Cholesterol metabolism and its implications in man.

Abstract

I-6 1 CHOLESTEROL METABOLISM AND ITS IMPLICATIONS IN MAN T. M. Sutherland*, P. Letchford* , Joan Arthur* and D.J. Farrell* I. INTRODUCTION It is generally, if not quite universally accepted that hypercholesterolemia and triglyceridemia are indicators and risk factors for coronary heart disease (see for example Shaper 1972, Vergroesen 1972). There is now a considerable literature beginning with the correlative observations of Walker and Arvidsson (1954) showing that the inclusion of fibrous materials in the diet may decrease plasma cholesterol levels. Trowell. (1972) and Story and Kritchkc'vsky ('1976) have extensively reviewed this literature. As might be expected not all sources and components of fibre are equally effective. Rolled oats (De Groat, Luyken & Pikaar, 1963; Luyken et al. 1962) and leguminous seeds (Anderson & Keys, 1965; Mather, Khan & Sharma 1968) have been shown to reduce blood cholesterol concentrations. Eastwood and co-workers (1973), Heatonand Pomare (1974) and Durrington, Wicks and Heaton (1975) obtained nil effects on cholesterol levels from the addition of wheat bran although Eastwood (1969) and Persson, Rabv, Fdnns-Beth and Jeason (1975) have reported significant falls. As recommendations to increase fibre intake as a preventative and therapeutic measure against low-fibre associated diseases are usually met by wheat bran used as a supplement and because this is such a variable product depending on grain sources and milling practice, it was felt worthwhile to examine the effects of increased wheat bran intake on blood cholesterollevels in humans under local Australian conditions. II. MATERIALS AND METHODS There were three experiments, all involved volunteers of both sexes mainly from the University of New England. A venous blood sample (5 ml) was withdrawn following an overnight fast to give the initial cholesterol concentration. Subsequent samples were taken at suitable .. intervals. Plasma was removed from whole blood following centrifugation and stored at -22OC for cholesterol analysis using an enzvmatic method . .L (Calbiochem) in a Centrificem System 400. (i) Experiment I . Twelve subjects ('5 males and 7 females), all but one aged less . than 25 years, were asked not to change their usual dietary habits but to take thrice daily, for 2 weeks, 3 bran snacks (Hygienic Food Supplies, .... West Ryde, N.S.W.) that contributed 2 g dietary fibre .. per day. Two subjects took instead 2 Bran Mini-Biscuits (Sanatarium Health Food Co., Sydney, N.S.W.). thrice daily; these also provided approximatelv the 2 g . of dietary fibre. Blood samples were taken at the &art of the experiment and again two weeks later. (ii) Experiment 2 Seventeen volunteers (12 females and 5 males) aged less than 40 * Department of Biochemistry & Nutrition, Universitv of New England, -. Armidale, N.S.W. 2351. 3.6 2 years were divided into two groups on the basis of an initial plasma Six subjects with a plasma cholesterol level cholesterol determination. below 6.2 m mol. z-1 were asked to take foods known to be naturally high in cholesterol. These were largely dairv. Iproducts, seafoods.and some . meats. The remainder (11) was asked to avoid these foods as far as possible. After three weeks on the dietary regimens all subjects took 12 g/d of wheat bran in equal amounts at each meal until the end of week 5. Both groups then reverted to their pre-experimental dietary habits. Venous blood samples were taken after 1, 3, 5 and 12 weeks. Diaries were kept by volunteers of foods' consumed each day. (iii) Experiment 3 _ Seventy three (51 males and 22 males) volunteers aged from 20 to 57 years were asked to continue their normal diet and eating pattern. With the exception of 6 control subjects, all were told to add 5-6 g of dietary fibre each day as wheat bran. This could be taken in several different food combinations, e.g. wholemeal flour, Ryvita biscuits, wholemeal bread, certain breakfast cereals, bran muffins, etc- A blood sample was taken at the start and after 3 weeks on the regimen. III. RESULTS (i) Experiment I + The mean pla?ma cholesterol at the start and 2 weeks later was 7.17-0.38 and.6.4100.39 m moles .z 'l respectively. The reduction was significant (P< 0.01). (ii) Experiment 2 . The effect of diet was important in changing plasma cholesterol concentrations. (Figure I). However, for the group with an initially high value, dietary change was apparently unable to reduce -.plasma cholesterol further after 1 week. The consumption of 12 g of wheat bran had an overriding influence in that plasma cholesterol of both croups declined significantly (P < 0.05) at the end of week 5. When told to resume their normal dietary habits, those that had a naturally low cholesterol remained at this level, while those with an elevated cholesterol showed a signficant increase at the end of week 12 compared with the mean value at week 5. (iii) Experiment 3 ' Data are from 65 volunteers who had an initial cholesterol concentration of 5.9 m moles .Z -1 ,. and 5.8 m moles .z -1 after 3 weeks during which time thev consumed an additional 5-6 g/d of dietary fibre. Those with an initialconcentration of 6 5 m moles .z -1 and above had a mean concentration of 7.2 m moles . z- I (N = 18); this was reduced significantly (P < 0.01) to 6.6 m moles .Z -1 at the end of 3 weeks. The six subjects used as controls showed no change (P < 0.05) during the 3-week period. IV. DISCUSSION Results of the first two experiments clearly demonstrate the ability of wheat fibre to reduce plasma cholesterol but only when the 163 Figure I. The cholesterol concentrations of Jii A and1X A with an initial low and high plasma cholesterol\ respectively. Wheat bran was introduced, thrice for 2 weeks when subjects had been on low 4 and cholesterol A diets for 3 weeks. At the end of and onwards diet was unspecified. subjects daily, high 5 weeks In xperiment 3, the unusually low average concentration of 5.9 m moles z- H coupled with the fact that many volunteers were already routinelv s taking considerable amounts of dietary fibre, as determined by a dietary survey of this group, indicates that the fibre addition was ineffective. However, bran was effective in lowering plasma cholesterol of those sub?' jects that had a slightly elevated level'at the start of Experiment 3. Fibre appears to exert its effects on cholesterol levels not by interfering with the absorption of dietary cholesterol but by increasing turnover (Balmer & Zilversmit, 1974). The most likely point for intervention is the enterohepatic circulation .of bile salts. In man some 30-40 mg of bile salts enter the gut daily of which only about 0.8 g w escape in the faeces and have to be replaced-by synthesis from cholesterol (Bergstrom & Danielsson, 1968). The simplest hypothesis for the fibre effect of increasing turnover and decreasing cholesterol level would be that binding of bile salts and acids to fibre prevents readsorption, necessitating increased conversion of cholesterolto bile acids. There is an appreciable amount of evidence supporting a mechanism of this kind. Animal experiments on fibre inclusion have shown increased faecal excretion of bile acids and sterols (Coleman and Baumann, 1957). Feeding of anion exchangers such as JW 135 (Bergen et al. 1959) . cholestyramine (Hashim & Van Itallie 1965) and colestipol (Parkinson et al. 1970) have all been shown to effectively lower plasma cholesterol concentrations. Eastwood and Boyd (1967) found appreciable quantities . of bile acids bound to nonabsorbable materials in the small intestine. Studies in vitro have confirmed the ability of dietary comlronents to bind bile salts (Birkner & Kern 1974, Kritchnevsky & Story 19i4 & 1975; Balmer & Zilversmit 1974). Story'and Kritchnevsky (1975) made comparative studies of bile acids and bile salts to bran, cellulose and lignin. Adsorption to cellulose was negligible but lignin was highly effective as an adsorbent. The adsorption to bran was higher than would be accounted for by its lignin content and the pattern of adsorption was different. The ratio of cholate to chenodeoxycholate adsorption was about 0,5 for bran but about 2 for lignin. Taurochenocholate and glycochenocholate were also much more strongly adsorbed to bran than the corresponding cholate or deoxycholate derivatives. On these properties one would expect the chenodeoxycholate pool to be the most depleted when bran is fed. In experiments in vivo Heaton (1972) found that the chenodeoxycholate pool fell on bran feeding. There are a number of possible explanations for this paradoxical finding. Deoxycholate is a secondary bile acid requiring,microbial activity for its formation. Pomare and Heaton (1974) have found the 7-a dehydroxylase activity to be depressed by bran feeding. This coupled with the diminished transit time caused by bran feeding would appreciably decrease the rate of formation of deoxycholate. Finally there is appreciable microbial digestion of structural carbohydrates of fibre (Williams 6 Olmsted 1936; Hummel, Shepherd & Macy 1943; Milton-Thompson St Lewis 1971; Southgate & Durnit! 1970; Southgate et al., 1976) so that binding at the site of readsorption of the bile salts may be determined largely by the lignin (deoxycholate preferring) residue. It would be interesting to have information on the relative effectiveness of cholate, deoxycholate and chenodeoxycholate and their *derivatives in the negative feed back control of B-hydroxy -@-methyl-glutary1CoA reductase, the rate-controlling enzyme of cholesterol biosynthesis and cholesterol 7a -hydroxylase, the rate limiting enzyme in bile acid formation* There are a number of unsatisfactory features in the simple bile salt binding hypothesis. Lignin which binds in vitro has been shown to be ineffective in vivo (Heaton & Barry 1971). The chemistry of the binding of bile salts by fibre is obscure. Oakenfull (1977) examined a range of purified fibres and fibre components and found binding only with lucerne and soya bean. The binding in these he attributed to the presence of saponins. Bile 'salts might bind ionically to fixed cations or hydronhobical. ly to non-polar materials. In the common bile salts there are at least two hydroxyls, the carbonyl and the -NH- group of the amide link as .. well as the carboxylate or sulphonate groups capable of hydrogen bonding. H bonds are weak and also directiona- so that multiple bondincr on a fixed carbohydrate matrix is unlikely. Eastwood andMitchell (1976) foun'd that although most natural fibre residues had appreciable cation exchange capacity they were unable to detect any anion exchange properties. This is perhaps not too surprising since no aminosugars have been detected in plant cell wall -- polysaccharides. The ability to sequester bile salts may be linked to the gelforming potential of the fibre. (Ershoff & Wells 1962). Pectin (Fisher et al. 1965; Anderson, Grande & Keys 1973); Jenkins et.al 1975) and Guar gum (Fahrenbach et al. 1965; Jenkins et al. 1975) have both been shown to be effective in lowering cholesterol levels in man. There is a wealth of evidence showing that pectin is particularly effective in preventing the hypercholesterolemia induced in animals by cholesterol feeding and may lead to increased faecal excretion of bile acids and sterols (Fisher et al. 1966; Lin et al. 1967). An observation indicating a more complex mechanism than sim-pie sequestration of cholesterol and bile salts is the reduction observed in the biosynthesis of liver cholesterol (Moday 1974). Recently guar gum has come into prominence as a possible means of controlling mild diabetes due to its effect in slowing glucose absorption. Insulin promotes hepatic lipogenesis and A. the synthesis triglyceride rich circulating lipoproteins (Farauahar et al. 1966) ; it is also one of the hormones involved in the induction of f+hydoxy B-methyl-glutaryl CoA reductase (EC1.1.1.34). A slower rate of glucose entry might be expected to lead to diminished insulin secretion. It is clear that a great deal more information is recruired before the relative importance of the various mechanisms proposed for the lowering of blood cholesterol,bile salt binding with increased drainage from the cholesterol pool, altered feed back control due changes in the proportions of the bile acids or altered hormonal control due to changes in rate and quantity of absorption of glucose and other metabolites, can be determined. There is a need for more detailed investigation in these areas but a1so.a need to widen the range of parameters measured in experiments on fibre for the true explanation for the effects may lie in an area as yet unexplored. The simple assay of plasma cholesterol in experiments of this type needs to be supplemented -.. by determination of high and low density lipoproteins and by glucose ._ and insulin tolerance tests. V. ACKNOWLEDGEMENTS The help and cooperation of Dr. Arthur Beresford of the Armidale and District Hospital, Mr. John Roberts of the Department of Physiology, the University of New England and MS Rosalind Angus, Mrs. Barbara Ward and Miss Shiralee Ford of the Department of Biochemistry and Nutrition, University of New England is gratefullv acknowledged. We also wish to thank all the volunteers who took part in these experiments. VI. REFERENCES ANDERSON, J-T., GRANDE, F. & KEYS, A. (1973). J. Am. Diet. Assoc. -- : 62 133 BALMER, J. & ZILVERSMIT, D.B. (1974). J. Nutr. --- : 1319 104 BERGEN, S-S., VAN ITALLIE, T-B., TENNENT, D.M. & SEBRELL, W.H. (1959) Proc.- Soc. exp. Biol. Med. --- : 676 102 BERGSTROM, S. & DANIELSSON, H. (1968). In 'Handbook of Physioloqv' CF.' Code, editor: Vol. 5, Section 6, P. 2391 (Williams & Wilkins, Baltimore). BIRKNER, H.J. & KERN, F. (1974). Gastroenterology 67 : 237 COLEMAN, D.L. & BAUMANN, CA. (1957). Arch. 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