143
Lopez de Romana G, Brown KH, Bravo N, Balazar OG, Kanashiro HC. The incidence and etiology of infantile diarrhoea and major routes of transmission in Huascar, Peru. Am J Epidemiol 1989; 129: 785-99. 5. Elegbe IA, Ojofeitimi EO. Early initiation of weaning and proliferation of bacteria in Nigerian infants. Clin Pediatr 1984; 23: 261-64. 4. Black RE,
6. Gordon JE, Chinese ID, Lyon JB. Weanling diarrhoea. Am J S Sci 1963; 245: 345-77. 7. Rowland MGM, Barrell RAE, Whitehall RG. Bacterial contamination in traditional Gambian weaning foods. Lancet 1978; i: 136-38. 8. Esrey SA, Feachem RG. Interventions of the control of diarrhoeal
diseases among young children. Promotion for Food
Hygiene 1989;
WHO/CDD/89.30. 9. Nyaga PM, Kagiko MM, Gathuma JM. Milk hygiene in nomadic herds in Kenya evaluated by bacterial isolation, bacterial viability trials in traditionally fermented milk and drug sensitivity. Bull Animal Health Prod Afr 1982; 30: 19-24. 10. Tomkins AM, Alnwick D, Haggerty P. Fermented foods for improving child feeding in eastern and southern Africa: a review. In: Alnwick D, Moses S, Schmidt OG, eds. Improving young child feeding in Eastern and Southern Africa: household level food technology. Proceedings of a workshop. Nairobi, 1988: 136-167. 11. Mensah PPA, Tomkins AM, Drasar BS, Harrison TJ. Effect of fermentation of Ghanaian maize dough on the survival and proliferation of4 strains of Shigella flexneri. Trans R Soc Trop Med Hyg 1989; 82: 635-36. 12. Sakoane AL, Walsh A. Bacterial properties of traditional sour porridges in Lesotho. In: Alnwick D, Moses S, Schmidt OG, eds. Improving young child feeding in Eastern and Southern Africa: household level food technology. Proceedings of workshop. Nairobi, 1988: 136-67. 13. Nout MJR, Hautvast JGAJ, Van der Haar F, Marks WEW, Rombarts FM. Energy, protein and microorganisms: the formulation and microbiological stability of cereal-based weaning foods. In: Alnwick D, Moses S, Schmidt OG, eds. Improving young child feeding in Eastern and Southern Africa: household level food technology. Proceedings of workshop. Nairobi, 1988: 245-260.
14. Harrigan WF, McCance ME. Laboratory Methods in Dairy Microbiology. London: Academic Press, 1976. 15. Thatcher FS, Clark DS. Microorganisms in foods. Vol I. Their significance and methods of enumeration. Toronto: University of Toronto Press, 1968. Alim Falkow S. Detection of
ARMA, So M, Samadpour-Motalebi M, enterotoxigenic. Escherichia coli by DNA colony hybridisation. J Infect Dis 1980; 142: 892-95. 17. Grunstein M, Hogness DS. Colony hybridisation: a method for the isolation of cloned DNAs that contain specific gene. Proc Natl Acad Sci 16.
Moseley SL, Hug I,
USA; 72: 3961-65. 18. Mason PJ, Williams JG. Hybridisation in the analysis of recombinant DNA. In: Hames BD, Higgins SJ, eds. Nucleic add hybridisation: a practical approach. Oxford: IRL Press, 1985: 113-25. 19. Gennaro ML, Greenway PJ, Broadbent DA. The expression of biologically active cholera toxin in Escherichia coli. Nucleic Acids Res 1982; 10: 4883-90. 20. Lathe R, Hirth P, De Wilde M, Lecog JP. Cell free synthesis of enterotoxin of Esherichia coli from a cloned gene. Nature 1980; 284: 473-74. 21. De Wilde M, Ysebert M, Hartford M. DNA sequence of STa2 enterotoxin gene from an Escherichia coli strain of human origin. In:
Levey SB, Clowes RC, Koenig EK, eds. Molecular biology, pathogenicity and ecology of bacterial plasmids. New York: Plenum Publishing Corporation, 1981: 596. 22. Feinberg AP, Vogelstein B. A technique for radiolabelling DNA restriction endonuclease fragments to high specific activity. Anal Biochem 1983; 132: 6-13. 23.
Banigo EOI, Muller HG. Carboxylic acid patterns J Sci Food Agric 1972; 23: 101-11.
in
ogi fermentation.
24. Andah A, Muller HG. Studies on koko, a Ghanaian fermented maize porridge. Ghana J Agric Sci 1973; 6: 103-08. 25. Akinrele IA. Fermentation studies on maize during the preparation of traditional African starch-cake food. J Sci Food Agric 1970; 21: 619-25. 26. Muller HG. Traditional cereal processing in Nigeria and Ghana. Ghana J Agric Sci 1970; 3: 187-95.
Transient release of lipid peroxides after coronary artery balloon angioplasty
Free radical
production may cause myocardial damage during reperfusion of ischaemic myocardial tissue; when free radicals interact with polyunsaturated fatty acids or their esters, lipid peroxides are produced. A product of lipid peroxidation, malondialdehyde, was measured in 10 subjects with stable angina who underwent angioplasty of a proximal high-grade stenosis (over 90%) of the left anterior descending coronary artery. In all subjects the duration of balloon occlusion was 60 s. Blood was withdrawn from the great cardiac vein immediately before balloon inflation (T0), immediately after balloon deflation (T60), 15 s after balloon deflation (T75), and 1 min after balloon deflation (T120). There was a significant increase in malondialdehyde at T60 compared with T0 for the first balloon inflation (mean increase 0·3 µmol/l [95% confidence limits 0·1, 0·5]), and at both T60 (0·31 µmol/l [0·15, 0·47]) and T75 (0·22 µmol/l [0·04. 0·40]) for the second balloon inflation. This model could be used to assess antioxidant effects of drugs.
Introduction
Oxygen-derived free radicals are produced after reperfusion of ischaemic myocardium in animals. Radicals have been directly detected by electron spin resonance,2 and freeradical damage products have also been measured.3 Myocardial reperfusion is associated with several potentially harmful effects, including transient impairment of myocardial function (myocardial "stunning"),4 an increased incidence of dysrhythmias,55 and possibly increased myocardial necrosis.6 These adverse effects can be reduced by treatment with various antioxidants at the time of myocardial reperfusion,’,8 which indicates that free-radical formation is an important factor in their pathophysiology.
ADDRESSES: Regional Medical Cardiology Centre, Royal Victoria Hospital, Belfast (M. J. D. Roberts, MRCP, T. G. Trouton, MD, M. M. Khan, FRCP, S. W. Webb, MD, C M. Wilson, MD, G. C. Patterson, MD, A. A. J. Adgey, MD); and Department of Chemical Pathology, Queen’s University, Belfast, UK (I. S. Young, MRCP, E. R. Trimble, MD). Correspondence to Dr A. A. J. Adgey, Regional Medical Cardiology Centre, Royal Victoria Hospital, Belfast BT12 6BA, UK.
144
But how far can the results of these animal experiments be extrapolated to man? In particular, most animal studies have used a period of complete coronary artery occlusion of sudden onset, followed by equally sudden myocardial reperfusion; this sequence would clearly be unusual in man, for whom coronary artery occlusion may be incomplete, and myocardial reperfusion gradual. Thus it is important to look
After the second balloon inflation at T7s the result in 1 patient was grossly aberrant (malondialdehyde concentration more than doubled after angioplasty) and was excluded from the analysis. The differences between the observations at each time-point were compared and 95% confidence limits calculated. Such differences between patients are statistically independent. Sequential balloon inflations were compared separately.
for evidence of free-radical formation in man under conditions of controlled transient coronary artery occlusion and myocardial reperfusion; such conditions are met by subjects who undergo percutaneous transluminal coronary angioplasty (PTCA) for stable angina. Free-radical species are very short-lived and can only be directly measured by electron spin resonance-a technique difficult to apply in man. However, when free radicals are produced they react with various molecules to form products which can be measured as an indirect indicator of free-radical activity. Lipid peroxides are produced when free radicals interact with polyunsaturated fatty acids or their esters. As an indicator of lipid peroxidation we have measured malondialdehyde, a product of free-radical attack on polyunsaturated fatty acids, in a group of patients who underwent PTCA. Serial samples of blood from the great cardiac vein were used to represent venous blood draining directly from ischaemic myocardial tissue as it undergoes
Results
reperfusion.
Mean malondialdehyde concentrations in arterial (2-15
pmol/1) and venous (2-03 Nmol/1) control samples were similar to values previously reported by other workers.10 There was no significant difference between initial (2-15 Ilffioljl) and final (1 -96 trnol/1) arterial or great-cardiac-vein malondialdehyde concentrations (2-03 and 2-02 J.Ul101/1, respectively). There was no significant arteriovenous gradient for malondialdehyde in pre-inflation or postinflation samples. Results for first and second balloon inflations are recorded in the table. Mean malondialdehyde concentrations were significantly raised at T 60 compared with To for the first balloon inflation (0-3 umol/1 [95% confidence limits 0-1, 05]), and were significantly raised in comparison with To at both T60 (0-31 Ilffioljl [015, 0-47]) and T7s (0-22 umol/1 [004, 0-4 for the second balloon inflation. By T 120 malondialdehyde concentrations had returned to baseline levels. Discussion
Patients and methods patients (aged 26-72 years) with stable angina who were to undergo angioplasty of a single stenosis (over 90%) of the left anterior descending coronary artery were recruited. Subjects with left bundle-branch block, valvular heart disease, or diabetes mellitus were excluded. Conventional angioplasty protocol was followed in all cases. Patients received 10 mg intravenous diazepam emulsion before the procedure, a heparin bolus of 10 000 units was given intravenously, and an infusion of dextran 40 was commenced at a 10
of 20 drops per min. In addition to arterial cannulation for balloon angioplasty, a 7 French multipurpose catheter was introduced via the femoral or brachial vein into the great cardiac vein. After a stabilisation period of 5 min, baseline blood samples were taken from the arterial and venous catheters and further control samples were taken 2 min later. Subsequent venous blood samples were taken immediately before balloon inflation (T), immediately after balloon deflation (T 60; the occlusion time was 60 s in all subjects); 15 s after balloon deflation (T7s); and 60 s after balloon deflation (T 120)’ This sequence of sampling was repeated for up to 3 subsequent inflations. Further arterial and venous samples were taken 5 min after completion of angioplasty. The protocol was approved by the ethical committee of the Queen’s University, Belfast, and patients gave full informed consent. Blood samples were taken into dipotassium ethylenediaminotetraacetate and stored on ice before separation of plasma at 2000 gat 4°C. Malondialdehyde content was measured by fluorometric assay of the adduct formed with thiobarbituric acid as described by Yagi.99 The intra-assay coefficient of variation was 5-5% and the interassay coefficient of variation 70% at a malondialdehyde concentration of rate
1-76
Eunol/1.
INCREASE IN GREAT-CARDIAC-VEIN MALONDIALDEHYDE (MDA) CONCENTRATION AFTER FIRST AND SECOND PTCA BALLOON INFLATIONS
Malondialdehyde is a measure of lipid peroxidation, which when polyunsaturated fatty acids are attacked by free
occurs
radicals
of
Raised concentrations of oxygen. have been observed in malondialdehyde reperfused myocardial tissue in animals,3and in man malondialdehyde concentrations are raised in several conditions where free-radical damage has been strongly implicated (eg, rheumatoid
arthritis" and diabetes mellitus’2). been measured by high-performance Malondialdehyde has liquid chromatography,l° and spectrophotometric13 or fluorometric9 measurement of its thiobarbituric acid adduct. Some interference has been observed in the spectrophotometric and fluorometric assaysl4 but, in general, malondialdehyde correlates well with other markers of free-radical activity.1s As we observed consistent changes in serial measurements taken over a short period of time, we do not believe that interference in the assay is likely to have been a significant problem in our subjects. In man, raised malondialdehyde concentrations have been reported in peripheral venous blood in patients in whom successful myocardial reperfusion was achieved after thrombolytic therapy for acute myocardial infarction in comparison with subjects in whom reperfusion did not occur.16 During PTCA we found a transient rise in malondialdehyde after balloon deflation in blood from the great cardiac vein, which returned to baseline concentrations within 1 min. This observation is consistent with the concept that a burst of free-radical production occurs during the reperfusion which follows the brief ischaemia of balloon inflation. It is also possible that free-radical production is secondary to cellular damage that occurs during the ischaemia-but such damage seems unlikely during this short period of ischaemia. Lipid peroxides are also known to be present in atherosclerotic plaque tissue17 and we cannot exclude the possibility that the rises in malondialdehyde concentration are due to mechanical disruption of plaque during balloon inflation. In addition, malondialdehyde is a potentially harmful
145
substance that may damage extracellular components,l8 which may themselves be cytotoxic to endothelial cells.19 Whatever the cause of the transiently raised malondialdehyde concentration, we believe that angioplasty to a high-grade proximal coronary artery stenosis, with subsequent release of the balloon, represents a setting in which the therapeutic usefulness of pharmacological agents as free-radical scavengers can be investigated in man. If such agents reduce the concentrations of biochemical markers of free-radical damage during coronary artery angioplasty, further support will be provided for trials of adjuvant antioxidant therapy with thrombolytic agents in acute myocardial infarction. REFERENCES 1. Dart RC, Sanders AB. Oxygen free radicals and myocardial reperfusion injury. Ann Emerg Med 1988; 17: 53-58. 2. Bolli R, Patel BS, Jeroudi MO, Lai EK, McCay PB. Demonstration of free radical generation in "stunned" myocardium of intact dogs with the use of the spin trap &agr;-phenyl N-tert-butyl nitrone. J Clin Invest 1988; 82: 476-85. 3. Guarnieri C, Flamigni F, Caldarera CM. Role of oxygen in the cellular damage induced by re-oxygenation of hypoxic heart. J Mol Cell Cardiol 1980; 12: 797-808. 4. Bolli R, Jeroudi MO, Patel BS, et al. Marked reduction of free radical generation and contractile dysfunction by antioxidant therapy begun at the time of reperfusion. Circ Res 1989; 65: 607-22. 5. Woodward B, Zakaria MNM. Effect of some free radical scavengers on reperfusion induced arrhythmias in the isolated rat heart. J Mol Cell
Cardiol 1985; 17: 485-93. 6. Ambrosio G, Becker LC, Hutchins GM, Weisman HF, Weisfeldt ML. Reduction in experimental infarct size by recombinant human superoxide dismutase: insights into the pathophysiology of reperfusion injury. Circulation 1986; 74: 1424-33.
7. Opie LH. Reperfusion injury and its pharmacologic modification. Circulation 1989; 80: 1049-62.
Myers ML, Bolli R, Lekich RF, Hartley CJ, Roberts R. Enhancement of recovery of myocardial function by oxygen free-radical scavengers after reversible regional ischaemia. Circulation 1985; 72: 915-21. 9. Yagi K. Assay for blood plasma or serum. Methods Enzymol 1984; 105: 8.
328-31.
10.
Wong SHY, Knight JA, Hopfer SM, Zahana O, Leach CN, Sunderman FW. Lipoperoxides in plasma as measured by liquid-chromatographic separation of malondialdehyde-thiobarbituric acid adduct. Clin Chem 1987; 33: 214-20. CR, Jackson PG, Highton J, van Rij AM. Lipid peroxidation and
11. Wade
in the synovial fluid and plasma of patients with rheumatoid arthritis. Clin Chim Acta 1987; 164: 245-50. 12. Sato Y, Hotta N, Sakamoto N, Matsuoka S, Ohishi N, Yagi K. Lipid peroxide level in plasma of diabetic patients. Biochem Med 1979; 21: 104-07. 13. Bird RP, Draper HH. Comparative studies on different methods of malondialdehyde determination. Methods Enzymol 1984; 105: 299305. 14. Knight JA, Pieper RK, McClellan L. Specificity of the thiobarbituric acid reaction: its use in studies of lipid peroxidation. Clin Chem 1988; 34: 2433-38. 15. Slater TF. Overview of methods used for detecting lipid peroxidation. Methods Enzymol 1984; 105: 283-93. 16. Davies SW, Ranjadayalan K, Wickens DG, Dormandy TL, Timmis AD. Free radicals, reperfusion and thrombolysis. Eur Heart J 1989; 10
malondialdehyde
17.
(suppl): 274. Ledwozyw A, Michalak J, Stepien A, Kadziolka A. The relationship between plasma triglycerides, cholesterol, total lipids and lipid peroxidation products during human atherosclerosis. Clin Chim Acta 1986; 155: 275-84.
18.
Fogelman AM, Shechter I, Seager J, Hokom M, Child JS, Edwards PA. Malondialdehyde alteration of low density lipoproteins leads to cholesteryl ester accumulation in human monocyte-macrophages. Proc Natl Acad Sci USA 1980; 77: 2214-18. SA, Galdal KS, Nilsen E. LDL-induced cytotoxicity and its inhibition by anti-oxidant treatment in cultured human endothelial cells and fibroblasts. Atherosclerosis 1983; 49: 23-30.
19. Evensen
BOOKSHELF Living Medicine Peter Richards. Cambridge: Cambridge University Press. 1990. Pp 187. 25,$39 hardback; /;8.95,$14.95 paperback. ISBN 0-521384788 and 0-521386284.
Living Medicine
has been written
partly to supplement Professor Richards’ already popular Learning Medicine. Whereas the earlier book is directed particularly to 17 and 18 year-olds considering a career in medicine, Living Medicine is addressed to students and house-officers who are planning their future careers. Written with clarity and a wry sense of humour, the book is short enough to be read in an evening. Its stated objective is to prepare senior students and house officers for their responsibilities and opportunities as doctors. Is this achieved? My house officer’s answer was guarded-yes, the book is entertaining, but too superficial in its kaleidoscopic survey of the many careers open to medical graduates. It alerts readers to the options, but little more. For a career that "dominates life" a fuller treatment would be preferable, if the book is to act as an important reference for students and new graduates. Nevertheless, there is a great deal to enjoy here. The pen portraits of different branches of medicine are miniature masterpieces. Not everyone will agree with Richards’ perception of their specialty-some even come across as decidedly grim. There is no attempt, for example, to disguise the arduous training necessary for general surgery. Running through the text is a lively appreciation of the source
conflicting demands of marriage, child-rearing, unsociable hours, hobbies, and life outside medicine. It is hard to discern whether the author advocates total dedication of one’s entire life to medicine or whether there is a covert admiration of those who do other things as well (as one of his "handful of doctors with a diploma in organ playing", I can vouch that a double act brings its own problems in plenty). As well as describing numerous possible careers, Richards writes with authority and wisdom about professional freedom and responsibility, quoting the Hippocratic oath and other key principles of conduct, confidentiality, and honesty. Further information on accreditation, General Medical Council regulations, and study leave is contained in appendices. Overall, medicine is presented as a hard task-master, but infinitely rewarding. There is little criticism of the present structure, although specialty overcrowding and over-long hours are acknowledged. The comment "appointment committees do not on the whole look imaginatively on missionary spirit or humanitarian concern" is unfortunately true, but it is sad that work abroad is seen as little more than an interruption to
diploma gathering. Wittily illustrated by
David Langdon, this enjoyable book can be recommended to a readership far wider than the students and house officers at which it is aimed. Southampton General Hospital, Southampton S09 4XY, UK
RICHARD GODFREY
Lopez de Romana G, Brown KH, Bravo N, Balazar OG, Kanashiro HC. The incidence and etiology of infantile diarrhoea and major routes of transmission in Huascar, Peru. Am J Epidemiol 1989; 129: 785-99. 5. Elegbe IA, Ojofeitimi EO. Early initiation of weaning and proliferation of bacteria in Nigerian infants. Clin Pediatr 1984; 23: 261-64. 4. Black RE,
6. Gordon JE, Chinese ID, Lyon JB. Weanling diarrhoea. Am J S Sci 1963; 245: 345-77. 7. Rowland MGM, Barrell RAE, Whitehall RG. Bacterial contamination in traditional Gambian weaning foods. Lancet 1978; i: 136-38. 8. Esrey SA, Feachem RG. Interventions of the control of diarrhoeal
diseases among young children. Promotion for Food
Hygiene 1989;
WHO/CDD/89.30. 9. Nyaga PM, Kagiko MM, Gathuma JM. Milk hygiene in nomadic herds in Kenya evaluated by bacterial isolation, bacterial viability trials in traditionally fermented milk and drug sensitivity. Bull Animal Health Prod Afr 1982; 30: 19-24. 10. Tomkins AM, Alnwick D, Haggerty P. Fermented foods for improving child feeding in eastern and southern Africa: a review. In: Alnwick D, Moses S, Schmidt OG, eds. Improving young child feeding in Eastern and Southern Africa: household level food technology. Proceedings of a workshop. Nairobi, 1988: 136-167. 11. Mensah PPA, Tomkins AM, Drasar BS, Harrison TJ. Effect of fermentation of Ghanaian maize dough on the survival and proliferation of4 strains of Shigella flexneri. Trans R Soc Trop Med Hyg 1989; 82: 635-36. 12. Sakoane AL, Walsh A. Bacterial properties of traditional sour porridges in Lesotho. In: Alnwick D, Moses S, Schmidt OG, eds. Improving young child feeding in Eastern and Southern Africa: household level food technology. Proceedings of workshop. Nairobi, 1988: 136-67. 13. Nout MJR, Hautvast JGAJ, Van der Haar F, Marks WEW, Rombarts FM. Energy, protein and microorganisms: the formulation and microbiological stability of cereal-based weaning foods. In: Alnwick D, Moses S, Schmidt OG, eds. Improving young child feeding in Eastern and Southern Africa: household level food technology. Proceedings of workshop. Nairobi, 1988: 245-260.
14. Harrigan WF, McCance ME. Laboratory Methods in Dairy Microbiology. London: Academic Press, 1976. 15. Thatcher FS, Clark DS. Microorganisms in foods. Vol I. Their significance and methods of enumeration. Toronto: University of Toronto Press, 1968. Alim Falkow S. Detection of
ARMA, So M, Samadpour-Motalebi M, enterotoxigenic. Escherichia coli by DNA colony hybridisation. J Infect Dis 1980; 142: 892-95. 17. Grunstein M, Hogness DS. Colony hybridisation: a method for the isolation of cloned DNAs that contain specific gene. Proc Natl Acad Sci 16.
Moseley SL, Hug I,
USA; 72: 3961-65. 18. Mason PJ, Williams JG. Hybridisation in the analysis of recombinant DNA. In: Hames BD, Higgins SJ, eds. Nucleic add hybridisation: a practical approach. Oxford: IRL Press, 1985: 113-25. 19. Gennaro ML, Greenway PJ, Broadbent DA. The expression of biologically active cholera toxin in Escherichia coli. Nucleic Acids Res 1982; 10: 4883-90. 20. Lathe R, Hirth P, De Wilde M, Lecog JP. Cell free synthesis of enterotoxin of Esherichia coli from a cloned gene. Nature 1980; 284: 473-74. 21. De Wilde M, Ysebert M, Hartford M. DNA sequence of STa2 enterotoxin gene from an Escherichia coli strain of human origin. In:
Levey SB, Clowes RC, Koenig EK, eds. Molecular biology, pathogenicity and ecology of bacterial plasmids. New York: Plenum Publishing Corporation, 1981: 596. 22. Feinberg AP, Vogelstein B. A technique for radiolabelling DNA restriction endonuclease fragments to high specific activity. Anal Biochem 1983; 132: 6-13. 23.
Banigo EOI, Muller HG. Carboxylic acid patterns J Sci Food Agric 1972; 23: 101-11.
in
ogi fermentation.
24. Andah A, Muller HG. Studies on koko, a Ghanaian fermented maize porridge. Ghana J Agric Sci 1973; 6: 103-08. 25. Akinrele IA. Fermentation studies on maize during the preparation of traditional African starch-cake food. J Sci Food Agric 1970; 21: 619-25. 26. Muller HG. Traditional cereal processing in Nigeria and Ghana. Ghana J Agric Sci 1970; 3: 187-95.
Transient release of lipid peroxides after coronary artery balloon angioplasty
Free radical
production may cause myocardial damage during reperfusion of ischaemic myocardial tissue; when free radicals interact with polyunsaturated fatty acids or their esters, lipid peroxides are produced. A product of lipid peroxidation, malondialdehyde, was measured in 10 subjects with stable angina who underwent angioplasty of a proximal high-grade stenosis (over 90%) of the left anterior descending coronary artery. In all subjects the duration of balloon occlusion was 60 s. Blood was withdrawn from the great cardiac vein immediately before balloon inflation (T0), immediately after balloon deflation (T60), 15 s after balloon deflation (T75), and 1 min after balloon deflation (T120). There was a significant increase in malondialdehyde at T60 compared with T0 for the first balloon inflation (mean increase 0·3 µmol/l [95% confidence limits 0·1, 0·5]), and at both T60 (0·31 µmol/l [0·15, 0·47]) and T75 (0·22 µmol/l [0·04. 0·40]) for the second balloon inflation. This model could be used to assess antioxidant effects of drugs.
Introduction
Oxygen-derived free radicals are produced after reperfusion of ischaemic myocardium in animals. Radicals have been directly detected by electron spin resonance,2 and freeradical damage products have also been measured.3 Myocardial reperfusion is associated with several potentially harmful effects, including transient impairment of myocardial function (myocardial "stunning"),4 an increased incidence of dysrhythmias,55 and possibly increased myocardial necrosis.6 These adverse effects can be reduced by treatment with various antioxidants at the time of myocardial reperfusion,’,8 which indicates that free-radical formation is an important factor in their pathophysiology.
ADDRESSES: Regional Medical Cardiology Centre, Royal Victoria Hospital, Belfast (M. J. D. Roberts, MRCP, T. G. Trouton, MD, M. M. Khan, FRCP, S. W. Webb, MD, C M. Wilson, MD, G. C. Patterson, MD, A. A. J. Adgey, MD); and Department of Chemical Pathology, Queen’s University, Belfast, UK (I. S. Young, MRCP, E. R. Trimble, MD). Correspondence to Dr A. A. J. Adgey, Regional Medical Cardiology Centre, Royal Victoria Hospital, Belfast BT12 6BA, UK.
144
But how far can the results of these animal experiments be extrapolated to man? In particular, most animal studies have used a period of complete coronary artery occlusion of sudden onset, followed by equally sudden myocardial reperfusion; this sequence would clearly be unusual in man, for whom coronary artery occlusion may be incomplete, and myocardial reperfusion gradual. Thus it is important to look
After the second balloon inflation at T7s the result in 1 patient was grossly aberrant (malondialdehyde concentration more than doubled after angioplasty) and was excluded from the analysis. The differences between the observations at each time-point were compared and 95% confidence limits calculated. Such differences between patients are statistically independent. Sequential balloon inflations were compared separately.
for evidence of free-radical formation in man under conditions of controlled transient coronary artery occlusion and myocardial reperfusion; such conditions are met by subjects who undergo percutaneous transluminal coronary angioplasty (PTCA) for stable angina. Free-radical species are very short-lived and can only be directly measured by electron spin resonance-a technique difficult to apply in man. However, when free radicals are produced they react with various molecules to form products which can be measured as an indirect indicator of free-radical activity. Lipid peroxides are produced when free radicals interact with polyunsaturated fatty acids or their esters. As an indicator of lipid peroxidation we have measured malondialdehyde, a product of free-radical attack on polyunsaturated fatty acids, in a group of patients who underwent PTCA. Serial samples of blood from the great cardiac vein were used to represent venous blood draining directly from ischaemic myocardial tissue as it undergoes
Results
reperfusion.
Mean malondialdehyde concentrations in arterial (2-15
pmol/1) and venous (2-03 Nmol/1) control samples were similar to values previously reported by other workers.10 There was no significant difference between initial (2-15 Ilffioljl) and final (1 -96 trnol/1) arterial or great-cardiac-vein malondialdehyde concentrations (2-03 and 2-02 J.Ul101/1, respectively). There was no significant arteriovenous gradient for malondialdehyde in pre-inflation or postinflation samples. Results for first and second balloon inflations are recorded in the table. Mean malondialdehyde concentrations were significantly raised at T 60 compared with To for the first balloon inflation (0-3 umol/1 [95% confidence limits 0-1, 05]), and were significantly raised in comparison with To at both T60 (0-31 Ilffioljl [015, 0-47]) and T7s (0-22 umol/1 [004, 0-4 for the second balloon inflation. By T 120 malondialdehyde concentrations had returned to baseline levels. Discussion
Patients and methods patients (aged 26-72 years) with stable angina who were to undergo angioplasty of a single stenosis (over 90%) of the left anterior descending coronary artery were recruited. Subjects with left bundle-branch block, valvular heart disease, or diabetes mellitus were excluded. Conventional angioplasty protocol was followed in all cases. Patients received 10 mg intravenous diazepam emulsion before the procedure, a heparin bolus of 10 000 units was given intravenously, and an infusion of dextran 40 was commenced at a 10
of 20 drops per min. In addition to arterial cannulation for balloon angioplasty, a 7 French multipurpose catheter was introduced via the femoral or brachial vein into the great cardiac vein. After a stabilisation period of 5 min, baseline blood samples were taken from the arterial and venous catheters and further control samples were taken 2 min later. Subsequent venous blood samples were taken immediately before balloon inflation (T), immediately after balloon deflation (T 60; the occlusion time was 60 s in all subjects); 15 s after balloon deflation (T7s); and 60 s after balloon deflation (T 120)’ This sequence of sampling was repeated for up to 3 subsequent inflations. Further arterial and venous samples were taken 5 min after completion of angioplasty. The protocol was approved by the ethical committee of the Queen’s University, Belfast, and patients gave full informed consent. Blood samples were taken into dipotassium ethylenediaminotetraacetate and stored on ice before separation of plasma at 2000 gat 4°C. Malondialdehyde content was measured by fluorometric assay of the adduct formed with thiobarbituric acid as described by Yagi.99 The intra-assay coefficient of variation was 5-5% and the interassay coefficient of variation 70% at a malondialdehyde concentration of rate
1-76
Eunol/1.
INCREASE IN GREAT-CARDIAC-VEIN MALONDIALDEHYDE (MDA) CONCENTRATION AFTER FIRST AND SECOND PTCA BALLOON INFLATIONS
Malondialdehyde is a measure of lipid peroxidation, which when polyunsaturated fatty acids are attacked by free
occurs
radicals
of
Raised concentrations of oxygen. have been observed in malondialdehyde reperfused myocardial tissue in animals,3and in man malondialdehyde concentrations are raised in several conditions where free-radical damage has been strongly implicated (eg, rheumatoid
arthritis" and diabetes mellitus’2). been measured by high-performance Malondialdehyde has liquid chromatography,l° and spectrophotometric13 or fluorometric9 measurement of its thiobarbituric acid adduct. Some interference has been observed in the spectrophotometric and fluorometric assaysl4 but, in general, malondialdehyde correlates well with other markers of free-radical activity.1s As we observed consistent changes in serial measurements taken over a short period of time, we do not believe that interference in the assay is likely to have been a significant problem in our subjects. In man, raised malondialdehyde concentrations have been reported in peripheral venous blood in patients in whom successful myocardial reperfusion was achieved after thrombolytic therapy for acute myocardial infarction in comparison with subjects in whom reperfusion did not occur.16 During PTCA we found a transient rise in malondialdehyde after balloon deflation in blood from the great cardiac vein, which returned to baseline concentrations within 1 min. This observation is consistent with the concept that a burst of free-radical production occurs during the reperfusion which follows the brief ischaemia of balloon inflation. It is also possible that free-radical production is secondary to cellular damage that occurs during the ischaemia-but such damage seems unlikely during this short period of ischaemia. Lipid peroxides are also known to be present in atherosclerotic plaque tissue17 and we cannot exclude the possibility that the rises in malondialdehyde concentration are due to mechanical disruption of plaque during balloon inflation. In addition, malondialdehyde is a potentially harmful
145
substance that may damage extracellular components,l8 which may themselves be cytotoxic to endothelial cells.19 Whatever the cause of the transiently raised malondialdehyde concentration, we believe that angioplasty to a high-grade proximal coronary artery stenosis, with subsequent release of the balloon, represents a setting in which the therapeutic usefulness of pharmacological agents as free-radical scavengers can be investigated in man. If such agents reduce the concentrations of biochemical markers of free-radical damage during coronary artery angioplasty, further support will be provided for trials of adjuvant antioxidant therapy with thrombolytic agents in acute myocardial infarction. REFERENCES 1. Dart RC, Sanders AB. Oxygen free radicals and myocardial reperfusion injury. Ann Emerg Med 1988; 17: 53-58. 2. Bolli R, Patel BS, Jeroudi MO, Lai EK, McCay PB. Demonstration of free radical generation in "stunned" myocardium of intact dogs with the use of the spin trap &agr;-phenyl N-tert-butyl nitrone. J Clin Invest 1988; 82: 476-85. 3. Guarnieri C, Flamigni F, Caldarera CM. Role of oxygen in the cellular damage induced by re-oxygenation of hypoxic heart. J Mol Cell Cardiol 1980; 12: 797-808. 4. Bolli R, Jeroudi MO, Patel BS, et al. Marked reduction of free radical generation and contractile dysfunction by antioxidant therapy begun at the time of reperfusion. Circ Res 1989; 65: 607-22. 5. Woodward B, Zakaria MNM. Effect of some free radical scavengers on reperfusion induced arrhythmias in the isolated rat heart. J Mol Cell
Cardiol 1985; 17: 485-93. 6. Ambrosio G, Becker LC, Hutchins GM, Weisman HF, Weisfeldt ML. Reduction in experimental infarct size by recombinant human superoxide dismutase: insights into the pathophysiology of reperfusion injury. Circulation 1986; 74: 1424-33.
7. Opie LH. Reperfusion injury and its pharmacologic modification. Circulation 1989; 80: 1049-62.
Myers ML, Bolli R, Lekich RF, Hartley CJ, Roberts R. Enhancement of recovery of myocardial function by oxygen free-radical scavengers after reversible regional ischaemia. Circulation 1985; 72: 915-21. 9. Yagi K. Assay for blood plasma or serum. Methods Enzymol 1984; 105: 8.
328-31.
10.
Wong SHY, Knight JA, Hopfer SM, Zahana O, Leach CN, Sunderman FW. Lipoperoxides in plasma as measured by liquid-chromatographic separation of malondialdehyde-thiobarbituric acid adduct. Clin Chem 1987; 33: 214-20. CR, Jackson PG, Highton J, van Rij AM. Lipid peroxidation and
11. Wade
in the synovial fluid and plasma of patients with rheumatoid arthritis. Clin Chim Acta 1987; 164: 245-50. 12. Sato Y, Hotta N, Sakamoto N, Matsuoka S, Ohishi N, Yagi K. Lipid peroxide level in plasma of diabetic patients. Biochem Med 1979; 21: 104-07. 13. Bird RP, Draper HH. Comparative studies on different methods of malondialdehyde determination. Methods Enzymol 1984; 105: 299305. 14. Knight JA, Pieper RK, McClellan L. Specificity of the thiobarbituric acid reaction: its use in studies of lipid peroxidation. Clin Chem 1988; 34: 2433-38. 15. Slater TF. Overview of methods used for detecting lipid peroxidation. Methods Enzymol 1984; 105: 283-93. 16. Davies SW, Ranjadayalan K, Wickens DG, Dormandy TL, Timmis AD. Free radicals, reperfusion and thrombolysis. Eur Heart J 1989; 10
malondialdehyde
17.
(suppl): 274. Ledwozyw A, Michalak J, Stepien A, Kadziolka A. The relationship between plasma triglycerides, cholesterol, total lipids and lipid peroxidation products during human atherosclerosis. Clin Chim Acta 1986; 155: 275-84.
18.
Fogelman AM, Shechter I, Seager J, Hokom M, Child JS, Edwards PA. Malondialdehyde alteration of low density lipoproteins leads to cholesteryl ester accumulation in human monocyte-macrophages. Proc Natl Acad Sci USA 1980; 77: 2214-18. SA, Galdal KS, Nilsen E. LDL-induced cytotoxicity and its inhibition by anti-oxidant treatment in cultured human endothelial cells and fibroblasts. Atherosclerosis 1983; 49: 23-30.
19. Evensen
BOOKSHELF Living Medicine Peter Richards. Cambridge: Cambridge University Press. 1990. Pp 187. 25,$39 hardback; /;8.95,$14.95 paperback. ISBN 0-521384788 and 0-521386284.
Living Medicine
has been written
partly to supplement Professor Richards’ already popular Learning Medicine. Whereas the earlier book is directed particularly to 17 and 18 year-olds considering a career in medicine, Living Medicine is addressed to students and house-officers who are planning their future careers. Written with clarity and a wry sense of humour, the book is short enough to be read in an evening. Its stated objective is to prepare senior students and house officers for their responsibilities and opportunities as doctors. Is this achieved? My house officer’s answer was guarded-yes, the book is entertaining, but too superficial in its kaleidoscopic survey of the many careers open to medical graduates. It alerts readers to the options, but little more. For a career that "dominates life" a fuller treatment would be preferable, if the book is to act as an important reference for students and new graduates. Nevertheless, there is a great deal to enjoy here. The pen portraits of different branches of medicine are miniature masterpieces. Not everyone will agree with Richards’ perception of their specialty-some even come across as decidedly grim. There is no attempt, for example, to disguise the arduous training necessary for general surgery. Running through the text is a lively appreciation of the source
conflicting demands of marriage, child-rearing, unsociable hours, hobbies, and life outside medicine. It is hard to discern whether the author advocates total dedication of one’s entire life to medicine or whether there is a covert admiration of those who do other things as well (as one of his "handful of doctors with a diploma in organ playing", I can vouch that a double act brings its own problems in plenty). As well as describing numerous possible careers, Richards writes with authority and wisdom about professional freedom and responsibility, quoting the Hippocratic oath and other key principles of conduct, confidentiality, and honesty. Further information on accreditation, General Medical Council regulations, and study leave is contained in appendices. Overall, medicine is presented as a hard task-master, but infinitely rewarding. There is little criticism of the present structure, although specialty overcrowding and over-long hours are acknowledged. The comment "appointment committees do not on the whole look imaginatively on missionary spirit or humanitarian concern" is unfortunately true, but it is sad that work abroad is seen as little more than an interruption to
diploma gathering. Wittily illustrated by
David Langdon, this enjoyable book can be recommended to a readership far wider than the students and house officers at which it is aimed. Southampton General Hospital, Southampton S09 4XY, UK
RICHARD GODFREY
