Conjugated linoleic acids, atherosclerosis, and hepatic very-low-density lipoprotein metabolism

¹ From the Department of Biochemistry and Molecular Biology, Dalhousie University, Halifax, Canada

² Presented at the workshop "The Role of Conjugated Linoleic Acid in Human Health," held in Winnipeg, Canada, March 13–15, 2003.

³ Supported by grants (to RSM) from the Canadian Institutes of Health Research (MOP-42492) and from the Dairy Farmers of Canada.

⁴ Address reprint requests to RS McLeod, Department of Biochemistry and Molecular Biology, Dalhousie University, Halifax, Nova Scotia, Canada B3H 1X5. E-mail: rmcleod2{at}dal.ca.

ABSTRACT

Conjugated linoleic acids (CLAs) are isomeric forms of the 18:2 fatty acid that contain conjugated sites of unsaturation. Although CLAs are minor components of the diet, they have many reported biological activities. For nearly a decade, the potential for CLA to modify the atherosclerotic process has been examined in animal models, and studies of supplementation of the human diet with CLA were started with the anticipation that such an intervention could also reduce the risk of cardiovascular disease. Central to the hypothesis is the expectation that dietary modification could alter plasma lipid and lipoprotein metabolism toward a more cardioprotective profile. This review examines the evidence in support of the hypothesis and the mechanistic studies that lend support for a role of CLA in hepatic lipid and lipoprotein metabolism. Although there are still limited studies in strong support of a role for CLA in the reduction of early atherosclerotic lesions, there has been considerable progress in defining the mechanisms of CLA action. CLA could primarily modulate the metabolism of fatty acids in the liver. The tools are now available to examine isomer-specific effects of CLA on hepatic lipid and lipoprotein metabolism and the potential of CLA to modify hepatic gene expression patterns. Additional animal and cell culture studies will increase our understanding of these unusual fatty acids and their potential for health benefits in humans.

Key Words: Conjugated linoleic acid • CLA • atherosclerosis • very-low-density lipoprotein • LDL • VLDL • fatty acid • triacylglycerols • liver

INTRODUCTION

Conjugated linoleic acid (CLA) is the term used to describe a group of positional and geometric isomers of linoleic acid (LA; 18:2 c9,c12). CLAs are produced naturally by bacterial hydrogenation and isomerization in the gut of ruminant animals, or they can be generated chemically by alkali isomerization of LA. Although there are as many as 28 possible isomeric forms of CLA (1), by far the most abundant isomer in nature is the c9,t11-CLA. In the human diet, CLAs are consumed in milk fat and in meats derived from ruminant animals whereby they represent 0.5–2% of fatty acids. More than 70% of the CLA in these foods is the c9,t11-isomer. By contrast, CLA, which is produced chemically, is available commercially, and is the most common form used for dietary supplementation, is a "mixed isomer" preparation, usually containing 40% c9,t11-CLA, 40% t10,c12-CLA, and 20% other isomers. In 1979, Pariza et al (2) described an extract of cooked ground beef that inhibited mutagenesis in mice, and they were subsequently able to identify the active agent as CLA (3). Since then, additional biological effects of CLA have been described, and evidence now suggests that these fatty acids function as modulators of immune responses, cell growth, nutrient utilization, nutrient storage, and lipid metabolism [reviewed in Parodi (4), Pariza (5), Pariza et al (6), Kritchevsky (7), MacDonald (8), Pariza et al (9), and Belury (10)]. The list of purported benefits of CLA is impressive. However, the wide range of biological activities may not be the result of a single entity, or even the most common natural isomer, because, essentially, all the experiments before the late 1990s were performed with use of mixed isomers, each with potentially different properties. With the recent commercial availability of purified preparations of the c9,t11- and t10,c12-isomers of CLA, studies of the mechanism of CLA action that used purified isomeric forms of CLA is an active area (6). Recent evidence has suggested that both the c9,t11-CLA and the t10,c12-isomer have important biological activities. In particular, many laboratories are interested in the potential benefits of CLA with regard to atherosclerosis, because of the enormous societal effect of the disease. At least a part of the beneficial effects of CLA on the development of early atherosclerotic lesions could be the result of modification of hepatic lipoprotein metabolism by CLA. This review considers recent observations in this research area.

CONJUGATED LINOLEIC ACID AND ATHEROSCLEROSIS

To our knowledge, there are no epidemiologic studies to support a role for dietary CLA in the prevention of atherosclerosis. This lack of support is due, in part, to the analytic challenges involved in the measurement of CLA and its isomers. In addition, studies in animal models indicate that the protection afforded by dietary CLA could occur at amounts above those usually consumed by humans.

Animal studies suggested that CLA supplementation decreases the development of early atherosclerotic lesions, although in most studies the reduction in lesions was modest. All of the published studies have used the chemically prepared mixture of CLA isomers. As a result, no clear consensus has yet emerged on the beneficial effects of CLA, and some are of the opinion that it is still too early to draw any conclusion (11, 12).

In rabbits fed a high-fat diet (14% by weight, predominantly hydrogenated coconut oil) containing 0.1% cholesterol (13), supplementation with 0.5 g/d of an isomeric mixture of CLA (40% c9,t11-CLA, 40% t10,c12-CLA, and 20% other isomers) significantly reduced aortic atherosclerosis after 22 wk, based on the extent of aortic surface with lipid deposition and connective tissue involvement. More recent work provided evidence to suggest that low amounts of CLA (0.1–1% of calories) could even reverse early atherosclerotic lesions in the rabbit model (14). Indeed, as little as 0.05% CLA in the diet reduced the severity of lesions (graded visually using a 0–4 scale) (15), and the higher the CLA content of the diet, the lower the lesion severity score. The latter study is important because it is the first to indicate that CLA concentrations that can be achieved in a normal human diet can also influence atherogenesis in an animal model.

In hamsters (16), a mixed isomer CLA preparation (supplemented to 0.06%, 0.11%, or 1.1% of food energy) was provided in a 10% saturated fat, 0.12% cholesterol diet. After 11 wk, atherosclerosis was reduced (as measured by en face staining of the aortic region for lipid accumulation), although in this study the nonconjugated form of LA also reduced fatty accumulations. There was no evidence for a dose-response relation between CLA and the decrease in fatty streaks. Direct comparison of the effects of LA and CLA (again as mixed isomer) in hamsters on an even higher-fat diet (20% hydrogenated coconut oil, 0.12% cholesterol) (17) indicated that CLA was more effective than LA at reducing the extent of atherosclerotic lesions. Thus, although the protection afforded by CLA is not different from LA on some diets, the benefits can be more readily observed when the diet contains a higher saturated fat component.

In contrast with the rabbit and hamster studies cited earlier, one study in mice suggested that CLA has no effect on, or could even promote, atherosclerotic lesion development. In C57BL/6 mice fed an atherogenic diet, CLA supplementation at 2.5–5 g/d increased the development of fatty streak lesions in the aorta (18). Because there is considerable variability both within and among experimental model systems, the ability of CLA to modulate atherosclerosis must still be considered an open question.

The differences in diets and methods for assessment of atherosclerosis and the marginal benefits of CLA in some of the studies have so far left some doubt as to the overall potential of CLA in the management of atherosclerotic risk in humans. When evaluating experimental models, one should consider variables such as species susceptibility to disease, length of study, and the dietary saturated fat content in addition to CLA content and isomer composition. In addition, the method of assessment of fatty streak lesions and the site of lesions should be evaluated in each model system. Fatty streak lesions in hamsters form primarily in the ascending aorta (PL Mitchell, MA Langille, DL Currie, RS McLeod, unpublished observations, 2003), whereas the early lesions form nearer the aortic root in mice (19). Lesions in rabbits affect primarily the aortic arch and the descending thoracic aorta (13). Every effort should be made to control for observer bias to ensure that the atherosclerotic lesion assessment is as objective as possible.

We initiated in vivo studies of the role of the isomeric forms of CLA in atherogenesis in hamsters (Mitchell et al, unpublished observations, 2003). The high-fat (20% by weight) diets of Syrian golden hamsters were supplemented with 1% (by weight) LA, c9,t11-CLA, or t10,c12-CLA for 10 wk (n = 6 per group). There was no significant difference in the food consumption or final body weight of the animals on the 3 different diets at the end of the study. However, animals supplemented with t10,c12-CLA had higher plasma triacylglycerol (259 ± 19 mg/100 mL) and HDL-cholesterol concentrations (HDL cholesterol = 122 ± 9 mg/100 mL) than did LA-supplemented animals (triacylglycerol = 156 ± 26 mg/100 mL; HDL cholesterol = 95 ± 4 mg/100 mL) (P < 0.02, t test) (all values are ± SEM) when compared with the LA group (60% ± 11% of sections). Although only the lesion frequency in the t10,c12-CLA group was significantly different from the LA group (P < 0.05, t test), both CLA isomers had a tendency to decrease the extent of atherosclerosis in hamsters. These observations suggest that both of the commonly studied isomeric forms of CLA can affect atherogenesis even when improvements in the lipoprotein profile are not clearly evident. Furthermore, synergistic effects of isomers should not be discounted, because the equivalent amount of pure isomer could have quite different effects alone and in an isomeric mixture ( In animal models CLAs have shown variable effects on plasma lipids. Plasma triacylglycerol and cholesterol were reduced when mixed isomer CLA was added in the first rabbit and hamster atherosclerosis studies described earlier (13, 16). However, in a subsequent rabbit study CLA increased both cholesterol and triacylglycerol (14). In hamsters the plasma triacylglycerol increased and cholesterol decreased in one study (17), whereas in another study (15) CLA did not significantly change either lipid. In the single study of mouse atherosclerosis, CLA decreased triacylglycerols but did not affect cholesterol (18). One can only conclude that changes in plasma lipids were not consistent and do not appear to correlate with the presence of atherosclerosis in the CLA supplementation studies.

In assessing the changes caused by CLA it is important to include an appropriate control group supplemented with a nonconjugated diene fatty acid. In hamsters it was shown (21) that, compared with a group that received no fatty acid supplement, plasma cholesterol and triacylglycerol were significantly lower in CLA-supplemented groups. However, similar changes were observed in a group of animals supplemented with LA. In a separate study (22), hamsters were fed a 13% fat (by weight) diet supplemented with 0.66% (wt:wt) of either c9,t11-CLA or t10,c12-CLA, or with an equal mixture of both. Both the mixed isomer preparation and the t10,c12-CLA preparation decreased LDL cholesterol (by 20%) and HDL cholesterol (by 10%) but increased VLDL triacylglycerol (by 70%). The c9,t11-CLA by itself had no significant effect on plasma lipid limits. Gavino et al (20) found that feeding hamsters a mixed isomer of CLA in a high-fat diet significantly lowered plasma triacylglycerol and cholesterol concentrations compared with pure c9,t11-CLA or with an LA-supplemented diet. Although they did not directly examine the t10,c12-isomer alone, their study suggested that the active component was the t10,c12-isomer, perhaps in synergy with the c9,t11-isomer. More studies of the comparison of the common isomers of CLA with each other and with LA are necessary to adequately demonstrate the biological importance of the individual isomers of CLA.

In the limited human studies performed so far, few changes were observed in the plasma lipid concentrations. Blankson et al (23) found no changes in plasma lipids at up to 6.8 g/d CLA supplement for 12 wk. Smedman and Vessby (24) found that feeding 53 healthy men and women 4.2 g/d of a CLA supplement for 12 wk did not affect serum lipid or lipoprotein concentrations even though body fat was decreased by 3.8%. A 2-mo trial of CLA supplementation (3.9 g/d) in 17 normolipidemic women (25) did not show any changes in plasma lipids or lipoproteins, although the CLA preparation used in that study contained only 10–15% of each of the c9,t11- and t10,c12-isomers. One study (26) found that 8 wk of CLA supplements (0.7 g/d for 4 wk and then 1.4 g/d for 4 wk) decreased plasma cholesterol, plasma triacylglycerols, and HDL cholesterol in normolipidemic volunteers. Taken together, these observations indicate that, if there are changes associated with mixed isomer CLA supplements in humans, they are small and not necessarily toward a more cardioprotective lipid profile.

A recent study compared 2 different isomer mixtures in a human trial. Mixed isomer CLA (80% c9,t11-CLA plus 20% t10,c12-CLA; 80:20) feeding for 8 wk decreased VLDL cholesterol, and a 50:50 mixture decreased fasting plasma triacylglycerols in a study of 51 normolipidemic human subjects (27). There were no effects on HDL cholesterol, LDL cholesterol, glucose, or insulin concentrations, as well as body weight, in that study, suggesting that CLA could have cardioprotective effects on human lipoprotein metabolism. The investigators suggested that the t10,c12-isomer could mediate the triacylglycerol-lowering effects of CLA but that neither isomer seems to have an appreciable effect on cholesterol concentration in normolipidemic human subjects.

CONJUGATED LINOLEIC ACID AND HEPATIC VERY-LOW-DENSITY LIPOPROTEINS

One of the potential mechanisms by which CLA could modulate atherosclerosis is by modifying the production of atherogenic lipoproteins by the liver. Hepatic VLDLs are assembled by the noncovalent association of phospholipids and neutral lipids (predominantly triacylglycerol) with the large, hydrophobic apolipoprotein B (apo B) [reviewed in Yao and McLeod (28)]. In the circulation, VLDL is catabolized to LDL, and an increase in the concentration of LDL is directly correlated with an increased risk of developing cardiovascular disease in humans. Assembly of VLDL appears to occur in 2 steps (29, 30). Initially, apo B associates with a small amount of neutral lipid to form a nascent lipoprotein particle within the lumen of the endoplasmic reticulum. Subsequently, a large amount of triacylglycerol is added to this precursor to form the mature VLDL. Kinetic evidence has shown that the majority (70%) of the triacylglycerol that is secreted in VLDL must first enter cytosolic stores in the hepatocyte before subsequent hydrolysis and re-esterification into secretory VLDL-triacylglycerol (31-34). Thus, hepatic VLDLs contain esters (predominantly triacylglycerol) of the fatty acids consumed in the diet and, at least temporarily, stored in the liver. Plasma LDL concentrations are affected by the fatty acid composition and the cholesterol content of the diet (35), and unesterified fatty acids are known to increase the assembly and secretion of VLDL from cultured hepatoma cells (29, 36, 37). Enhanced fatty acid flux to the liver was hypothesized to play a role in diseases characterized by enhanced secretion of hepatic VLDL (38).

The mechanism or mechanisms by which CLA can affect VLDL secretion is (are) not known, but there are indications from cell culture studies that CLA could affect hepatic VLDL assembly and secretion. Oleic acid (18:1) is known to enhance the secretion of apo B from HepG2 cells by decreasing the posttranslational degradation of the nascent polypeptide (39, 40). In contrast, oleate stimulates triacylglycerol secretion but not the secretion of apo B in primary rat hepatocytes (41). Further work is required to determine whether CLA can affect the assembly of hepatic VLDL by altering the number or the triacylglycerol content of the hepatic lipoproteins.

To gain insight into the mechanistic features of CLA action in the liver, the human hepatoma cell line HepG2 was used to model the hepatic assembly of apo B–containing lipoproteins. Experiments in HepG2 cells suggested that t10,c12-CLA decreases apo B (42) and triacylglycerol (43) secretion but could increase (43) or decrease (42) triacylglycerol biosynthesis. The secretion of apo B and triacylglycerol in the presence of the c9,t11-CLA isomer was similar to LA (43). These data suggest that there are isomer-specific effects of CLA on hepatic lipid and apo B metabolism that could lead to a beneficial lowering of hepatic VLDL secretion. However, if the t10,c12-isomer decreases triacylglycerol secretion without decreasing its synthesis, there is the potential for hepatic accumulation of triacylglycerol, as described in mice (44). One should bear in mind, however, that HepG2 cells may not be an ideal model for the study of VLDL assembly and secretion because the mobilization of triacylglycerol from cytosolic stores is already severely compromised (45, 46). Further work in other hepatic cell models is clearly needed to fully understand the effect of CLA isomers on hepatic lipid and lipoprotein metabolism.

We studied the ability of CLA to stimulate the assembly of hepatic VLDL in rat hepatoma cell McA-RH7777. Both c9,t11-CLA and t10,c12-CLA are incorporated into cellular triacylglycerol, as demonstrated by molecular species analysis of the lipid extracts from cells after treatment of the cultures with exogenous CLA. McA-RH7777 cells secrete authentic VLDL when oleic acid is added to the culture medium (29). Likewise, when CLA was added as the sole source of exogenous fatty acid, c9,t11-CLA and t10,c12-CLA isomers also stimulated the secretion of VLDL (AM LeBlanc, DL Currie, RS McLeod, unpublished observations, 2003). This secretion was despite the observation that the t10,c12-isomer was a less efficient stimulus for triacylglycerol secretion than was the c9,t11-isomer (43). Studies are currently under way to examine the effects of the different CLA isomers and mixtures of isomers on the esterification and oxidation of other fatty acids in rat hepatoma cells.

It is important to distinguish between the hepatic secretion of triacylglycerol and the secretion of its apo B component, because the liver can secrete VLDL with a variable neutral lipid load. In HepG2 cells, both CLA isomers stimulated triacylglycerol biosynthesis (43), but only the t10,c12-isomer decreased triacylglycerol secretion. If this situation were to persist, there is potential for the accumulation of triacylglycerol in the liver, causing hepatomegaly as a result of steatosis. In an important study in mice (44), the t10,c12-CLA isomer caused massive fatty liver and hyperinsulinemia, whereas the c9,t11-isomer did not have this effect. The investigators suggested that the hyperinsulinemia could be responsible for the increase in hepatic neutral lipid stores by way of increased fatty acid uptake and lipogenesis. Similar observations of severe hepatomegaly and a 10-fold increase in liver triacylglycerol stores were reported in another mouse study that used mixed isomer CLA (47).

Nevertheless, the hepatic steatosis found in mice was not observed in other animals. Sprague Dawley rats are less susceptible to the CLA-induced hepatic steatosis (at least with mixed isomer preparations) (48, 49). A 30% decrease in hepatic triacylglycerol and a 26% decrease in liver cholesterol were observed in Otsuka Long Evans Tokushima Fatty (OLETF) rats fed dietary CLA (50). Changes in liver triacylglycerol concentrations with CLA supplementation may not be as pronounced in human studies. Although changes in liver physiology must be monitored, one might anticipate that the changes observed in mice are related to the high rate of adipose tissue triacylglycerol turnover in this animal. As a proportion of the total body triacylglycerol, the adipose tissue alterations in human studies are anticipated to be much less than those in mice.

Central to the effects of CLA appears to be their modulation of the metabolic fate of hepatic fatty acids. Belury and Kempa-Steczko (51) suggested, based on studies in rats and mice, that CLA could affect the metabolism of fatty acids in the liver and modify the fatty acid composition of triacylglycerol and phospholipids in the cell, perhaps by stimulation of peroxisome proliferator-activated receptor (PPAR)–responsive genes (48, 52). Hamster experiments indicated that CLA does not increase enzymatic activities associated with peroxisome proliferation (22). Mixed CLA isomers fed at 1% concentration to rats decreased the secretion of triacylglycerol and cholesterol and increased ketone production (53) in part by increasing ß-oxidation and decreasing esterification. Moya-Camarena et al (48) found that acyl-CoA oxidase messenger RNA concentrations were increased 3-fold in CLA-fed rats and in OLETF rats. Dietary CLA increased the activity of mitochondrial carnitine palmitoyltransferase and decreased that of phosphatidate phosphohydrolase in the liver (50). These observations suggest that, at least in rats, CLA can both increase fatty acid oxidation and decrease esterification to triacylglycerol (54). Both of the common CLA isomers are rapidly imported into isolated mitochondria but are oxidized more slowly than either LA or palmitoleic acid (55), and it was suggested that treatment of the cells with CLA could decrease the oxidation of other fatty acids. Takahashi et al (47) found that CLA feeding increased the hepatic enzymes involved in both fatty acid synthesis (eg, acetyl-CoA carboxylase, fatty acid synthase) and fatty acid oxidation (eg, acyl-CoA oxidase, carnitine palmitoyltransferase). Banni et al (56) found that CLA feeding increased the conjugated fatty acid content of the liver neutral lipid pool, suggesting that both isomers are substrates for esterification into triacylglycerol and cholesteryl esters. In contrast, both the c9,t11-CLA and the t10,c12-CLA were more readily oxidized than linoleic acid in rats (57). Thus, the net effects of CLA on liver lipoprotein production could reflect how the liver metabolizes other fatty acids in the presence of CLA.

Changes in fatty acid metabolism were detected in adipocyte and intestinal enterocyte models. In Wistar rats (58) the t10,c12-isomer increased epididymal adipose tissue carnitine palmitoyltransferase-I activity by 30%, even though this rat strain was not very responsive to CLA feeding. The t10,c12-CLA isomer increased both the esterification and the oxidation of tracer oleic acid in 3T3-L1 preadipocytes (59), and in human adipose tissue cultures, the t10,c12-CLA isomer, but not the c9,t11-isomer, decreased triacylglycerol biosynthesis (60). On the basis of observations in 3T3-L1 cells, it was suggested that CLA could compete with LA for incorporation into triacylglycerol and phospholipids (61). In the intestinal enterocyte model Caco-2 (62), chronic supplementation with t10,c12-CLA increased de novo triacylglycerol biosynthesis but did not affect triacylglycerol secretion, whereas supplementation with the c9,t11-CLA isomer did not affect triacylglycerol metabolism.

On the basis of the work so far it is difficult to generalize about the effects of CLA on hepatic fatty acid metabolism. Several studies indicated that there are paradoxical increases in the ability of the hepatocyte to both oxidize fatty acids and incorporate them into esters such as triacylglycerol. Whether oxidation or storage prevails in a particular study could, in part, be related to the other fatty acid components of the diet. Additional metabolic factors could play a role, such as the t10,c12-CLA–induced hyperinsulinemia (44). Further mechanistic studies in cell culture could shed additional light onto the role of CLA in the fate of hepatic fatty acids.

MECHANISMS OF CONJUGATED LINOLEIC ACID ACTION

Considerable effort is now in progress to understand the mechanisms of the changes associated with CLA supplementation. Much of the work so far centers on the role of CLA as PPAR ligands. PPAR is an important member of this nuclear receptor family that is expressed in the liver and modulates hepatic lipid metabolism. With use of a scintillation proximity assay, CLAs were shown to be high-affinity fatty acid ligands for PPAR. The results also suggested that the c9,t11-CLA isomer could be a more potent ligand and activator of PPAR than t10,c12-CLA (52). Although CLA can activate PPARs, this activation did not cause peroxisome proliferation in rats (48). Furthermore, a study in PPAR-null mice indicated that the changes caused by a 0.5% CLA diet (in which body fat content and plasma triacylglycerol were reduced) were independent of PPAR (63). Thus, although liver PPAR is an important target of CLA, at least some of the effects of CLA could involve other mechanisms.

Another potential target of CLA that has received considerable attention is stearoyl-CoA desaturase (SCD). This enzyme is responsible for the introduction of a cis- double bond at the C9 position of fatty acyl-CoA, primarily stearoyl-CoA (18:0) and palmitoyl-CoA (16:0). Both common CLA isomers inhibit SCD activity and reduce levels of the protein in breast tumor cell lines (64), and the t10,c12-CLA isomer (but not c9,t11-CLA) decreased SCD activity and the production of monounsaturated fatty acids in HepG2 cells (65). t10,c12-CLA caused a dose-dependent decrease in the expression of SCD1 messenger RNA in 3T3-L1 cells, and this change was greater than the effect of the c9,t11-isomer (66). Phenotypic changes were also evident, as t10,c12-CLA treatment produced adipocytes with smaller lipid droplets than did the c9,t11-CLA treatment. It was suggested (67) that the c-12 double bond is important in the inhibition of SCD. CLA also affects desaturases other than SCD, as the t10,c12-isomer also decreased the activities of ⁵- and ⁶-desaturases in HepG2 cells (68). Furthermore, in rat liver microsomes (69), both c9,t11-CLA and t10,c12-CLA also decreased ⁶-desaturation.

Other potential mechanisms of CLA action were suggested. Dietary CLA decreased intestinal sterol O-acyltransferase in hamsters (21). Hepatic expression of sterol regulatory element-binding protein-1c and liver X receptor- was down-regulated by the c9,t11-isomer in the ob/ob mouse (70), whereas the t10,c12-isomer had no effect on these 2 transcripts. These observations suggest that CLA could affect cellular cholesterol metabolism as well and that by way of changes in hepatic gene expression, the c9,t11-CLA could, in the absence of other isomers, improve hepatic lipid metabolism.

CONCLUSIONS

In summary, despite nearly a decade of study of the role of CLA in modulating the development of atherosclerosis and hepatic lipoprotein metabolism, considerable uncertainty still exists as to the potential cardiovascular benefits or risks associated with these unusual fatty acids. No consistent improvement in the plasma lipid or lipoprotein profiles has been detected in animal or human studies, although emerging evidence suggests that CLA could affect the metabolism of fatty acids in the liver. More work is needed to understand the mechanisms of CLA action in the hepatocyte, especially with regard to isomer-specific effects on hepatic lipid metabolism and the role of gene expression patterns that can be altered by CLA. Of some concern are the observations of hepatic steatosis, particularly in mouse studies, when there are substantial reductions in adipose tissue mass. Considerable progress was made in studies of the mechanistic bases of CLA action in the liver and the adipocyte, especially the role of PPARs. As animal and cell culture observations increase our knowledge of how CLA affects lipid and lipoprotein metabolism, it will become easier to assess the potential risks and benefits of dietary CLA in humans.

ACKNOWLEDGMENTS

The authors had no existing or potential conflicts of interest with the financial sponsors of the research and writing of this article.

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