A wide variety of disturbances in carnitine metabolism of humans has been reported during the past decade. (Broquist & Borum, 1982; Borum, 1983). Many of ...
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Edwards, M. R.. Bird. M. I. & Saggerson, E. D. (1985) Biocheni. J . 230. 169-179 Fiol. C. J. & Bieber. L. L. (1984) J . B i d . Chem. 259. 13084-13088 Flatt, J. P. (1970) J . Lipid Rrs. 1 I. I3 I 143 Flatt, J. P. & Ball, E. G . (1964) J . Biol. Chem. 239, 675--685 Gamble, M . S. & Cook. G. A. (1985) J . Biol. Chem. 260. 95169519 Harano, Y.. Kowal, J.. Yamazaki, R., Lavine, L. & Miller, M. (1972) Arch. Biochem. Biophj~s.153, 426 437 Harper. R. D. & Saggerson, E. D. (1975) Biochem. J . 152, 485- 494 Harper, R. D. & Saggerson. E. D. (1976) J . Lipid Res. 17, 516526 McGarry, J. D. & Foster, D. W. (1980) Annu. Rev. Biochem. 49, 395- 420 McGarry, J. D., Leatherman, G. F. & Foster, D. W. (1978) J . B i d . Chem. 253, 4128-4136 McGarry, J. D., Mills, S. E., Long, C. S. & Foster, D. W. (1983) Biochrm. J. 214. 21-28 Mills. S. E.. Foster, D. W. & McGarrv. J. D. (1983) Biochem. J . 214. 83 91 Mills, S. E., Foster. D. W. & McGarry. J . D. (1984) Biochem. J. 219. 601 608 Nichols. D. & Locke. R. (1983) in Mummuliun Thermogenesis (Girardier, L. & Stock, M., eds.). pp. 8 -49, Chapman and Hall, London Paulson. D. J.. Ward. K. M. & Shug, A. L. (1984) FEES L a / [ . 176. 381- 384 Rath, E. A., Salmon, D. M. W. & Hems. D. A. (1979) FEBS Lett. 108, 33 36 Robinson, I. N. & Zammit, V. A. (1982) Biochem. J . 206. 177-179 Saggerson, E. D. (1982) Biochrm. J. 202. 397 405 Saggerson. E. D. &Carpenter, C. A. (1981~)FEBS Lett. 129.225 228 Saggerson. E. D. &Carpenter, C . A. (198lh) FEES Let/. 129,229-232 Saggerson. E. D. & Carpenter. C. A. ( I 981c.)FEBS Let/. 132. I66168 Saggerson. E. D. & Carpenter. C . A. (1982) Biochem. J . 204, 373-375 Saggerson. E. D. & Carpenter, C. A. (1983) Biochem. J. 210, 591-597 Saggerson. E. D. & Carpenter. C. A. (1986) Biocheni. J . 236, 137-141 Saggerson. E. D.. Carpenter. C . A. & Tselentis, B. S. (1982) Biochem. J. 208. 667 672 Saggerson. E. D., Bird, M. 1.. Carpenter. C. A,. Winter, K. A. & Wright. J. J. (1984) Biochem. J. 224. 2 0 1 ~206 Smith. S. J. & Saggerson, E. D. (1979) Int. J . Biochem. 10. 785-790 Stakkestad, J. A. & Bremer. J. (1983) Biochim. Biophys. Acra 750, 244-252 Stephens. T. W., Cook. G. A. & Harris, R. A. (1983) Biochem. J. 212. 521 524 Trayhurn. P. (1981) Biochim. Biophys. Acru 664. 549-560 West. D. W..Chase, J. F. A. &Tubbs. P. K. (1971) Biochem. Biophys. Rex. Commun. 42. 912 918 Zammit. V. A. (1986) Biochrm. Soc. Truns. 14. 676 679 Zammit, V. A. & Corstorphine. C. G . (1985) Biochem. J. 231,343--347 ~
Fig. 2. Effect of p H on rat heart CPT, activity CPT, activity was measured as described in Fig. 1 using- 40.W M palmitoyl-CoA with ( 0 ) or without (0) 3 pM-malonyl-CoA. ( a ) Absolute activities; ( b ) activities expressed relative to the pH = 7.4 value. The values are means of two similar experiments.
relevant to control of the enzyme in skeletal muscle in exercise or in ischaemic heart muscle. I thank the Medical Research Council for financial support.
Bergseth. S.. Lund, H. & Bremer, J. (1986) Biochem. Soc. Truns. 14. 671 672 Bieber. L. L. & Fiol. C. J. (1986) Biochem. Soc. Trans. 14, 674 676 Bird, M. I . & Saggerson. E. D. (1984) Biochem. J. 222. 639 647 Bird. M. I. & Saggerson. E. D. (1985) Biochrm. J. 230. I61 I67 Bird. M. 1.. Munday. L. A., Saggerson. E. D. & Clark. J. B. (1985) Biochrm. J . 226, 323-330 Bremer, J. (1981) Biochim. Bic1phy.s. Actu 665, 628 631 Buckley. M. G . & Rath. E. A. (1985) Biochem. Soc. Truns. 13.946 947 Chase. J. F. A. & Tubbs, P. K. (1969) Biochrm. J . I 1 I , 225 235 Chase. J. F. A. & Tubbs, P. K. (1972) Birichrm. J. 129. 55 65 Cook, G. A. (1984) J. B i d . Chem. 259. 12030 12033
Disturbances in carnitine metabolism PEGGY R. BORUM 409 Food Science Building, University of Florida. Guinesvilli~.FL 3261 I , U . S . A . A wide variety of disturbances in carnitine metabolism of humans has been reported during the past decade (Broquist & Borum, 1982; Borum, 1983). Many of the disturbances have been recognized in the clinical setting and associated with a spectrum of clinical symptoms and abnormal clinical laboratory results. Our understanding of carnitine metabolism does not permit an explanation for many of these disturbances or for the association of many of the observed symptoms and abnormal laboratory results with disturbances in carnitine metabolism. Carnitine has a more pivotal function in energy metabolism than has been previously recognized. Causes of disturbances in carnitine metabolism Table 1 lists several areas of metabolism which should be considered in the search for the biochemical lesion resulting in an observed disturbance of carnitine metabolism.
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Altered activity of one or more steps of the carnitine biosynthetic pathway is a prime candidate for causing a disturbance in carnitine metabolism. The intermediates of the pathway and the proteins catalysing each step have been identified (Bremer. 1983; Borum, 1983). Our understanding of the regulation of the pathway is limited. Activity values for a particular enzyme determined by an assay in vitro using a homogenized biopsy sample, may not be indicative of the
Table 1. C'auses of' disturbances in carnitine metabolism ( I ) Altered carnitine biosynthetic pathway (2) Altered transport of carnitine to its site of action (3) Altered enzymes that use carnitine as a
.,P -I , hst .,...r2 . tp .I
(4) Altered interaction of carnitine with 'other' metabolic pathways
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682 activity under physiological conditions with physiological regulation factors in place. Additional information concerning each enzyme protein, the kinetic characteristics of the enzymic reaction, and the mechanism of regulating the enzymic steps of the carnitine biosynthetic pathway are needed. Altered transport of carnitine from its site of synthesis or its site of entry into the body to its site of action has been implicated in patients with normal carnitine concentrations in some tissue or fluid sample and low carnitine concentrations in other samples (Broquist & Borum, 1982). Research concerning the mechanism of carnitine transport from one tissue to another is an active area of research, but our knowledge is rudimentary at present (Bremer, 1983; Borum, 1983). Additional information concerning the transport mechanism and its regulation are required before meaningful investigations testing possible alterations in carnitine transport as the biochemical lesion resulting in disturbances in carnitine metabolism can be performed in the clinical setting . Alterations in one of the proteins that use carnitine as a substrate or alterations in the regulation of one of the enzyme reactions will also result in disturbances in carnitine metabolism, although the carnitine concentration may be normal or elevated (Bremer, 1983). These enzymes have been discussed at length by other contributors to this Colloquium, and the discussion will not be repeated here. Many observations of the past decade can be explained only by postulating a metabolic involvement of carnitine in metabolic pathways not usually associated with the wellestablished functions of carnitine. A biochemical lesion affecting one of these pathways might in turn alter the involvement of carnitine resulting in a disturbance of carnitine metabolism. At the present time this area includes hypotheses and associations with a limited amount of hard data that can be utilized in determining the cause of a disturbance in carnitine metabolism. Symptoms and abnormal laboratory results associated with disturbances in carnitine metabolism Table 2 lists some of the clinical symptoms, and Table 3 lists some of the abnormal clinical laboratory results, associated with disturbances in carnitine metabolism. The first description of a patient with carnitine insufficiency (Engel & Angelini, 1973) included muscle weakness and skeletal muscle type I fibres filled with lipid-containing vacuoles associated with the disturbances in carnitine metabolism. These symptoms and abnormal laboratory results were consistent with the well-established function of carnitine in transporting long-chain fatty acids into the matrix of the mitochondria. One could rationalize that the carnitine insufficiency severely impaired the capability of the skeletal muscle to utilize fatty acids for metabolic energy and as a result accumulating triglycerides formed lipid droplets in the
Table 2. Clinical symptoms associated with disturbances in carnitine metabolism (I) (2) (3) (4) (5) (6)
Muscle weakness Cardiac hypertrophy and dysfunction Poor appetite Enlarged liver Central nervous system disturbances Acute severe symptoms during periods of starvation and/or infectious illness (7) Muscle pain and dark urine after periods of severe exercise and starvation
Table 3. Abnormal clinical laboratory results associated with disturbances in carnitine metabolism (I) (2) (3) (4) (5) (6) (7)
Lipid accumulation in tissue Hypertriglyceridaemia Hypoglycaemia Hypoketonaemia Hyperammonaemia Negative nitrogen balance Energy deficit
tissue. The muscle weakness could be explained by lack of adequate metabolic energy for contraction and perhaps even mechanical interference of the contraction process by the lipid vacuoles. As more patients with carnitine insufficiency were described, many of the symptoms could continue to be rationalized, at least in part, by the wellestablished function of carnitine. Cardiac hypertrophy (with lipid vacuoles in cardiac tissue at autopsy), enlarged fatty liver and hypertriglyceridaemia were not inconsistent with an impaired ability to utilize fatty acids. When patients were described with non-ketotic hypoglycaemia associated with disturbances in carnitine deficiency (Broquist & Borum, 1982; Borum, 1983), it was possible to suggest that the impairment in the ability to produce ketone bodies was the result of the inability to transport fatty acids into the mitochondria to serve as a substrate of ketogenesis, and that the same metabolic lesion resulted in a reduction in the production of NADH, which in turn is needed for reducing equivalents in gluconeogenesis. However, the fact that other compounds such as amino acids provide substrates for ketogenesis, which according to the well-established function of carnitine would not require carnitine and thus would not be affected by carnitine insufficiency, raises the question of whether or not the disturbance in carnitine metabolism may have an additional effect on ketogenesis. Research in the 1960s suggested that additions of carnitine to liver slices or kidney cortex slices stimulates gluconeogenesis (Benmiloud & Freinkel, 1967) and raises the same question of whether or not carnitine has an effect on gluconeogenesis in addition to stimulating /I-oxidation of fatty acids with production of NADH. Several patients with disturbances in carnitine metabolism have been shown to have central nervous system disorders and hyperammonaemia. The symptoms may be very similar to the symptoms of Reye’s syndrome (Borum, 1983). Many patients with disturbances in carnitine metabolism have also been shown to have negative nitrogen balance. Animals maintained on carnitine-free total parenteral nutrition have improved nitrogen balance when carnitine is added to the total parenteral nutrition solutions (Bohles et al., 1984). Critically ill patients who have experienced a great deal of metabolic stress, such as patients with both severe trauma and sepsis, have a wide variety of symptoms which may be the result of a metabolic energy deficit, as demonstrated by a reduction in muscle ATP and phosphocreatine (Liaw, 1985). These patients have also been shown to have disturbances in carnitine metabolism. The symptoms listed above are only associated with disturbances in carnitine metabolism at the present time and the relationship may be nothing more than unrelated association. However, it is also quite possible and perhaps likely that the relationship is more than merely association. These clinical observations should be used to assist basic scientists in designing experiments to delineate metabolic functions for carnitine that have remained unrecognized to this point. Elucidation of the metabolic functions of carnitine In addition to the well-established function of carnitine in
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616th MEETING, LONDON transporting long-chain fatty acids into the matrix of the mitochondria, it now appears that the role of carnitine in transporting short-chain acyl groups into the matrix of the mitochondria is more important than previously recognized. The dependence of medium-chain and short-chain acyl groups upon carnitine for intercompartment transport appears to be increased in metabolic conditions characterized by reduced energy charge potential (Otto, 1984). Disturbances in carnitine metabolism may more severely affect the individual with hypermetabolism than the individual with limited metabolic stress. Disturbances in carnitine metabolism may affect a variety of metabolic pathways involving acyl compounds in addition to fatty acid oxidation when hypermetabolism is present. The functions of carnitine may actually vary in response to the metabolic state of the individual. For example, it was suggested several years ago (Pearson & Tubbs, 1967) that carnitine functions in the buffering of bound coenzyme A to free coenzyme A. A disturbance in carnitine metabolism altering carnitine’s capacity to buffer the ratio of bound coenzyme A to free coenzyme A may result in no detectable pathology in the individual experiencing limited metabolic stress but may be expressed as severe clinical symptoms when the individual experiences metabolic stress. Accumulation of acyl-CoA compounds has been shown to inhibit a growing list of enzymes that have critically important roles in metabolism (Stumpf et al., 1985). A disturbance in carnitine metabolism may not be detectable in the individual unless the stimulus (such as cardiac ischaemia) is present which induces the accumulation of the acyl-CoA compounds. When acyl-CoA compounds accumulate to toxic levels, carnitine appears to function in the removal of the compounds by converting them to acylcarnitine, which is excreted in the urine. During these physiological situations, it is difficult to discern whether the stimulus causing an accumulation of acyl-CoA compounds is also causing a disturbance in carnitine metabolism, or whether the disturbance in carnitine metabolism was present but undetected before the stress of the stimulus causing accumulation of acyl-CoA, or whether protection against accumulation of acyl-CoA is a function of carnitine which requires physiological concentrations of carnitine that are higher than can be maintained by combined normal endogenous synthesis and the quantity of carnitine consumed in a typical diet. In the clinical setting the first or third possibility would indicate that all patients with accumulation of acyl-CoA compounds need treatment for the disturbance in carnitine metabolism, but the second possibility would indicate that only patients with a preexisting disturbance in carnitine metabolism would need treatment for the disturbance in carnitine metabolism in addition to the treatment for the cause of the accumulation of acyl-CoA compounds.
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Our laboratory has recently suggested that carnitine may function in storing readily transportable metabolic energy (Borum, 1986). The suggestion that carnitine can store metabolic energy is based upon the fact that the acylcarnitine molecule maintains the same high metabolic energy that was in the precursor acyl-CoA molecule. The suggestion that the metabolic energy stored in the acylcarnitine molecule is readily transportable is based upon the fact that metabolic mechanisms are known which are needed to transport an acylcarnitine synthesized using the metabolic energy in one cellular compartment to another compartment within the cell or perhaps even to another cell. Once the acylcarnitine has reached the cellular compartment which needs additional high energy compounds, the acylcarnitine can be converted to acyl-CoA and utilized. Once again this function of carnitine may be of critical importance to the patient with a decreased energy charge potential, such as the septic severely traumatized patient who must ration all available metabolic energy. However, this function of carnitine may have little importance in the healthy individual with a surplus of metabolic energy. Observations of disturbances in carnitine metabolism are forcing us to recognize that carnitine has a pivotal role in energy metabolism, which includes metabolic functions that are only beginning to be understood. These observations also demonstrate that a healthy animal model may not be appropriate for investigations concerning some of the functions of carnitine. The functions of carnitine may be expressed according to the state of metabolic stress being experienced. Investigations of the disturbances in carnitine metabolism hold great promise for obtaining information needed both by the basic scientist and the clinician. I thank Ms. Robin Adkins and Ms. Tracie Poole for their assistance in preparing the manuscript. This work was supported in part by funds from the Florida Agricultural Experiment Station.
Benmiloud, M . & Freinkel, N . (1967) Mefaholism 16,658-669 Bohles. H..Segerer, H . & Fekl. W. (19x4) J . Parenrer. Enrer. Nufr. 8. 9-13
Borum. P. R. (1983) Annu. Rev. Nutr. 3,233-259 Borum. P. R . (1986) in Clinicul A.specr.s of Human Curnitine Deficiency (Borum, P. R., ed.), Pergamon Press, New York, in the press Bremer. J . (1983) Physiol. Rev. 63, 1420-1480 Broquist. H.P. & Borum. P. R. (1982) Adv. Nutr. Res. 4, 181-204 Engel, A. G . & Angelini, C. (1973) Science 179, 899-902 Liaw. K . Y . (1985) J . Purmter. Enter. Nurr. 9, 28-33 Otto, D. A. (1984) J . Biol. Chem. 259, 5490-5494 Pearson. D. J . & Tubbs, P. K . (1967) Biochem. J . 105, 953-963 Stumpf, D. A.. Parker, W. D. & Angelini, C. (1985) Neurology 35, 1041 1045
