Andrzej Pronczuk

Differences in the plasma transport and tissue concentrations of tocopherols and tocotrienols : observations in humans and hamsters

Abstract Certain aspects of tocopherol and tocotrienol absorption, plasma transport, and tissue distribution were examined in humans and hamsters. Plasma transport differed in that tocopherols were found primarily in low density lipoprotein and high density lipoprotein in association with plasma surface components, whereas tocotrienols disappeared from plasma with chylomicron clearance. In keeping with transport by triglyceride-rich lipoproteins, tocotrienols were deposited in conjunction with triglycerides in the adipose tissue of hamsters. In hamsters, tocopherols were the only tocol readily detected in all tissues, except adipose during tocotrienol supplementation. In fasting humans, the plasma tocotrienol concentration was not significantly increased after tocotrienol supplementation, whereas the platelet concentration of δ-tocotrienol doubled. Furthermore, tocotrienol intake did not appear to modulate the plasma cholesterol concentration in normolipemic hamsters. Thus, the transport, tissue concentration, and relative biologic function of tocopherol and tocotrienol appear somewhat disparate and possibly unrelated.

Dietary myristic, palmitic, and linoleic acids modulate cholesterolemia in gerbils.

Previous studies with cebus monkeys and review of published human data indicated that 85% to 90% of the variation in plasma cholesterol (TC) could be explained on the basis of dietary myristic (14:0) and linoleic (18:2) acid intake in the absence of cholesterol, and that 16:0 contributed to cholesterolemia as dietary cholesterol was increased. Because monkeys are a limited resource, a more convenient, sensitive model was sought for investigating these dietary fatty acid and plasma lipid relationships. Accordingly, this report describes the results of multiple regression analysis of the TC response to dietary fatty acids based on 319 young, male Mongolian gerbils fed a total of 59 purified diets supplying about 40% energy as fat from single or blended fat sources. Gerbils (6‐16 animals per dietary group) were fed purified diets (21 with 0.01 to 0.08% cholesterol and 38 cholesterol‐free) for 4‐week periods. When cholesterol‐free diets were fed, dietary 14:0 and 18:2 together accounted for 89% of the observed variation in TC. Although 14:0 consumption increased TC in a linear manner, the independent ability of 18:2 to lower cholesterol was nonlinear and exhibited a threshold effect at 5‐6% dietary energy, above which further lowering of TC was less remarkable. In gerbils consuming cholesterol‐supplemented diets, 87% of the observed variation in TC could be accounted for by a regression equation that included 14:0, palmitic acid (16:0), and a log function of 18:2 plus dietary cholesterol itself. These results demonstrate the applicability of gerbils for such studies and confirm previous observations in monkeys and humans that dietary 14:0 and 18:2 are the main fatty acids modulating plasma cholesterol under normocholesterolemic circumstances (i.e., when consuming low‐cholesterol diets and lipoprotein metabolism is normal) whereas 16:0 also appears modestly hypercholesterolemic when LDL receptors are compromised (i.e., when dietary cholesterol or certain metabolic factors have encumbered lipoprotein metabolism).—Pronczuk, A., Khosla, P., Hayes, K. C. Dietary myristic, palmitic, and linoleic acids modulate cholesterolemia in gerbils. FASEB J. 8, 1191‐1200 (1994)

Species variation in the atherogenic profile of monkeys: Relationship between dietary fats, lipoproteins, and platelet aggregation

Because lipoproteins and platelet aggregation have been implicated in atherogenesis, relative differences in the response of these variables to dietary fat saturation were compared in three species of monkeys differing in their susceptibility to atherosclerosis (cebus, rhesus, and squirrel monkeys). Both long-term (8–12 years) and short-term (8 weeks) responses to diets containing 31% fat calories were examined in the same monkeys. As expected, long-term feeding of coconut oil by comparison to corn oil produced significantly higher plasma concentrations of total cholesterol, LDL cholesterol, apoB, and triglycerides, as well as higher ratios of LDL/HDL cholesterol and apo B/apo A-I. These responses were characteristic of all species with cebus being most responsive and rhesus the least. The shortterm plasma cholesterol response to animal fats (butter, lard, beef tallow) was significantly less than that to coconut oil. When fish oil was substituted for two-thirds of either corn oil or coconut oil, exceptional decreases occurred in plasma cholesterol and triglycerides, as well as in HDL cholesterol and apo A-I concentrations despite the fact that the fish oil diets contained more saturated fat and less polyenes than the corn oil diet. Platelet aggregation tended to increase with saturated fat consumption and greatly decreased with fish oil intake in all monkeys, although cebus monkeys were ten-fold more resistant to platelet aggregation than the other two species. The molecular species of platelet phosphatidylcholine (PC) varied with both the dietary fat fed and species of monkey. An inverse correlation (r=−0.60; p<0.001) was found between changes in one such PC molecular species (18∶0−20∶4) induced by diet and the platelet aggregation threshold. These results demonstrate that the lipemic and platelet responses to dietary saturated fat depend upon both the type of fat (i.e., the specific combination of dietary fatty acids, including the chain length of saturated fatty acids and the degree of polyunsaturation) and the species of monkey (genetic component) in which the response is elicited.

Dietary palmitic acid (16:0) enhances high density lipoprotein cholesterol and low density lipoprotein receptor mRNA abundance in hamsters.

Abstract In order to examine the qualitative effect of different fats and specific fatty acids on plasma lipids and lipoprotein metabolism, six low fat, cholesterol-free diets were fed to young male hamsters (10/group) for a 4-week period. Fat blends were formulated with coconut oil, palm oil, soybean oil, high oleic acid safflower oil, butter, corn oil, and canola oil. Diets contained 13% energy as fat and dietary polyunsaturate/saturate ratios ranged from 0.12 to 1.04, one of which incorporated the American Heart Association-recommended concentrations of saturates, monoenes, and polyenes and another reflected the current American Fat Blend. In three diets the polyunsaturate/monounsaturate/saturate ratio was held constant while only the 12:0, 14:0, and 16:0 were varied. Plasma lipoproteins and apoproteins were assessed in conjunction with the abundance of specific hepatic and intestinal mRNA for the low density lipoproteins (LDL) receptor and various apolipoproteins associated with cholesterol metabolism. The plasma cholesterol response was lowest with the American Heart Association blend and equally elevated by the more saturated, low polyene diets (polyunsaturate/saturate, 0.12–0.38). Replacing 12:0 plus 14:0 from coconut oil with 16:0 as palm oil induced a significant increase in high density lipoprotein (HDL) cholesterol with a trend toward decreased LDL. These shifts in lipoprotein cholesterol were corroborated by measures of the LDL/HDL ratio, the plasma apolipoprotein B/apolipoprotein A1 ratio, and differences in the synthesis of apolipoproteins and the LDL receptor based on estimates of the mRNA for these proteins in the liver and gut, using specific cDNA probes for apolipoprotein A1, apolipoprotein B, apolipoprotein E, and the LDL receptor. Although it has been suggested that dietary polyenes lower total plasma cholesterol, including HDL, and that saturated fat increases both these pools of cholesterol, the current data represent the first evidence that a specific saturated fatty acid, i.e., palmitic acid, may enhance HDL production.

A rationale for plasma cholesterol modulation by dietary fatty acids: Modeling the human response in animals

Abstract At low levels of dietary cholesterol intake ( 500 mg/day), the plasma cholesterol response is no longer described accurately by dietary 14:0 and 18:2 alone. In such situations 16:0 appears to contribute to plasma cholesterol elevation. The hypercholesterolemic potential of 16:0, possibly reflecting a synergism between dietary cholesterol and 16:0, is thought to reside, in part, in the ability of 16:0 to increase the transport of very low density lipoprotein (VLDL) apoB. Increased production of VLDL, coupled with impaired LDL receptor activity, results in an expansion of the LDL pool when the ability to clear VLDL remnants is impaired. Evidence is also available to suggest that the position of saturated fatty acids on the TG molecule affects its hypercholesterolemic ability. An argument is made for selecting animal models for investigation of the fat saturation effect based initially on the total plasma cholesterol (TC) response, with subsequent emphasis being placed on lipoproteins and the actual control mechanism(s) once the generic similarity in the TC response with that in humans has been established.

Replacing Trans Fat: The Argument for Palm Oil with a Cautionary Note on Interesterification

To replace dietary trans fatty acids (TFA), two practical options exist: revert to a natural saturated fat without cholesterol (most likely palm oil or its fractions) or move to a newer model of modified fat hardened by interesterification (IE). This review summarizes the relative risks for cardiovascular disease inherent in these options. Interestingly, both types of fat have been the subject of nutritional scrutiny for approximately the last 40 years, and both have positive and negative attributes. Only during that period has palm oil production developed to the point where it has become the major edible oil in world markets, making clinical studies of it an important objective. On the other hand, approximately 25 human studies have fed interesterified fat in one form or another over this period, some for weeks, some as a single meal. Two types of diet designs exist. Several fed a small amount of interesterified fat, usually incorporated within a margarine, and stayed below the radar of biological detection of any abnormal metabolism. A few fed interesterified fat that incorporated stearic acid, as interesterified 18:0 (IE-18:0), even comparing it to trans fat and saturated fat, as a major part of total daily calories to assess its metabolic impact per se. These latter 5 to 6 studies clearly reveal negative biological effects on lipoproteins, blood glucose, insulin, immune function, or liver enzymes when relatively high intake of IE-18:0 or palmitic acid (IE-16:0) were fed in fats with sn2–saturated fatty acids. High intake of 18:0 in natural fats can depress total lipoproteins, while IE-18:0 and IE-16:0 at high levels adversely affect lipoprotein metabolism. Still other studies have supplied interesterified fat as a single meal or fed such fat daily only in a single snack, as opposed to incorporating the fat into the entire fat pool consumed at all meals in association with most foods (which is the more physiological approach and more apt to elicit effects). Even in meal studies, IE-18:0 typically delayed fat absorption postprandially, indicating its effect on fat metabolism originating, in part, in the intestine. Mainly 2 saturated fatty acids (18:0 or 16:0) have been interesterified to harden oils, using the 16:0 from fully hydrogenated palm oil or 18:0 from fully hydrogenated soybean oil as the source material. It is not clear that IE-16:0 is as problematic as IE-18:0, but IE-16:0 has been studied less. Levels between 8% energy (%E) and 12%E from 18:0 as interesterified fat (the typical diet provides about 2%E–4%E as 18:0 from natural fats) show the most effect. Detection of adverse effects would seem to start around 7%E–8%E as IE-18:0, but one can assume that effects are initiated, even if undetected, at a lower intake, similar to the situation with TFA. Thus, although an intake of 1%E to 4%E from IE-18:0 does not appear to influence lipoproteins, it is not necessarily the only system affected. The negative effects of IE-18:0 may be alleviated or masked by dilution with other fats, especially by adding 18:2-rich polyunsaturated oils to the diet. This is similar to the trans fat story, i.e., if a limited intake of TFA is heavily diluted with other oils, the consumption of TFA fails to be detected as an adverse effect. Accordingly, more research is warranted to determine the appropriateness of interesterified fat consumption, particularly before it becomes insidiously embedded in the food supply similar to TFA and intake levels are achieved that compromise long-term health.

An animal model of spontaneous metabolic syndrome: Nile grass rat

Metabolic syndrome (MetS) is a prevalent and complex disease, characterized by the variable coexistence of obesity, dyslipidemia, hyperinsulinaemia, and hypertension. The alarming rise in the prevalence of metabolic disorders makes it imperative to innovate preventive or therapeutic measures for MetS and its complications. However, the elucidation of the pathogenesis of MetS has been hampered by the lack of realistic models. For example, the existing animal models of MetS, i.e., genetically engineered rodents, imitate certain aspects of the disease, while lacking other important components. Defining the natural course of MetS in a spontaneous animal model of the disease would be desirable. Here, we introduce the Nile grass rat (NGR), Arvicanthis niloticus, as a novel model of MetS. Studies of over 1100 NGRs in captivity, fed normal chow, revealed that most of these animals spontaneously develop dyslipidemia (P < 0.01), and hyperglycemia (P < 0.01) by 1 yr of age. Further characterization showed that the diabetic rats develop liver steatosis, abdominal fat accumulation, nephropathy, atrophy of pancreatic islets of Langerhans, fatty streaks in the aorta, and hypertension (P < 0.01). Diabetic NGRs in the early phase of the disease develop hyperinsulinemia, and show a strong inverse correlation between plasma adiponectin and HbA1c levels (P < 0.01). These data indicate that the NGR is a valuable, spontaneous model for exploring the etiology and pathophysiology of MetS as well as its various complications.—Noda, K., Melhorn, M. I., Zandi, S., Frimmel, S., Tayyari, F., Hisatomi, T., Almulki, L., Pronczuk, A., Hayes, K. C, Hafezi‐Moghadam, A. An animal model of spontaneous metabolic syndrome: Nile grass rat. FASEBJ. 24, 2443–2453 (2010). www.fasebj.org

Saturated fatty acids and LDL receptor modulation in humans and monkeys

It has been known for 40 years that dietary saturated fat (SAT FAT) increases plasma cholesterol, including LDL-C and HDL-C. In humans, where LDL-C is typically > 90 mg/dl this SAT FAT effect largely reflects changes in LDL-C pool size. The original human studies suggested that LDL-C expansion during SAT FAT consumption reflected reduced LDL clearance (LDL receptor activity) in hyperlipemics and increased LDL production rates in normolipemics (LDL-C < 100 mg/dl) . This dual explanation is supported by data from several animal models where specific saturated fatty acids (SFAs) have been the focus. However, the situation is complicated by the fact that polyunsaturated fatty acids (PUFAs) oppose SFAs, i.e. PUFAs decrease LDL-C and increase LDL receptor (LDLr) activity, so the effect of SAT FAT intake may represent the combined influence of increased SFAs and decreased PUFAs. In fact, careful scrutiny of primate data suggests a negligible effect of saturated fat on LDL clearance (and receptor activity) in the absence of dietary cholesterol when PUFA intake is adequate (5-10%en) and the lipoprotein profile is relatively normal (LDL-C < 90 mg/dl), i.e. normolipemic situations at the time of dietary intervention. In such cases increases in LDL-C due to SFAs (particularly 12:0+14:0) appear to reflect LDL overproduction associated with a shift in cholesterol from tissues to the plasma cholesteryl ester (CE) pool (both LDL-C and HDL-C) without altering whole-body cholesterol balance. The reason for this shift, which is accompanied by an increase in the plasma oleic/linoleic CE ratio, is unknown but may reflect a decreased rate of CE hydrolysis by the liver. When individuals or animals are rendered hyperlipemic by other factors (e.g. chronic caloric and dietary cholesterol excesses in humans or by cholesterol feeding in animals) specific SFAs (particularly 16:0) can contribute to decreased LDLr activity initiated by a primary factor, such as dietary cholesterol. However, LDLr down-regulation by dietary cholesterol greatly exceeds any contribution from SFAs.

Nutritional correlates and dynamics of diabetes in the Nile rat (Arvicanthis niloticus): a novel model for diet-induced type 2 diabetes and the metabolic syndrome.

BackgroundThe prevalence of Metabolic Syndrome and related chronic diseases, among them non-insulin-dependent (type 2) diabetes mellitus, are on the rise in the United States and throughout the world. Animal models that respond to environmental stressors, such as diet, are useful for investigating the outcome and development of these related diseases.ObjectiveWithin this context, growth and energy relationships were characterized in the Nile rat, an exotic African rodent, as a potential animal model for diet-induced type 2 diabetes mellitus and Metabolic Syndrome.MethodsCompiled data from several studies established the relationship between age, body weight gain (including abdominal adiposity), food and water consumption, and blood glucose levels as determinants of diabetes in male and female Nile rats. Glucose Tolerance Testing, insulin, HbA1c, blood pressure measurements and plasma lipids further characterized the diabetes in relation to criteria of the Metabolic Syndrome, while diet modification with high-fat, low-fiber or food restriction attempted to modulate the disease.ResultsThe Nile rat fed lab chow demonstrates signs of the Metabolic Syndrome that evolve into diet-induced non-insulin-dependent (type 2) diabetes mellitus characterized by hyperinsulinemia with rising blood glucose (insulin resistance), abdominal adiposity, and impaired glucose clearance that precedes increased food and water intake, as well as elevated HbA1c, marked elevation in plasma triglycerides and cholesterol, microalbuminuria, and hypertension. Males are more prone than females with rapid progression to diabetes depending on the challenge diet. In males diabetes segregated into early-onset and late-onset groups, the former related to more rapid growth and greater growth efficiency for the calories consumed. Interestingly, no correlation was found between blood glucose and body mass index (overall adiposity) in older male Nile rats in long term studies, whereas blood glucose and the perirenal fat pad, as well as liver and kidney weight, were positively related to early-onset diabetes. Rats weaned early (4-5 wks) and challenged with a high-fat Western-type diet developed diabetes faster, and body fat accumulation was more apparent, whereas food restriction curtailed it.ConclusionThe Nile rat fed typical rodent diets develops hyperinsulinemia that precedes hyperglycemia (insulin resistance) leading to diet-induced type 2 diabetes associated with hypertriglyceridemia, hypercholesterolemia, and hypertension. Dietary modulation affected growth rate (weight gain and central adiposity) to impact disease progression. This rodent model represents a novel system of gene-diet interactions affecting energy utilization that can provide insight into the prevention and treatment of the type 2 diabetes and Metabolic Syndrome.

Betaine in sub-acute and sub-chronic rat studies.

To evaluate the sub-acute and sub-chronic effect of large doses of betaine, this trimethyl glycine compound was fed to rats. Initial studies at BIBRA in the UK evaluated intakes of 0, 1, 2, and 5% betaine added to a maintenance chow designed for use in toxicology studies. Male and female Sprague-Dawley rats were followed for up to 90 days. No toxicity occurred, but at higher betaine intakes several serum chemistries were altered slightly, the MCV, MCH, and MCHC of red cells were reduced, and hepatocytes developed fatty droplets in direct proportion to betaine intake. Females were more affected than males. In a second study to assess reversibility in females, betaine effects were induced for 28 days, followed by a 28 day betaine-free period. All perturbations, except the reduced MCV and MCH, were reversed. As a follow up to BIBRA investigations, both 28 and 90 day feeding trials were conducted at Brandeis University using a rat chow with higher levels of energy, protein, and fat, with betaine added at 0, 0.5, 0.75, 1.0 and 5.0% of the diet. A similar broad range of clinical chemistries and physiological parameters were monitored, and hepatic lipid droplets were investigated in more detail. Liver lipid was actually reduced by betaine, and no significant adverse effects of clinical importance resulted from any dose. However, the MCV was again reduced at 5% betaine in the 28 day study. By 90 days all parameters were normal and comparable to controls. Based on these collective data, it was concluded that even at these high doses, betaine is nontoxic. Differences observed between the BIBRA and Brandeis studies were attributed to differences in the dietary formulations. Significant betainexdietxgrowth interactions were thought to reflect primary disparities in protein and energy concentrations, more than the addition of betaine per se.