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STARCH EMBOLI IN TRANSPLANTED KIDNEYS
1009 his significant X2 also to satisfy the second condition; the other half would have the maximum and minimum periods adjacent. Therefore, rejecting these, the p values return to those normally applicable to a 2 x 2 X2 test. Naturally, we agree that further investigation of this important question is highly desirable, and would not claim that our results are any more " conclusive " than most other pieces of research. We have learnt from Medawar and Popper that research proceeds from hypothesis to experiment and that experimental results can prove a hypothesis to be wrong but never prove it to be right. However, statistics properly used provide- some idea of the likelihood that a hypothesis may be correct. We believe our statistical analysis to be entirely appropriate to the testing of our particular hypothesis. This analysis shows it to be unlikely (though not impossible) that the seasonal variation in osteomalacia which we have reported is due to chance. M.R.C. Mineral Metabolism Unit, General Infirmary, Great George Street, Leeds 1.
INTRAUTERINE
D. H. MARSHALL B. E. C. NORDIN.
STARCH EMBOLI
IN TRANSPLANTED KIDNEYS
found in 63%of gloin two children whose allografted kidneys transplants came from a single donor. A third child had starch emboli in 35% of the glomeruli of his transplanted kidney. The grafts had been preserved in a cold, pulsatile perfusing machine, the Belzer T1450, before transplantation. The first two kidneys had been manipulated within the casette of the Belzer T1450 because of leakage. Within 90 minutes of transplantation open kidney biopsy was performed on all three grafts. The glomerular capillaries and some afferent arterioles contained corn-starch granules, 10-25 µ in diameter, identical to those found on surgical gloves. Electron microscopy revealed that neutrophilic granulocytes and platelet thrombi were closely associated with the starch granules (see accompanying figure). Glomerular capillaries without starch contained platelet thrombi with partial degranulation of some platelets. Granulocytes were not present in peritubular capillaries. The starch emboli persisted in the kidneys: 24% of glomeruli from the infarcted kidney of patient 1 contained starch emboli at six days; 12% of glomeruli from needle biopsy specimen of patient 2 at 22 days, and 1 % from his
SIR,—Corn-starch emboli
were
meruli of
DEVICE, PROSTAGLANDINS, AND COPPER
SIR,—In 1971 Chaudhuri1 proposed a hypothesis to explain the mechanism of action of the intrauterine device (l.u.D.). He suggested that the I.U.D., by virtue of a traumatic action on the endometrium, resulted in the local release of prostaglandins and that it was these substances which prevented conception. Since that time studies in several animal species have provided evidence to support this.2,3 In view of the relatively small surface area of the coppercontaining I.U.D.s now in use, Chaudhuri’s hypothesis would appear at first sight to be untenable as an explanation of the mechanism of action of this l.U.D. type. However, recent studies on the influence of copper ions on prostaglandin biosynthesis provide possibilities for an extension of the above hypothesis to support a role for prostaglandins in the contraceptive effect of copper-containing l.U.D.S. Maddox,4confirming earlier findings,5 has shown that copper ions can shift prostaglandin biosynthesis from prostaglandin E2 (P.G.E2) to P.G.F2. production. Hence the presence of a copper-containing l.U.D. in the uterus could be expected to stimulate local P.G.F2a production (with a simultaneous decrease in P.G.E2 production). p.G.F211 now seems to be established as the luteolytic hormone in the sheep, and evidence exists to support a similar role in other subprimate species. Recent findings in man implicate this prostaglandin in human luteolysis.6 The above effects of copper ions and the recent isolation of an enzyme capable of the reduction of P.G.E2 to p.G.F2u? indicate to us that research into the mechanism of action of copper-containing i.u.D.s should be directed towards a search for the presence of a similar enzyme in human endometrium. Studies on the effects of copper ions on endometrial P.G.-synthetase and any possible P.G.EP.G.F-converting enzyme may prove to be of particular relevance in providing a biochemical explanation for the mechanism of action of the copper-containing l.U.D.S. St
George’s Hospital Medical School, Hyde Park Corner, London SW1.
J. V. H. FORDER.
1. Chaudhuri, G. Lancet, 1971, i, 480. 2. Chaudhuri, G. Prostaglandins, 1973, 3, 773. 3. Saksena, S. K., Lau, I. F., Castracane, V. O. ibid. 1974, 5, 97. 4. Maddox, I. S. Biochim. biophys. Acta, 1973, 306, 74. 5. Lee, R. E., Lands, W. E. M. ibid. 1972, 260, 203. 6. Powell, W. S., Hammarström, S., Samuelson, B., Sjöberg, B. 7.
Lancet, 1974, i, 1120. Hensby, C. N. Biochim. biophys. Acta, 1974, 348, 145.
Electron micrograph montage of glomerular capillary loops shows starch (s) in lower loop and platelet thrombus distending the upper loop. x 2750.
To the left of the starch granule, which is folded and distorted because it could not be fixed by glutaraldehyde, is a polymorphonuclear leucocyte (g). Many platelets (p) contain granules, while others are degranulated. Arrows point to glomerular basement membrane. u-urinary space. b-Bowman’s capsule.
1010
nephrectomy at seven weeks had starch emboli. At 22 days the starch granules were completely engulfed by large mononuclear cells. The first kidney infarcted because of technical error. The second kidney functioned immediately, had adequate blood-flow by xenon washout and angiography, developed anuria at 2t days, and later had a modest return of urinary output. Nephrectomy was performed because of rejection and recurrence of initial disease, oxalate nephritis. Direct cross-match of donor leucocytes with recipient’s serum was negative. Hyperacute rejection was unlikely on the basis of clinical course and laboratory findings. In-vitro incubation at 37 °C of 0-45 ml. citrated human platelet-rich plasma (P.R.P.) with 0-05 ml. of starch (1 mg. per ml.) in 0-15M sodium chloride showed no platelet aggregation at various times up to 60 minutes. The
reconstituted plasma clotting-time, obtained by adding calcium chloride to the mixture of P.R.P. and starch, was shortened 20-45% indicating activation of either platelets system, or both. orCorn-starch the clotting emboli in the glomerular capillaries of renal transplants have been previously recorded by Min et a1. Active phagocytosis of the starch by granulocytes and much later by macrophages was found. Neither phagocytosis of the starch nor the associated platelet thrombi were noted by Min et a1. or by Lamb and Roberts2 in an experimental study in rats. We found granulocytes closely associated with starch emboli within ten minutes following intra-aortic injection into rabbits. In-vitro tests substantiated our observation that the starch granules could be involved with the formation of platelet thrombi - but the mechanism remains unexplained and is still being investigated. The starch probably contaminated the recirculating corn perfusate during manipulations performed on the kidneys in the cold perfusing apparatus and could probably be prevented either by thorough washing and drying of surgical gloves, or by placing a final filter in the perfusion apparatus. Temple University Health I. B. ELFENBEIN Sciences Center, R. F. MCALACK 3400 North Broad Street, D. C. B. MILLS Philadelphia, Pennsylvania 19140, U.S.A., and E. MUNOZ-GHANNAM Albert Einstein Medical Center, Northern Division, Philadelphia, Pennsylvania, U.S.A
J. E.
a
defined or
closed institutions. Department of Epidemiology
and
Preventive Medicine, University of Glasgow,
Glasgow.
GORDON T. STEWART.
MILDER
SIR,—In his comment on the paper by Professor Holland and Professor Whitehead (Aug. 17, p. 391), Dr Lehman (Sept. 14, p. 647) draws attention to the limitation of value imposed by the prevalence of the disease in question. There are further limitations which should be noted, especially when tests are used in screening programmes. If the prevalence of disease-(a+c)/N in Holland and Whitehead’s notation-is 10% with specificity and sensitivity of the test each as high as 90%, as in the favourable instance cited by Lehman, there will in the first 1000 persons tested be a larger number (100) of false results than of true positives (90). As the prevalence falls, the proportion of false results rises sharply. Thus, at 1% prevalence, there will be 9 true positives compared with 100 that are false in 1000 persons tested. It is extremely difficult to make allowance for this when incidence varies, as in prospective surveys, except by imposing the general condition that if the result of a test is going to be more often true than false, the ratio of false to true positives must be less than (100 -P)/
2.
cutting score (e.g., measurement of bloodpressure intelligence). They are of special importance where decisions are influenced by the results, as in certain surgical interventions, treatment of symptomless conditions, various prophylactic procedures, and referral of patients to with
M. COTÉ.
PREDICTIVE VALUE OF LABORATORY TESTS
1.
P where P is the best estimate of the true prevalence at the time of testing-i.e., N/(a+c)-1in Holland and Whitehead’s example. In the absence of experimental validation of any given test, the argument used by Holland and Whitehead must be restricted to situations where the sensitivity a x 100/(a-c) of the test is governed by intrinsic biological factors. If there are additional variables of selective bias (e.g., selfselection by patients in screening programmes or caseselection by doctors) or technical errors (seldom less than 5%), the predictive value of the test is subject to further, more complex, restrictions which may or may not be eased by repetitions in search of accuracy. Holland and Whitehead acknowledge the need to check " accuracy by the most definitive investigation available ", but, in this respect, accuracy depends upon the availability of independent criteria. They are semantically correct in " saying that discrimination is based upon sensitivity and specificity values ", but in practice these values are usually ascertained by results obtained in populations already tested, whereas accuracy depends upon the incidence of disease in that about to be tested as well as upon the other variables mentioned above. If incidence falls rapidly, as in some disease-control programmes, tests with high specificity may become uneconomic and impracticable for logistic as well as epidemiological reasons. These considerations apply not only to new laboratory tests. They apply equally to the predictive value of changes in the ST segment in electrocardiography, as is well shown in the comment by Dr Wilson and others (Sept. 14, p. 665) on the paper by Dr Morris and his colleagues (Aug. 17, p. 372). They apply in general to any test or procedure
Min, K-W., Jackson, J. Y., Gyorkey, F., Bergeron, J., MartinezMaldonado, M. Kidney International, 1972, 2, 291. Lamb, D., Roberts, G. J. clin. Path. 1972, 25, 876.
ESTIMATION OF BLOODCARBOXYHÆMOGLOBIN
SIR,—Estimation of the blood-level of carboxyhxmoglobin (COHb) as an index of the subject’s degree of inhalation of tobacco smoke is
now
a
well-established
technique. It was employed successfully by Dr Turner and his colleagues (Sept. 28, p. 737). Unfortunately, the estimations were carried out by means of the I.L. CO Oximeter, model 182 (Instrumentation Laboratories Inc.). In this laboratory we have had the opportunity to examine many examples of this instrument and we have reached the following conclusions: (1) The instrument is inherently unstable, particularly when measuring " CO ", and tends to lose calibration despite frequent use of the alleged " zeroing solution ". This instability has the greatest effect on readings at the lower end of the scale. (2) Variations in the oxygen saturation of the blood-sample may affect the " CO " reading. This defect is often not immediately obvious but can be responsible for errors of up to 6% COHb.
Although we still employ the instrument for multiple analyses of COHb, we now believe that accurate readings can only be obtained by: (a) frequent calibration at at least two points with blood-samples analysed for COHb by an alternative method of greater accuracy (such as that of Commins and Lawther 1); (b) reduction of all hxmo1.
Commins,
B.
T., Lawther, P. J. Br. J. Industr. Med. 1965, 22, 139.
INTRAUTERINE
D. H. MARSHALL B. E. C. NORDIN.
STARCH EMBOLI
IN TRANSPLANTED KIDNEYS
found in 63%of gloin two children whose allografted kidneys transplants came from a single donor. A third child had starch emboli in 35% of the glomeruli of his transplanted kidney. The grafts had been preserved in a cold, pulsatile perfusing machine, the Belzer T1450, before transplantation. The first two kidneys had been manipulated within the casette of the Belzer T1450 because of leakage. Within 90 minutes of transplantation open kidney biopsy was performed on all three grafts. The glomerular capillaries and some afferent arterioles contained corn-starch granules, 10-25 µ in diameter, identical to those found on surgical gloves. Electron microscopy revealed that neutrophilic granulocytes and platelet thrombi were closely associated with the starch granules (see accompanying figure). Glomerular capillaries without starch contained platelet thrombi with partial degranulation of some platelets. Granulocytes were not present in peritubular capillaries. The starch emboli persisted in the kidneys: 24% of glomeruli from the infarcted kidney of patient 1 contained starch emboli at six days; 12% of glomeruli from needle biopsy specimen of patient 2 at 22 days, and 1 % from his
SIR,—Corn-starch emboli
were
meruli of
DEVICE, PROSTAGLANDINS, AND COPPER
SIR,—In 1971 Chaudhuri1 proposed a hypothesis to explain the mechanism of action of the intrauterine device (l.u.D.). He suggested that the I.U.D., by virtue of a traumatic action on the endometrium, resulted in the local release of prostaglandins and that it was these substances which prevented conception. Since that time studies in several animal species have provided evidence to support this.2,3 In view of the relatively small surface area of the coppercontaining I.U.D.s now in use, Chaudhuri’s hypothesis would appear at first sight to be untenable as an explanation of the mechanism of action of this l.U.D. type. However, recent studies on the influence of copper ions on prostaglandin biosynthesis provide possibilities for an extension of the above hypothesis to support a role for prostaglandins in the contraceptive effect of copper-containing l.U.D.S. Maddox,4confirming earlier findings,5 has shown that copper ions can shift prostaglandin biosynthesis from prostaglandin E2 (P.G.E2) to P.G.F2. production. Hence the presence of a copper-containing l.U.D. in the uterus could be expected to stimulate local P.G.F2a production (with a simultaneous decrease in P.G.E2 production). p.G.F211 now seems to be established as the luteolytic hormone in the sheep, and evidence exists to support a similar role in other subprimate species. Recent findings in man implicate this prostaglandin in human luteolysis.6 The above effects of copper ions and the recent isolation of an enzyme capable of the reduction of P.G.E2 to p.G.F2u? indicate to us that research into the mechanism of action of copper-containing i.u.D.s should be directed towards a search for the presence of a similar enzyme in human endometrium. Studies on the effects of copper ions on endometrial P.G.-synthetase and any possible P.G.EP.G.F-converting enzyme may prove to be of particular relevance in providing a biochemical explanation for the mechanism of action of the copper-containing l.U.D.S. St
George’s Hospital Medical School, Hyde Park Corner, London SW1.
J. V. H. FORDER.
1. Chaudhuri, G. Lancet, 1971, i, 480. 2. Chaudhuri, G. Prostaglandins, 1973, 3, 773. 3. Saksena, S. K., Lau, I. F., Castracane, V. O. ibid. 1974, 5, 97. 4. Maddox, I. S. Biochim. biophys. Acta, 1973, 306, 74. 5. Lee, R. E., Lands, W. E. M. ibid. 1972, 260, 203. 6. Powell, W. S., Hammarström, S., Samuelson, B., Sjöberg, B. 7.
Lancet, 1974, i, 1120. Hensby, C. N. Biochim. biophys. Acta, 1974, 348, 145.
Electron micrograph montage of glomerular capillary loops shows starch (s) in lower loop and platelet thrombus distending the upper loop. x 2750.
To the left of the starch granule, which is folded and distorted because it could not be fixed by glutaraldehyde, is a polymorphonuclear leucocyte (g). Many platelets (p) contain granules, while others are degranulated. Arrows point to glomerular basement membrane. u-urinary space. b-Bowman’s capsule.
1010
nephrectomy at seven weeks had starch emboli. At 22 days the starch granules were completely engulfed by large mononuclear cells. The first kidney infarcted because of technical error. The second kidney functioned immediately, had adequate blood-flow by xenon washout and angiography, developed anuria at 2t days, and later had a modest return of urinary output. Nephrectomy was performed because of rejection and recurrence of initial disease, oxalate nephritis. Direct cross-match of donor leucocytes with recipient’s serum was negative. Hyperacute rejection was unlikely on the basis of clinical course and laboratory findings. In-vitro incubation at 37 °C of 0-45 ml. citrated human platelet-rich plasma (P.R.P.) with 0-05 ml. of starch (1 mg. per ml.) in 0-15M sodium chloride showed no platelet aggregation at various times up to 60 minutes. The
reconstituted plasma clotting-time, obtained by adding calcium chloride to the mixture of P.R.P. and starch, was shortened 20-45% indicating activation of either platelets system, or both. orCorn-starch the clotting emboli in the glomerular capillaries of renal transplants have been previously recorded by Min et a1. Active phagocytosis of the starch by granulocytes and much later by macrophages was found. Neither phagocytosis of the starch nor the associated platelet thrombi were noted by Min et a1. or by Lamb and Roberts2 in an experimental study in rats. We found granulocytes closely associated with starch emboli within ten minutes following intra-aortic injection into rabbits. In-vitro tests substantiated our observation that the starch granules could be involved with the formation of platelet thrombi - but the mechanism remains unexplained and is still being investigated. The starch probably contaminated the recirculating corn perfusate during manipulations performed on the kidneys in the cold perfusing apparatus and could probably be prevented either by thorough washing and drying of surgical gloves, or by placing a final filter in the perfusion apparatus. Temple University Health I. B. ELFENBEIN Sciences Center, R. F. MCALACK 3400 North Broad Street, D. C. B. MILLS Philadelphia, Pennsylvania 19140, U.S.A., and E. MUNOZ-GHANNAM Albert Einstein Medical Center, Northern Division, Philadelphia, Pennsylvania, U.S.A
J. E.
a
defined or
closed institutions. Department of Epidemiology
and
Preventive Medicine, University of Glasgow,
Glasgow.
GORDON T. STEWART.
MILDER
SIR,—In his comment on the paper by Professor Holland and Professor Whitehead (Aug. 17, p. 391), Dr Lehman (Sept. 14, p. 647) draws attention to the limitation of value imposed by the prevalence of the disease in question. There are further limitations which should be noted, especially when tests are used in screening programmes. If the prevalence of disease-(a+c)/N in Holland and Whitehead’s notation-is 10% with specificity and sensitivity of the test each as high as 90%, as in the favourable instance cited by Lehman, there will in the first 1000 persons tested be a larger number (100) of false results than of true positives (90). As the prevalence falls, the proportion of false results rises sharply. Thus, at 1% prevalence, there will be 9 true positives compared with 100 that are false in 1000 persons tested. It is extremely difficult to make allowance for this when incidence varies, as in prospective surveys, except by imposing the general condition that if the result of a test is going to be more often true than false, the ratio of false to true positives must be less than (100 -P)/
2.
cutting score (e.g., measurement of bloodpressure intelligence). They are of special importance where decisions are influenced by the results, as in certain surgical interventions, treatment of symptomless conditions, various prophylactic procedures, and referral of patients to with
M. COTÉ.
PREDICTIVE VALUE OF LABORATORY TESTS
1.
P where P is the best estimate of the true prevalence at the time of testing-i.e., N/(a+c)-1in Holland and Whitehead’s example. In the absence of experimental validation of any given test, the argument used by Holland and Whitehead must be restricted to situations where the sensitivity a x 100/(a-c) of the test is governed by intrinsic biological factors. If there are additional variables of selective bias (e.g., selfselection by patients in screening programmes or caseselection by doctors) or technical errors (seldom less than 5%), the predictive value of the test is subject to further, more complex, restrictions which may or may not be eased by repetitions in search of accuracy. Holland and Whitehead acknowledge the need to check " accuracy by the most definitive investigation available ", but, in this respect, accuracy depends upon the availability of independent criteria. They are semantically correct in " saying that discrimination is based upon sensitivity and specificity values ", but in practice these values are usually ascertained by results obtained in populations already tested, whereas accuracy depends upon the incidence of disease in that about to be tested as well as upon the other variables mentioned above. If incidence falls rapidly, as in some disease-control programmes, tests with high specificity may become uneconomic and impracticable for logistic as well as epidemiological reasons. These considerations apply not only to new laboratory tests. They apply equally to the predictive value of changes in the ST segment in electrocardiography, as is well shown in the comment by Dr Wilson and others (Sept. 14, p. 665) on the paper by Dr Morris and his colleagues (Aug. 17, p. 372). They apply in general to any test or procedure
Min, K-W., Jackson, J. Y., Gyorkey, F., Bergeron, J., MartinezMaldonado, M. Kidney International, 1972, 2, 291. Lamb, D., Roberts, G. J. clin. Path. 1972, 25, 876.
ESTIMATION OF BLOODCARBOXYHÆMOGLOBIN
SIR,—Estimation of the blood-level of carboxyhxmoglobin (COHb) as an index of the subject’s degree of inhalation of tobacco smoke is
now
a
well-established
technique. It was employed successfully by Dr Turner and his colleagues (Sept. 28, p. 737). Unfortunately, the estimations were carried out by means of the I.L. CO Oximeter, model 182 (Instrumentation Laboratories Inc.). In this laboratory we have had the opportunity to examine many examples of this instrument and we have reached the following conclusions: (1) The instrument is inherently unstable, particularly when measuring " CO ", and tends to lose calibration despite frequent use of the alleged " zeroing solution ". This instability has the greatest effect on readings at the lower end of the scale. (2) Variations in the oxygen saturation of the blood-sample may affect the " CO " reading. This defect is often not immediately obvious but can be responsible for errors of up to 6% COHb.
Although we still employ the instrument for multiple analyses of COHb, we now believe that accurate readings can only be obtained by: (a) frequent calibration at at least two points with blood-samples analysed for COHb by an alternative method of greater accuracy (such as that of Commins and Lawther 1); (b) reduction of all hxmo1.
Commins,
B.
T., Lawther, P. J. Br. J. Industr. Med. 1965, 22, 139.