjugated linoleic acid (CLA) is associated with decreased 12-O-tetradecanoyl-phorbol-13-acetate (TPA)-induced tumor promotion in mouse skin. In addition, CLA decreases TPA-induced prostaglandin E synthesis and ornithine decarboxylase activity in cultured keratinocytes compared with linoleic acid (LA) and arachidonic acid (AA). When LA or CLA was added to keratinocyte cell cultures, the amounts of each of these cellular fatty acids increased significantly in a dose-dependent manner. Furthermore, LA treatment was associated with increased cellular AA while the AA content of keratinocytes was reduced when cultures were treated with CLA. Moreover, CLA (16 microg/ml) was more potent than LA at decreasing the level of 14C-AA incorporated into cellular phosphatidylcholine. In order to determine the effect of CLA on arachidonate-derived PGE2, the release of 14C-AA and 14C-PGE2 synthesis was measured in cultures pre-treated with LA/14C-AA or CLA/14C-AA for 12 h. The amount of 14C-AA release induced by TPA in CLA/14C-AA pre-treated cultures was significantly lower than cultures pre-treated with LA/14C-AA. Furthermore, TPA-induced 14C-PGE2 was significantly lower in cultures pre-treated with CLA/14C-AA compared with cultures pre-treated with LA/14C-AA. The effects of LA and CLA on AA composition of phospholipids and subsequent arachidonate-derived PGE2 synthesis will provide insight into the anti-promoter mechanisms of CLA.

Dietary conjugated linoleic acid normalizes impaired glucose tolerance in the Zucker diabetic fatty fa/fa rat.

Conjugated linoleic acid (CLA) is a naturally occurring fatty acid which has anti-carcinogenic and anti-atherogenic properties. CLA activates PPAR alpha in liver, and shares functional similarities to ligands of PPAR gamma, the thiazolidinediones, which are potent insulin sensitizers. We provide the first evidence that CLA is able to normalize impaired glucose tolerance and improve hyperinsulinemia in the pre-diabetic ZDF rat. Additionally, dietary CLA increased steady state levels of aP2 mRNA in adipose tissue of fatty ZDF rats compared to controls, consistent with activation of PPAR gamma. The insulin sensitizing effects of CLA are due, at least in part, to activation of PPAR gamma since increasing levels of CLA induced a dose-dependent transactivation of PPAR gamma in CV-1 cells cotransfected with PPAR gamma and PPRE X 3-luciferase reporter construct. CLA effects on glucose tolerance and glucose homeostasis indicate that dietary CLA may prove to be an important therapy for the prevention and treatment of NIDDM.

Conjugated linoleic acid modulation of phorbol ester-induced events in murine keratinocytes.

Recent work in our lab has shown that the chemoprotective fatty acid, conjugated linoleic acid (CLA), inhibits phorbol ester skin tumor promotion in mice. Because little is known about the deposition of CLA into tissues as well as its biological activity, this study compared the incorporation and biological activity of CLA to linoleic acid (LA; 18:2, c9,c12) and arachidonic acid (AA; 20:4 c5,c8,c11,c14) in cultured keratinocytes. When keratinocytes (HEL-30) were grown in media containing 14C-CLA for various periods, more than 50% of the 14C-CLA was incorporated into cellular lipids by 9 h. The distribution of CLA in phospholipid classes was similar to LA, Approximately 50% of 14C-LA and 14C-CLA were incorporated into phosphatidylcholine (PC), while the remainder was taken up by phosphatidylethanolamine (PE) and phosphatidylserine/phosphatidylinositol (PS/PI). In contrast, 14C-AA was more equitably distributed into PC, PE, or PS/PI (27, 30, or 38%, respectively). When keratinocytes were prelabeled with radiolabeled fatty acids, phorbol ester-induced release of 14C-CLA was 1.5 times higher than 14C-LA and 14C-AA. However, 14C-prostaglandin E (PGE) release in 14C-CLA prelabeled cultures was 6 and 13 times lower than cultures treated with 14C-LA and 14C-AA, respectively. Moreover, the ability of non-radiolabeled CLA to support ornithine decarboxylase activity, a hallmark event of tumor promotion, was significantly lower than in LA- and AA-treated cultures. These studies suggest that CLA inhibits skin tumor promotion, in part, through a PGE-dependent mechanism.

Calcium bioavailability of vegetarian diets in rats: potential application in a bioregenerative life-support system.

Calcium bioavailability of vegetarian diets containing various proportions of candidate crops for a controlled ecological life-support system (CELSS) was determined by femur 45Ca uptake. Three vegetarian diets and a control diet were labeled extrinsically with 45Ca and fed to 5-wk old male rats. A fifth group of rats fed an unlabeled control diet received an intraperitoneal (IP) injection of 45Ca. There was no significant difference in mean calcium absorption of vegetarian diets (90.80 +/- 5.23%) and control diet (87.85 +/- 5.25%) when calculated as the percent of an IP dose. The amounts of phytate, oxalate, and dietary fiber in the diets did not affect calcium absorption.