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Dietary and nutritional aspects of fatty acid binding proteins
Chemisto' and Physics o f Lipids. 38 (1985) 131 - 135 Elsevier Scientific Publishers Ireland Ltd.
131
DIETARY AND NUTRITIONAL ASPECTS OF F A T T Y ACID BINDING PROTEINS
RIAZ-UL-HAQ and EARL SHRAGO Departments o f Nutritional Sciences and Medicine University of Wisconsin, Madison, W153706 (U.S.A.) Received April 23rd, 1985 Information on cytosolic fatty acid binding proteins (FABP) related to dietary and pharmacological manipulations is discussed in terms of FABP function. FABP present in liver, heart, intestinal mucosa and omental fat responds to different diets. A parallel change occurs in tissue levels of FABP and metabolism of fatty acids. It seems FABP might play a role in lipid metabolism by interacting with membrane bound enzymes. The available data also support the argument in favor of FABP involvement in intracellular transport, compartmentalization and channeling of fatty acids. Keywords: fatty acid binding protein; nutrition; lipid metabolism; diabetes; clofibrate. During the last decade fatty acid binding protein (FABP) has been identified in the cytosol of various tissues o f the rat. More recently it has also been reported in human liver and adipose tissue [1,2]. The protein has been cited by several laboratories to play a role in the intracellular metabolism of long chain fatty acids such as cellular uptake [3], esterification [4,5] and oxidation [6,7]. Since many o f the enzymatic reactions of fatty acid metabolism respond to dietary manipulation, it seems logical to correlate these variations in enzymatic activity with the tissue levels o f FABP. It is important to know concentration of FABP under different nutritional states in order to understand its overall physiological function. The purpose o f this communication is to summarize and conceptualize the FABP data related to diet and nutritional status of the animal. Intestinal FABP levels respond to dietary changes [8]. Rats kept on high fat diet for three weeks had significantly higher concentration of FABP in the middle and distal intestine as compared to control (Table I). However, the FABP concentration in the proximal segment was not affected by the high fat diet. It was proposed that FABP in the middle and distal intestine is present at basal levels, and as these segments become more involved in the absorption process, the FABP concentration increased in response to the high fat diet. Rat intestine contains two immunologically distinct proteins [9] and which one precisely changes with diet is not known at the present time. Chickens fed a diet containing 15% fat for 8 weeks show changes in intestinal FABP [10]. Unlike the rat, in broiler type birds the concentration of FABP increased in all three intestinal segments. An attempt has also been made to explore 0009-3084/85./$03.30 © 1985 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland
132 IABLE 1 fissue
Proximal intestine Middle imestine Distal intestir~e Liver t.iver Liver Intestinal mucosa Heart Omental fat Liver
Dietary treatment
High fat High fat High fat High fat High carbohydrate High fat High tat High fat High fat 5% Linoleate
FABP conc./binding activity values are mean ± S.E. Control
Treatment
14.9 15.9 7.3 0.29 0.29 0.071 0.328 0.685 3.28 37.1
14.4 21.0 10.5 0.43 0.35 0.027 0.088 0.418 1.037 27.3
_+1.7 +0.8 __.0.9 ±0.01 -+0.0l _+0.027 _+0.121 _+0.27 ±3.22 ± 8.6
± 1.6 +_2 _+0.9 +-0.02 +_0,02 +_0.006 +_0.038 +_0.157 +_0.76 ± 1.8
Ref.
8a
8a 8a 12b 12 b t3 c 13 c 13 c 13 c 18d
aug FABP/mg soluble protein. bnmol palmitoyl CoA bound to FABP fraction/mg protein. CProtein concentration (mg/ml cytosol) required for immunological detection of FABP. dpercentage of added palmitoyl CoA bound to FABP fraction. the differences in fat utilization in relation to intestinal FABP in birds obtained 1¥om different genetic sources. New Hampshire birds utilize fat more efficiently than broiler type birds in the first few weeks o f their life. It was speculated that FABP might be a limiting factor for the breed differences, as the broiler chick has less FABP at hatching time [1 1 ]. F a t t y acid binding protein concentration increased in both breeds kept on 12% corn oil diet, but the distribution o f FABP between breeds was different in the proximal and distal intestine. Addition o f sodium taurocholate to basal diet (3% fat) of birds increased the concentration o f FABP. This observation warrants further investigation in terms of FABP synthesis as a function o f bile salt. It has been shown that [~4C] palmitoyl CoA binding to liver FABP is significantly higher in certain experimental as compared to control groups [12]. Figure 1 shows that with the same anaount of cytosolic protein, palmitoyl CoA bound to FABP is higher in the high fat group (45%) in comparison to the control group. The change in binding activity can be due either to the increase in the quantity o f FABP or an increase in binding affinity of protein for palmitoyl CoA. It also cannot be excluded that both binding affinity and quantity of protein are responsible for the changes. However, other studies using quantitative radioimmunodiffusion techniques have shown that it is FABP concentration which is increased in response to a high fat diet [13]. FABP concentration was also increased in other tissues such as intestine, heart and omental fat besides liver (Table 1). F a t t y acid utilization which increases in different tissues in response to the diet is tbllowed by an increase in FABP concentration. The elevated levels of FABP oh-
133 H High fat o-o Control a-~ Absorbonce, 280nm
~opoo
2.5
80OO
2.0
6000
1.5
4000
1.0
2000
0.5
I0
20
30 40 50 Tube Number
60
Fig. 1. Sephadex G-75 chromatography of rat liver cytosol containing 52 mg protein mixed with [t4ClPalmitoyl CoA. Eluting buffer, 0.01 M sodium phosphate (pH 7.4) at a flow rate of 30 ml/h. served by different laboratories in the high fat diet group suggests the participation of this protein in the transport and oxidation of fatty acids. There are reports that FABP could be involved in the oxidation of fatty acids by heart mitochondria and liver peroxisomes [6,7]. The dietary induction of FABP is consistent with the concept that it might modulate the compartmentalization of fatty acids, thereby channeling them toward mitochondria particularly in the case of a high fat diet. A parallel increase in the concentration of FABP in both liver and heart of rats on a high fat diet (Table 1) suggests a genetic similarity of these proteins. On the other hand, a recent report concluded that FABP from rat liver and heart are different proteins [14}. However, this finding is not compatible with the observation that liver FABP mRNA is detectable in heart [15]. The presence of liver FABP mRNA in different organs warrants the need for further studies before a firm conclusion can be drawn on the structural and immunological identity of FABP in different tissues. It may be of interest that rat liver FABP concentration was also noted to be increased in response to a high carbohydrate diet (Table 1). Although the uptake of fatty acids might not be increased by a high carbohydrate diet, lipogenesis would be greater, and it is possible that liver FABP plays a role in the intracellular transport and storage of newly synthesized fatty acids. This observation is consistent with a study which showed that 5% linoleate in the diet decreased the amount of liver FABP along with reduction in the activities of key lipogenic enzymes [ 16]. A number of intriguing parallels exists between dietary changes and the physio-
134 logical aspects of fatty acid metabolism. As an example during lasting fatty acid synthesis decreases, while it increases in animals fed a high carbohydrate diet. Stein et al. [17] studied the effect o f 48-h fasting on the concentration of liver FABP. Total liver FABP concentration decreased about 50%, however, it should be noted that there was also a 36% decrease in liver weight of fasting animals. In another study in rats fasted for 48 h and then refed a fat free diet for the following 2 days results were more conclusive [18]. Specific precipitation of [u-t4C]palmitoyl CoA binding protein with anti-Z immunoglobulin G showed 89% more FABP bound palmitoyl CoA in refed than fasted rats (Table 1I). These responses in FABP concentration to starvation and refeeding further support the concept of involvement of FABP in fatty acid metabolism. Starvation is similar to uncontrolled diabetes in ternrs of fatty acid esterification and oxidation. In the diabetic liver [l-14C]oleic acid binding to FABP was found to be 37% less than that of the control liver [19]. The decreased binding of ligand was reversed by administration o f insulin to diabetic rats (Table 11). The mechanism of this change or potential role of FABP in diabetes is not clear, bul it is likely similar to that of fasting. It would be of interest to correlate the protein bound fatty acids in diabetic liver with the immunoprecipitable FABP. Besides dietary variations, pharmacological manipulations by hypolipidemic drugs seems to effect the hepatic content of FABP. Incorporation of Clofibrate (0.25%) into normal chow diet increased liver FABP by 3-fold [20], and the increase has been correlated with a higher uptake of fatty acids from perfusate [21]. Experiments in vivo showed no significant increase in the incorporation of [14C] oleic acid into triacylglycerol in spite of high FABP content in clofibtic acid fed rats [22]. On the other hand, in vitro studies reported a concomitanl increase in FABP content and biosynthesis of triacylglycerol in rats ted a diet containing cholestyramine [23]. The discrepancy between in vilro and whole aninral results may be due to differences in two drugs. This leads to the speculation that FABP may not always be directly involved in the synthesis of triacylglyccrol
TABLE 11 Tissue
Liver Liver Liver Liver Liver Liver
Dietary treatment
Starved 48 h Refed fat free diet 48 h Diabetic Diabetic + insulin Starved 48 h Refed 20 h
FABP binding activity values are mean _+S.E. Control
l'reatlnenl
27.5 ± 4.3 27.5 +4.3 27.5 ±4.3 27.5 _+4.3
51.9 73.5 17.2 ± 4.5 25.0 ± 0.9 17.8 _+1.5 19.2 + 1.7
anmol bound palmitoyl CoA precipitated from FABP using antibody. bpereentage of added oleic acid bound to FABP fraction.
Ref.
18a 18 a 19 b 19 b 19b 19 b
135 under different pharmacological conditions, but supports the notion of FABP participation in the compartmentalization and channeling o f fatty acids toward initochondria or smooth endoplasmic reticulum under various nutritional, hormonal and pharmacological conditions. The precise mechanism or factors responsible for the dietary induction of FABP ave unclear at the present time. It is not known which fatty acids or their CoA derivative induce the synthesis of FABP. Knowledge of FABP synthesis in relation to diet can be expanded utilizing the cloned cDNAs as a potential tool. FABP synthesis can be controlled at the gene transcription or mRNA translation level. Control of FABP synthesis by transcriptional or translational mechanism under various nutritional conditions still awaits future research in this direction.
Acknowledgements Work in author's laboratory was supported by U.S.P.H.S. N1H SCOR Grant 1 P50 HL 27358 and R01 GM-14033. References l K. Kamisaka, M. Hirano and M. Tsuru, Experientia, 34 (1978) 1265-1266. 2 R.U. Haq, L. Christodoulides, B. Ketterer and E. Shrago, Biochim. Biophys. Acta, 713 (1982) 193--198. 3 C.A. Goresky, D.S. Daly, S. Mishkin and I.M. Arias, Am. J. Physiol., 254 (1978) 542553. 4 M.Y.C. Wu-Rideout, C. Elson and E. Shrago, Biochem. Biophys. Res. Commun., 71 (1976) 809-816. 5 D.A. Bumett, N. Lysenko, J.A. Manning and R.K. Ockner, Gastroenterology, 77 (1979) 241-249. 6 N. Fournier, M. Geoffroy and J. Deshusses, Biochim. Biophys. Acta, 533 (1978) 457-464. 7 E.L. Appelkvist and G. Dallner, Biochim. Biophys. Acta, 617 (1980) 156-160. 8 R.K. Ockner and J.A. Manning, J. Clin. Invest., 54 (1974) 336 338. 9 D.H. Alpers, A.W. Strauss, R.K. Ockner, N.M. Bass and J.l. Gordon, Proc. Natl. Acad. Sci. U.S.A., 81 (1984) 313-317. 10 J.B.D. Katongole and B.E. March, Poultry Sci., 58 (1979) 372-375. 11 J.B.D. Katongole and B.E. March, Poultry Sci., 59 (1980) 819 827. 12 R.U. Haq and E. Shrago, Nutr. Res., 3 (1982) 329-333. 13 B. Riistow, R. Risse and D. Kunze, Acta Biol. Med. Germ., 41 (1982) 439-445. 14 B. Said and H. Schulz, J. Biol. Chem., 259 (1984) 1155-1159. 15 J.l. Gordon, N. Elshourbagy, J.B. Lowe, W.S. Liao, D.H. Alpers and J.M. Taylor, J. Biol. Chem., 260 (1985) 1995-1998. 16 G.R. Herzberg and M. Rogerson, Nutr. Res., 1 (1981) 601-607. 17 L.B. Stein, S. Mishkin, G. Fleischner, Z. Gatmaitan and I.M. Arias, Am. J. Physiol., 31 (1976) 1371-1376. 18 N. lritani, E. Fukuda and K. lnoguchi, J. Nutr. Sci. Vitaminol., 26 (1980) 271-277. 19 R. Brandes and R. Arad, Biochim. Biophys. Acta, 750 (1983) 334-339. 20 G. Fleischner, D.K.F. Meijer, W.G. Levine, Z. Gatmaitan, R. Gluck and I,M. Arias, Biochem. Biophys. Res. Commun., 67 (1975) 1401-1407. 21 G. Renaud, A. Foliot, R. Infante, Biochem. Biophys. Res. Commun., 80 (1978) 327-334. 22 Y. Kawashima, S. Nakagawa and H. Kozuka, J. Pharmacobiodyn., 5 (1982) 771-779. 23 H.J.M. Kempen, J.F.C. Glatz, J. DeLange and J.H. Veerkamp, Biochem. J., 216 (1983) 511-514.
131
DIETARY AND NUTRITIONAL ASPECTS OF F A T T Y ACID BINDING PROTEINS
RIAZ-UL-HAQ and EARL SHRAGO Departments o f Nutritional Sciences and Medicine University of Wisconsin, Madison, W153706 (U.S.A.) Received April 23rd, 1985 Information on cytosolic fatty acid binding proteins (FABP) related to dietary and pharmacological manipulations is discussed in terms of FABP function. FABP present in liver, heart, intestinal mucosa and omental fat responds to different diets. A parallel change occurs in tissue levels of FABP and metabolism of fatty acids. It seems FABP might play a role in lipid metabolism by interacting with membrane bound enzymes. The available data also support the argument in favor of FABP involvement in intracellular transport, compartmentalization and channeling of fatty acids. Keywords: fatty acid binding protein; nutrition; lipid metabolism; diabetes; clofibrate. During the last decade fatty acid binding protein (FABP) has been identified in the cytosol of various tissues o f the rat. More recently it has also been reported in human liver and adipose tissue [1,2]. The protein has been cited by several laboratories to play a role in the intracellular metabolism of long chain fatty acids such as cellular uptake [3], esterification [4,5] and oxidation [6,7]. Since many o f the enzymatic reactions of fatty acid metabolism respond to dietary manipulation, it seems logical to correlate these variations in enzymatic activity with the tissue levels o f FABP. It is important to know concentration of FABP under different nutritional states in order to understand its overall physiological function. The purpose o f this communication is to summarize and conceptualize the FABP data related to diet and nutritional status of the animal. Intestinal FABP levels respond to dietary changes [8]. Rats kept on high fat diet for three weeks had significantly higher concentration of FABP in the middle and distal intestine as compared to control (Table I). However, the FABP concentration in the proximal segment was not affected by the high fat diet. It was proposed that FABP in the middle and distal intestine is present at basal levels, and as these segments become more involved in the absorption process, the FABP concentration increased in response to the high fat diet. Rat intestine contains two immunologically distinct proteins [9] and which one precisely changes with diet is not known at the present time. Chickens fed a diet containing 15% fat for 8 weeks show changes in intestinal FABP [10]. Unlike the rat, in broiler type birds the concentration of FABP increased in all three intestinal segments. An attempt has also been made to explore 0009-3084/85./$03.30 © 1985 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland
132 IABLE 1 fissue
Proximal intestine Middle imestine Distal intestir~e Liver t.iver Liver Intestinal mucosa Heart Omental fat Liver
Dietary treatment
High fat High fat High fat High fat High carbohydrate High fat High tat High fat High fat 5% Linoleate
FABP conc./binding activity values are mean ± S.E. Control
Treatment
14.9 15.9 7.3 0.29 0.29 0.071 0.328 0.685 3.28 37.1
14.4 21.0 10.5 0.43 0.35 0.027 0.088 0.418 1.037 27.3
_+1.7 +0.8 __.0.9 ±0.01 -+0.0l _+0.027 _+0.121 _+0.27 ±3.22 ± 8.6
± 1.6 +_2 _+0.9 +-0.02 +_0,02 +_0.006 +_0.038 +_0.157 +_0.76 ± 1.8
Ref.
8a
8a 8a 12b 12 b t3 c 13 c 13 c 13 c 18d
aug FABP/mg soluble protein. bnmol palmitoyl CoA bound to FABP fraction/mg protein. CProtein concentration (mg/ml cytosol) required for immunological detection of FABP. dpercentage of added palmitoyl CoA bound to FABP fraction. the differences in fat utilization in relation to intestinal FABP in birds obtained 1¥om different genetic sources. New Hampshire birds utilize fat more efficiently than broiler type birds in the first few weeks o f their life. It was speculated that FABP might be a limiting factor for the breed differences, as the broiler chick has less FABP at hatching time [1 1 ]. F a t t y acid binding protein concentration increased in both breeds kept on 12% corn oil diet, but the distribution o f FABP between breeds was different in the proximal and distal intestine. Addition o f sodium taurocholate to basal diet (3% fat) of birds increased the concentration o f FABP. This observation warrants further investigation in terms of FABP synthesis as a function o f bile salt. It has been shown that [~4C] palmitoyl CoA binding to liver FABP is significantly higher in certain experimental as compared to control groups [12]. Figure 1 shows that with the same anaount of cytosolic protein, palmitoyl CoA bound to FABP is higher in the high fat group (45%) in comparison to the control group. The change in binding activity can be due either to the increase in the quantity o f FABP or an increase in binding affinity of protein for palmitoyl CoA. It also cannot be excluded that both binding affinity and quantity of protein are responsible for the changes. However, other studies using quantitative radioimmunodiffusion techniques have shown that it is FABP concentration which is increased in response to a high fat diet [13]. FABP concentration was also increased in other tissues such as intestine, heart and omental fat besides liver (Table 1). F a t t y acid utilization which increases in different tissues in response to the diet is tbllowed by an increase in FABP concentration. The elevated levels of FABP oh-
133 H High fat o-o Control a-~ Absorbonce, 280nm
~opoo
2.5
80OO
2.0
6000
1.5
4000
1.0
2000
0.5
I0
20
30 40 50 Tube Number
60
Fig. 1. Sephadex G-75 chromatography of rat liver cytosol containing 52 mg protein mixed with [t4ClPalmitoyl CoA. Eluting buffer, 0.01 M sodium phosphate (pH 7.4) at a flow rate of 30 ml/h. served by different laboratories in the high fat diet group suggests the participation of this protein in the transport and oxidation of fatty acids. There are reports that FABP could be involved in the oxidation of fatty acids by heart mitochondria and liver peroxisomes [6,7]. The dietary induction of FABP is consistent with the concept that it might modulate the compartmentalization of fatty acids, thereby channeling them toward mitochondria particularly in the case of a high fat diet. A parallel increase in the concentration of FABP in both liver and heart of rats on a high fat diet (Table 1) suggests a genetic similarity of these proteins. On the other hand, a recent report concluded that FABP from rat liver and heart are different proteins [14}. However, this finding is not compatible with the observation that liver FABP mRNA is detectable in heart [15]. The presence of liver FABP mRNA in different organs warrants the need for further studies before a firm conclusion can be drawn on the structural and immunological identity of FABP in different tissues. It may be of interest that rat liver FABP concentration was also noted to be increased in response to a high carbohydrate diet (Table 1). Although the uptake of fatty acids might not be increased by a high carbohydrate diet, lipogenesis would be greater, and it is possible that liver FABP plays a role in the intracellular transport and storage of newly synthesized fatty acids. This observation is consistent with a study which showed that 5% linoleate in the diet decreased the amount of liver FABP along with reduction in the activities of key lipogenic enzymes [ 16]. A number of intriguing parallels exists between dietary changes and the physio-
134 logical aspects of fatty acid metabolism. As an example during lasting fatty acid synthesis decreases, while it increases in animals fed a high carbohydrate diet. Stein et al. [17] studied the effect o f 48-h fasting on the concentration of liver FABP. Total liver FABP concentration decreased about 50%, however, it should be noted that there was also a 36% decrease in liver weight of fasting animals. In another study in rats fasted for 48 h and then refed a fat free diet for the following 2 days results were more conclusive [18]. Specific precipitation of [u-t4C]palmitoyl CoA binding protein with anti-Z immunoglobulin G showed 89% more FABP bound palmitoyl CoA in refed than fasted rats (Table 1I). These responses in FABP concentration to starvation and refeeding further support the concept of involvement of FABP in fatty acid metabolism. Starvation is similar to uncontrolled diabetes in ternrs of fatty acid esterification and oxidation. In the diabetic liver [l-14C]oleic acid binding to FABP was found to be 37% less than that of the control liver [19]. The decreased binding of ligand was reversed by administration o f insulin to diabetic rats (Table 11). The mechanism of this change or potential role of FABP in diabetes is not clear, bul it is likely similar to that of fasting. It would be of interest to correlate the protein bound fatty acids in diabetic liver with the immunoprecipitable FABP. Besides dietary variations, pharmacological manipulations by hypolipidemic drugs seems to effect the hepatic content of FABP. Incorporation of Clofibrate (0.25%) into normal chow diet increased liver FABP by 3-fold [20], and the increase has been correlated with a higher uptake of fatty acids from perfusate [21]. Experiments in vivo showed no significant increase in the incorporation of [14C] oleic acid into triacylglycerol in spite of high FABP content in clofibtic acid fed rats [22]. On the other hand, in vitro studies reported a concomitanl increase in FABP content and biosynthesis of triacylglycerol in rats ted a diet containing cholestyramine [23]. The discrepancy between in vilro and whole aninral results may be due to differences in two drugs. This leads to the speculation that FABP may not always be directly involved in the synthesis of triacylglyccrol
TABLE 11 Tissue
Liver Liver Liver Liver Liver Liver
Dietary treatment
Starved 48 h Refed fat free diet 48 h Diabetic Diabetic + insulin Starved 48 h Refed 20 h
FABP binding activity values are mean _+S.E. Control
l'reatlnenl
27.5 ± 4.3 27.5 +4.3 27.5 ±4.3 27.5 _+4.3
51.9 73.5 17.2 ± 4.5 25.0 ± 0.9 17.8 _+1.5 19.2 + 1.7
anmol bound palmitoyl CoA precipitated from FABP using antibody. bpereentage of added oleic acid bound to FABP fraction.
Ref.
18a 18 a 19 b 19 b 19b 19 b
135 under different pharmacological conditions, but supports the notion of FABP participation in the compartmentalization and channeling o f fatty acids toward initochondria or smooth endoplasmic reticulum under various nutritional, hormonal and pharmacological conditions. The precise mechanism or factors responsible for the dietary induction of FABP ave unclear at the present time. It is not known which fatty acids or their CoA derivative induce the synthesis of FABP. Knowledge of FABP synthesis in relation to diet can be expanded utilizing the cloned cDNAs as a potential tool. FABP synthesis can be controlled at the gene transcription or mRNA translation level. Control of FABP synthesis by transcriptional or translational mechanism under various nutritional conditions still awaits future research in this direction.
Acknowledgements Work in author's laboratory was supported by U.S.P.H.S. N1H SCOR Grant 1 P50 HL 27358 and R01 GM-14033. References l K. Kamisaka, M. Hirano and M. Tsuru, Experientia, 34 (1978) 1265-1266. 2 R.U. Haq, L. Christodoulides, B. Ketterer and E. Shrago, Biochim. Biophys. Acta, 713 (1982) 193--198. 3 C.A. Goresky, D.S. Daly, S. Mishkin and I.M. Arias, Am. J. Physiol., 254 (1978) 542553. 4 M.Y.C. Wu-Rideout, C. Elson and E. Shrago, Biochem. Biophys. Res. Commun., 71 (1976) 809-816. 5 D.A. Bumett, N. Lysenko, J.A. Manning and R.K. Ockner, Gastroenterology, 77 (1979) 241-249. 6 N. Fournier, M. Geoffroy and J. Deshusses, Biochim. Biophys. Acta, 533 (1978) 457-464. 7 E.L. Appelkvist and G. Dallner, Biochim. Biophys. Acta, 617 (1980) 156-160. 8 R.K. Ockner and J.A. Manning, J. Clin. Invest., 54 (1974) 336 338. 9 D.H. Alpers, A.W. Strauss, R.K. Ockner, N.M. Bass and J.l. Gordon, Proc. Natl. Acad. Sci. U.S.A., 81 (1984) 313-317. 10 J.B.D. Katongole and B.E. March, Poultry Sci., 58 (1979) 372-375. 11 J.B.D. Katongole and B.E. March, Poultry Sci., 59 (1980) 819 827. 12 R.U. Haq and E. Shrago, Nutr. Res., 3 (1982) 329-333. 13 B. Riistow, R. Risse and D. Kunze, Acta Biol. Med. Germ., 41 (1982) 439-445. 14 B. Said and H. Schulz, J. Biol. Chem., 259 (1984) 1155-1159. 15 J.l. Gordon, N. Elshourbagy, J.B. Lowe, W.S. Liao, D.H. Alpers and J.M. Taylor, J. Biol. Chem., 260 (1985) 1995-1998. 16 G.R. Herzberg and M. Rogerson, Nutr. Res., 1 (1981) 601-607. 17 L.B. Stein, S. Mishkin, G. Fleischner, Z. Gatmaitan and I.M. Arias, Am. J. Physiol., 31 (1976) 1371-1376. 18 N. lritani, E. Fukuda and K. lnoguchi, J. Nutr. Sci. Vitaminol., 26 (1980) 271-277. 19 R. Brandes and R. Arad, Biochim. Biophys. Acta, 750 (1983) 334-339. 20 G. Fleischner, D.K.F. Meijer, W.G. Levine, Z. Gatmaitan, R. Gluck and I,M. Arias, Biochem. Biophys. Res. Commun., 67 (1975) 1401-1407. 21 G. Renaud, A. Foliot, R. Infante, Biochem. Biophys. Res. Commun., 80 (1978) 327-334. 22 Y. Kawashima, S. Nakagawa and H. Kozuka, J. Pharmacobiodyn., 5 (1982) 771-779. 23 H.J.M. Kempen, J.F.C. Glatz, J. DeLange and J.H. Veerkamp, Biochem. J., 216 (1983) 511-514.