Biochemistry and Nutrition & Food Science, University of. Surrey, Guildford, U.K.; and 2Department of Molecular. Sciences, Division of Biology, Aston University,.
Bioscience Reports 5, 701-705 (1985) Printed in Great Britain
701
E I I e c t s of I a t t y acid chain length and saturation on g a s t r i c inhibitory polypeptide release in o b e s e hyperglycaemic
(oh/oh) mice
Piotr KWASOWSKI 1, Peter R. FLATT I, Clifford 3. BAILEY2 and Vincent MARKS 1 IDepartment of Biochemistry, Divisions of Clinical Biochemistry and Nutrition & Food Science, University of Surrey, Guildford, U.K.; and 2Department of Molecular Sciences, Division of Biology, Aston University, Birmingham, U.K. (Received 12 August 1985)
Plasma gastric inhibitory polypeptide (GIP) responses to e q u i m o l a r i n t r a g a s t r i c a l l y administered emulsions of f a t t y acids (2.62 mmol/7.5 ml/kg) were examined in 18 h fasted obese hyperglycaemic (ob/ob) mice. Propionic acid (C3:0), a saturated short-chain f a t t y acid, and capric acid (C10:0), a saturated medium chain f a t t y acid, did not s i g n i l i c a n t l y s t i m u l a t e GIP release. H o w e v e r , the saturated long-chain f a t t y acid stearic acid (C18"0), and especially the unsaturated long-chain f a t t y acids oleic (C18:1), linoleic (C18:2) and linolenic (C18:3) acids produced a marked GIP response. The results show that chain length and to a lesser extent the degree of saturation are important determinants of f a t t y a c i d - s t i m u l a t e d GIP release. The GIP-release action of long-chain, but not short-chain, fatty acids m a y be r e l a t e d to differences in their intracellular handling. Gastric inhibitory polypeptide (GIP) is secreted by the K-cells of the intestinal mucosa during the alimentary processing of food (Brown, 1982; Creutzfeldt, 1982; Marks & Morgan, 1983). In addition to its possible role in the control of gastric acid secretion (Brown, 1982), GIP e x e r t s a g l u c o s e d e p e n d e n t i n s u l i n - r e l e a s i n g e f f e c t on the pancreatic islet B-cells, and constitutes a physiological component of the enteroinsular axis (Brown, 1982; Creutzfeldt, 1982; Marks & Morgan, 1983). Studies in many species, including man, have shown that GIP secretion is stimulated by carbohydrates, proteins and fats (Brown, 1982). Dietary fats provide an especially potent stimulus for GIP release (Brown et al., 1975; Falko et al., 1973; O'Dorisio et al., 1976), but the mechanism is not established. Thus, in contrast to the prominent effect of triglycerides, variable GIP responses have been observed to f a t t y acids and mono- and di-glycerides (Ross & Shaffer, 1981; Williams et al., 19gl). Recent studies have identified the genetically
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KWASOWSKI ET AL.
obese hyperglycaemic (ob/ob) mouse as a valuable model to investigate GIP release. This model exhibits intestinal hypertrophy (Flatt et al., 1983) and enteroendocrine hyperplasia (Polak et al., 1975), with enhanced basal and nutrient-stimulated GIP concentrations (Flatt et al., 1983, 198#). The present study investigates the effects of fatty acid chain length and saturation on GIP release in ob/ob mice. Materials
and Methods
Groups of Aston obese hyperglycaemic (ob/ob) mice from the colony maintained at Surrey University were used at 12-18 weeks of age. The origin and characteristics of Aston ob/ob mice have been described elsewhere (Flat & Bailey, 1981; Bailey et al., 1982). Mice were housed in an air-conditioned room at 22 + 2~ with a lighting schedule of 12 h light (0700-1900 h) and 12 h darkness. A standard pellet diet (Spratts Laboratory Diet 1, Lillico Ltd., Reigate, U.K.) and tap water were supplied ad libitum. This diet comprised 3.5% fat, 21.5% protein and #g% carbohydrate (digestible energy 1#.2 MJ/kg) with added fibre, vitamins and minerals. F a t t y acids (Sigma Chemical Company Ltd., Poole, U.K.) were administered intragastrically as an aqueous emulsion (2.62 mmol/7.5 ml/kg) to conscious 18 h fasted mice. The dose corresponded with the overall composition of the fat emulsion (Intralipid, Kabi Vitrum L t d . , Ealing, U.K.) previously tested in ob/ob mice (Flair et al., 1984). The f a t t y acids (with numbers of carbon atoms: unsaturated carbon-carbon bonds, and dose in mg/kg) were: propionic acid (C3:0, 19# mg/kg), capric acid (Cl0:0, #51 mg/kg), stearic acid (Clg:0, 7#6 mg/kg), oleic acid (C18:I, 741 mg/kg), linoleic acid (cIg:2, 735 mg/kg) and linolenic acid (cIg:3, 730 mg/kg). Blood samples (60 lal) were taken from the tail-tip of conscious mice immediately before and at 30, 60, 90 and 120 min after fatty acid administration. Plasma GIP was measured by radioimmunoassay (Morgan et al., 197g) using donkey anti-rabbit g a m m a globulin antiserum (Guildhay Antisera, University of Surrey, Guildford, U.K.) for separation of free from bound antigen. Porcine GIP (Professor 3.C. Brown, University of British Columbia, Canada) was used to prepare 1251-GIP and as standard. Immunoadsorbed GIP-free human plasma was added to the tubes of the standard curve used to minimize non-specific interference, and parallelism was demonstrated between the standard curve and serially diluted ob/ob mouse plasma. The GIP antiserum (RIC 34/11I; Guildhay Antisera) recognizes both 5000 and g000 molecular forms of GIP. It exhibits negligible cross reactivity (less than i % ) with other e n t e r o - p a n c r e a t i c hormones, including cholecystokinin, glucagon, gut g l u c a g o n - l i k e immunoreactivity (enteroglucagon), insulin, pancreatic p o l y p e p t i d e , proinsulin C-peptide, secretin and vasoactive intestinal polypeptide. Groups of data were compared by analysis of variance (ANOVA). Differences were considered to be significant if P < 0.05, and these were confirmed using Student's paired and unpaired t-tests as appropriate. Results
Mean basal plasma GIP concentrations in the experimental groups of 18 h f a s t e d ob/ob mice were 526-614 pg/ml (Fig. 1). Oral administration of propionic acid, a saturated short-chain (C3:0) f a t t y
FATTY
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AND
GASTRIC INHIBITORY PEPTIDE
Propionic acid (C3:0) a691 + 350
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Fig. i. Effects of fatty acid chain length and s a t u r a t i o n on plasma GIP concentrations in 18 h fasted ob/ob mice. Fatty acids were administered intragastrically as an aqueous emulsion at a dose of 2.62 mmol/7.5 ml/kg. Numbers of carbon atoms and unsaturated carbon-carbon bonds are s h o w n in parentheses. A values represent the increment above basal for the GIP response expressed as pg.ml-l.h, calculated as described in the Results. Values are presented as means • SEM of 5-6 mice.
acid, and capric acid, a saturated medium-chain (C10:0) fatty acid, did not significantly alter plasma GIP concentrations. However, the saturated and unsaturated long-chain fatty acids stearic acid (Clg:0), oleic acid ( C l g : l ) , linoJeic acid (CIg:2) and linolenic acid (CIg:3) produced a 3-#-fold elevation (P < 0.0l) of plasma GIP concentrations between 60 and 120 rain. Mean increments above basal areas for the GIP responses were calculated as the sum of values at 30-120 min minus # times the basal value, and divided by 2 to give final expression as pg.ml-l-h. The increments above basal were: propionic acid 691 -+ 350 (mean • SEM), capric acid 216 • 2gl, stearic acid 13#g • #36, oleic acid 2399 • 392, linoleic acid 2550 +- 691, and linolenic acid 150g • 665. The GIP response to oleic acid was significantly greater (P < 0.05-0.01) than the responses to prop•177 capric and stearic acids, but not significantly greater than to linoleic and linolenic acids. Discussion
Fat-stimulated GIP release is dependent on of triglycerides and absorption of the digestion GIP release is compromised in conditions of such as cystic fibrosis and chronic pancreatitis Ebert & Creutzfeldt, 19g0), and in the latter pancreatic enzymes partially restore the GIP
intraluminaI hydrolysis products. Accordingly, impaired fat digestion (Ross & Shaffer, 19gl; condition extracts of responses to oral fat
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KWASOWSKI ET AL.
(Ebert & Creutzfeldt, lgg0). In contrast, retardation of fat absorption with cholestyramine impairs GIP release (Ebert & Creutzfeldt, t980). Previous studies on the relative importance of triglyceride d i g e s t i o n p r o d u c t s in f a t - s t i m u l a t e d GIP r e l e a s e have produced equivocal results. Thus, medium-chain triglycerides, monoglycerides, glycerol and both saturated and unsaturated long-chain f a t t y acids such as palmitic, oleic and linoleic acids have all been reported to lack stimulatory e f f e c t s on GIP release (Ross & Shaffer, 19gl; Williams et al., 1991). In the present study, oral administration of the short-chain and medium-chain fatty acids, propionic (C3:0) and capric (CI0:0) acids, failed to produce a significant GIP response. However, equimolar a d m i n i s t r a t i o n of a long-chain saturated f a t t y acid (stearic acid, CIg:0) and especially the long-chain unsaturated f a t t y acids (oleic CI8:I, linoleic Clg:2, and linolenic C18.3 acids) produced a marked GIP response. Short-chain and medium-chain f a t t y acids are absorbed m o r e r a p i d l y t h a n l o n g - c h a i n f a t t y acids, and the more potent GIP-releasing unsaturated long-chain f a t t y acids are absorbed faster than their saturated counterparts (Cl&ment, 1980; Thomson & Dietschy, 1981). This indicates that GIP release is not related only to the rate of cellular uptake of f a t t y acids, an event which does not demand energy expenditure (Cl6ment, 1980; Thomson & Dietschy, 198I). It is t h e r e f o r e likely t h a t the stimulus for fat-induced GIP release is generated during the intracellular handling and metabolism of f a t t y acids. Short- and medium-chain f a t t y acids are transferred across the intestinal epithelium without esterification. However, long-chain f a t t y acids are conveyed to the smooth endoplasmic reticulum for esterification prior to incorporation into chylomicrons and exocytosis into the intercellular compartment (Clement, 1980; Thomson & Dietschy, 19gl). The GIP-releasing action of f a t t y acids may be coupled therefore to the extent of esterification, an energy-consuming metabolic process c o n f i n e d to l o n g - c h a i n f a t t y acids ( C l e m e n t , 1980; Thomson & Dietschy, 19gl). Previous studies on the mechanism of carbohydrates t i m u l a t e d GIP r e l e a s e have shown that only actively transported sugars, such as glucose and galactose, stimulate GIP secretion (Sykes et al., 19g0). Thus there may be a common link between metabolic and secretory events responsible for nutrient-regulation of GIP release from the intestinal K-cells.
References Bailey CJ, Flatt PR & Atkins TW (1982) Influence of genetic background and age on the expression of the obese hyperglycaemic syndrome in Aston ob/ob mice. Int. J. Obes. 6, 11-21. Brown JC (1982) Gastric Inhibitory Polypeptide. Monographs on Endocrinology, vol. 24, pp 1-88, Springer-Verlag, Berlin. Brown JC, Dryburgh JR, Ross SE & Dupre J (1975) Identification and actions of gastric inhibitory polypeptide. Recent Prog. Horm. Res. 31, 487-532. Cl6ment J (1980) Intestinal absorption of triglycerols. Reprod. Nutr. Develop. 20, 1285-1307.
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Creutzfeldt W (1982) Gastrointestinal peptides - role in pathophysiology and disease. Scand. J. Gastroenterol. 17, suppl. 77, 7-20. Ebert R & Creutzfeldt W (1980) Decreased GIP secretion through impairment of absorption. In: Frontiers in Hormone Research, vol 7, pp 192-201 (Creutzfeldt W, ed), Karger, Basel. Falko JM, Crockett SE, Cataland S & Mazzaferri EL (1976) Gastric inhibitory polypeptide (GIP). Intestinal distribution and stimulation by amino acids and medium chain triglycerides. J. Clin. Endocrinol. Metab. 41, 260-265. Flatt PR & Bailey CJ (1981) Abnormal plasma glucose and insulin responses in heterozygous lean (ob/+) mice. Diabetologia 20, 573-577. Flatt PR, Bailey CJ, Kwasowski P, Swanston-Flatt SK & Marks V (1983) Abnormalities of GIP in spontaneous syndromes of diabetes and obesity in mice. Diabetes 32, 433-435. Flatt PR, Bailey CJ, Kwasowski P, Page T & Marks V (1984) Plasma immunoreactive gastric inhibitory polypeptide in obese hyperglycaemic (ob/ob) mice. J. Endocrinol. IO1, 249-256. Marks V & Morgan LM (1984) The enteroinsular axis. In: Recent Advances in Diabetes, vol I, pp 55-71 (Nattrass M & Santiago JV, eds), Churchill Livingstone, Edinburgh. Morgan LM, Morris BA & Marks V (1978) Radioimmunoassay of gastric inhibitory polypeptide. Ann. Clin. Biochem. 15, 172-177. O'Dorisio TM, Cataland S, Stevenson M & Mazzaferri EL (1976) Gastric inhibitory polypeptide (GIP). Intestinal distribution and stimulation by amino acids and medium chain triglycerides. Am. J. Dig. Dis. 21, 761-765. Polak JM, Pearce AGE, Grimelius L & Marks V (1975) Gastrointestinal apudosis in obese hyperglycaemic mice. Virchows Arch. (Cell. Pathol.) 19, 135-150. Ross SA & Shaffer EA (1981) The importance of triglyceride hydrolysis for the release of gastric inhibitory polypeptide. Gastroenterology 80, 108-111. Sykes S, Morgan LM, English J & Marks V (1980) Evidence for preferential secretion of gastric inhibitory polypeptide in the rat by actively transported carbohydrates and their analogues. J. Endocrinol. 85, 201-207. Thomson ABR & Dietschy JM (1981) Intestinal lipid absorption: major extracellular and intracellular events. In Physiology of the Gastrointestinal Tract, vol 2, pp 1147-1220 (Johnson LR, ed), Raven Press, New York. Williams RJ, May, JM & Biebroeck JB (1981) Determinants of gastric inhibitory polypeptide and insulin secretion. Metabolism 30, 36-40.
