1. Field of the Invention
The present invention relates generally to the fields of lipid metabolism and dietary supplementation. More particularly, it concerns compositions and methods for inhibiting an increase in serum arachidonic acid of a mammal to which .gamma.-linolenic acid (GLA) has been provided.
2. Description of Related Art
Arachidonic acid (AA) is a polyunsaturated fatty acid found in relatively small quantities in membranes of mammalian cells. Research over the last four decades has shown that the in vivo modulation of levels of arachidonic acid and oxygen-containing derivatives of arachidonic acid (known as eicosanoids) is intimately liked to human disease (for a review, see Samuelsson et al., 1987 and Chilton et al., 1997). For example, during inflammation, low levels of certain arachidonic acid derivatives render a protective response leading to enhanced disease resistance. However, these same molecules induce an autotoxic response leading to a variety of inflammatory disorders when produced in excessive quantities. Over the past three decades, the therapeutic utility of blocking the metabolism of arachidonic acid through multiple pathways including 5-lipoxygenase and cyclooxygenase I and II has become evident for the treatment of a wide range of inflammatory disorders.
Since arachidonic acid or its precursors found in cells and tissues must be derived from diets, it follows that diet may affect diseases controlled by arachidonic acid or its derivatives. This relationship was suggested in the 1960s by studies which showed differences in frequencies of inflammatory disorders among Greenland Eskimos and Danes (Chilton et al., 1996a; Dyerberg and Bang, 1979). Later studies showed similar differences between Japanese and Americans. These differences (Danes and Americans have much higher frequencies of inflammatory disorders including asthma, arthritis, psoriasis and acute myocardial infarction) were attributed, in large part, to the consumption by Danes and Americans, on Western diets, of high dietary quantities of precursor fatty acids of arachidonic acid (termed n-6 fatty acids) and arachidonic acid, offset by the low consumption of n-3 fatty acids.
Based on these observations, a number of dietary fatty acid reduction and supplementation strategies were undertaken in an attempt to influence arachidonic acid metabolism, eicosanoid production and clinical outcomes. These studies carried out over the last two decades have revealed that controlling dietary fatty acid intake in a number of animal models has great potential in reducing eicosanoid synthesis and ameliorating inflammation in models which mimic human arthritis, asthma or glomerulonephritis (Prickett et al., 1981; Kelley et al., 1985; Lefkowith et al., 1990; Rovin et al., 1990; Hurd et al. 1981).
These studies demonstrated that the formation of derivatives of AA and the subsequent effects of these compounds (eicosanoids) on cells and tissues are central processes in inflammation and allergy. Dietary fatty acid reduction and supplementation strategies have been utilized in animals and humans in an attempt to modulate cellular AA levels and metabolism and to ameliorate clinical inflammatory disorders. However, dietary modifications in humans on Western diets have shown only modest efficacy. If these observations are to prove useful in the treatment of such disorders, it is necessary to find more efficient dietary strategies to reduce eicosanoid generation in humans and to determine the mechanism(s) leading to this reduction.
In terms of inflammation, at least four dietary reduction and supplementation strategies have been utilized in both animals and humans in an attempt to influence eicosanoid production and clinical outcomes. One strategy has been to supplement "normal" diets with n-3 fatty acids. Here, there has been some controversy as to how effective these fatty acids are in reducing lipid mediators (eicosanoids) of inflammation (Chilton et al., 1993; Sperling et al., 1987; Strasser et al, 1984; Kojima et al., 1991; Galloway et al., 1985; Mori et al., 1987; Ahmed and Holub, 1984; Payan et al., 1986; Rosenthal, and Hills, 1986; Triggiani et al., 1990). For example, several studies report only modest inhibition of leukotrienes and PAF after n-3 fatty acid supplementation, while other investigations report more dramatic reductions (Chilton et al., 1993; Sperling et al., 1987; Strasser et al., 1984). The basis for these discrepancies is unclear at this time. In addition to eicosanoids, n-3 fatty acids have been shown to affect processes such as gene expression, cytokine generation and programmed cell death in a number of in vitro and in vivo settings (Endres et al., 1989; Clarke and Jump, 1996; Chandrasekar et al., 1995; Fernandes et al., 1994).
A second strategy to affect changes in AA metabolism in humans has been to remove dietary essential fatty acids from the diet. This eliminates sources of cellular AA that are derived from dietary linoleic acid (LA). Severe restrictions of LA intake in infants result in significant falls in levels of prostaglandin metabolites (Friedman et al., 1978). Wene and colleagues studied healthy men on fat-free eucaloric diets and found that LA levels in serum components fell dramatically within seven days of starting the diet (Wene et al., 1975). However, if calorie intake was then reduced (intermittent fasting), LA levels in serum increased. This LA repletion is probably due to mobilization of fatty acids from adipose tissue triglycerides.
A third strategy to reduce AA metabolism has been to restrict preformed AA in diets of humans. There are several conflicting studies in humans restricting preformed AA by the chronic avoidance of animal tissue with results varying from increases to moderate reductions in serum AA levels (Phinney et al., 1990; Sanders et al., 1978; Melchert et al., 1987). In contrast to studies restricting dietary AA, humans supplemented with AA (an additional 6 g/day) exhibit a pronounced increase in AA levels within plasma triglycerides, phospholipids, cholesterol esters, and platelet phospholipids (Seyberth et al., 1975). This increase within complex lipids is accompanied by an increase in eicosanoid generation and a marked decrease in the ADP threshold dose required to induce platelet aggregation.
A fourth strategy that has been utilized to influence AA metabolism is to supplement normal diets with oils (primrose and borage) rich in gamma linolenic acid (18:3, n-6). Such oils have been shown to improve clinical symptoms of patients with atopic dermatitis and rheumatoid arthritis (Leventhal et al., 1993; Miller et al., 1990; Horrobin, 1992; Zibok and Fletcher, 1992; Tate et al., 1989). The mechanisms by which GLA influences these inflammatory disorders has not been elucidated. In fact, it is paradoxical that providing a dietary precursor of AA, GLA, attenuates inflammation. It is known that a portion of GLA provided is elongated (by 2 carbons) in vivo to form dihomogammalinolenic acid (DGLA) (Horrobin, 1992; Zibok and Fletcher, 1992; Tate et al., 1989). DGLA can then be metabolized to oxygenated products, 15-OH-20:3. (15 HETrE) and prostaglandin E.sub.1 by 15 lipoxygenase and cyclooxygenase, respectively (Miller et al., 1990; Horrobin, 1992). PGE.sub.1 has been found to be anti-inflammatory in a variety of in vitro systems and animal models (Kerins et al., 1991). GLA supplementation also has been shown to reduce the capacity of some cells to produce AA-derived eicosanoids (Leventhal et al., 1993; Zibok and Fletcher, 1992).
Over the last six years, the inventor's laboratory has provided humans, on controlled diets, with a wide range of dietary fatty acid supplements and supplement combinations in an attempt to affect AA metabolism in humans (Chilton et al., 1993; Triggiani et al., 1990; Chilton-Lopez et al., 1996; Johnson et al., 1997). In these studies, the inventor has utilized well defined diets (prepared and fed in a General Clinical Research Center [GCRC]) and measurement techniques (negative ion chemical ionization GC/MS), which precisely determine fatty acid and eicosanoid levels in serum and inflammatory cells.
Although much work has been performed on the dietary supplementation of fats, many questions remain to be answered, including the determination of the capacity of different inflammatory cells to synthesize (elongate and desaturate) polyunsaturated fatty acids; the major mechanism(s) by which analogs (which can be induced by dietary supplementation) of AA influence eicosanoid generation and the development of dietary strategies that will produce natural antagonists of AA in inflammatory cells thereby reducing the synthesis of pro-inflammatory eicosanoids without increasing serum levels of AA. These and other questions are addressed by the present invention.