Serum angiotensin converting enzyme in sarcoidosisits value in present clinical practice

Transcription

1 Ann Clin Biochem 1989; 26: Review Article Serum angiotensin converting enzyme in sarcoidosisits value in present clinical practice P R STUOOY and R BIRD From the Harefield Hospital, Harefield, Middx UB9 6JH and "Endocrtne Laboratory, Department of Chemical Pathology, Archway Wing, Whittington Hospital, London NJ9 5NF, UK Angiotensin converting enzyme (ACE) has a central role in blood pressure SUMMARY. homeostasis. It is present in low and fairly constant concentration in the serum and in high concentration in the pulomonary capillary bed. Unusually high serum angiotensin converting enzyme (SACE) activity is present in active sarcoidosis, an observation now confirmed by many investigators. In spite of its lack of specificity as a test for sarcoidoisis, SACE provides a good monitor ofdisease activity which clinicians find useful in the management ofpatients with sarcoidosis. There continues to be considerable interest in SACE in sarcoidosis and with the recent development of simpler assays, more centres may be expected to offer SACE measurements as a service. In this paper we discuss the indications for estimating SACE in sarcoidosis and its relevance to current clinical practice. Angiotensin converting enzyme (ACE) is a halide-activated exopeptidase responsible for the conversion ofthe decapeptide angiotensin I to the octapeptide angiotensin II with the release of the dipeptide His-Leu from the carboxyl terminus.p It also inactivates bradykinin by a similar mechanisrn.>! ACE therefore has a central role in the control of blood pressure. The rapid conversion ofangiotensin I to angiotensin II was assumed to take place in the circulation until Ng and Vane" recognised that the enzyme activity present in plasma was insufficient to account for the speed of in vivo conversion, and demonstrated that most conversion occurred during a single passage through the lungs. ACE is a membrane-bound glycoprotein; labelled antibody studies on lung tissue localise its activity to the luminal surface of the pulmonary endothelium." The demonstration of satisfactory blood pressure maintenance during cardiopulmonary bypass indicates that ACE is present in non-pulmonary vascular beds," Subsequent This paper was prepared at the invitation of the Clinical Laboratory Investigations working party of the Scientific Committee of the Association of Clinical Biochemists, but does not necessarily reflect their views. Correspondence: Dr R Bird. studies demonstrated low or moderate ACE activity in the vascular beds ofmost organs. The highest levels are found in the lungs. Measurement of serum ACE (SACE) activity may give useful information for research in hypertension and in experimental lung injury. Hypoxic or chemical lung injury may result in transient alterations in SACE activity, presumably following damage to the pulmonaryvascular endothelium." In 1975 Lieberman, in a chance observation, found a possible clinical role for SACE. His investigations into the cause of low blood pressure in patients with chronic lung disease unexpectedly revealed high SACE activity in a small group of patients with sarcoidosis." This observation has since been confirmed by many authors.":" The raised SACE activity in sarcoidosis is due to activation ofthemonocyte macrophage system thatprovides the 'building blocks' from which the granulomas are constructed. Circulating blood monocytes, the precursors of the epithelioid cells found in the sarcoid granulomas, contain almost no ACE, but on culture demonstrate the capacity to secrete the enzyme." Immunofluorescence studies show both angiotensin II, a cleavage product of ACE catalysis, and ACE in the epithelioid cells of granulomas. IS T lymphocytes 13

2 14 Studdy and Bird modulate ACE synthesis in the monocyte in culture and may do so in the sarcoid granuloma, for ACE is most abundant in the peripheral part of the granuloma where T lymphocytes and epithelioid cells are in close contact." Methods for measuring SACE A number ofdifferent methods ofassaying SACE are available. Most investigators have used the substrate hippuryl-l-histidyl-l-ieucine either in a spectrophotometric procedure based on that of Cushman and Cheung!' or as the spectrofluorimetric method described by Friedland and Silverstein." Assays using radiolabelled substrates have been described by Ryan et al. 18 and Rohrbach." Ronc-Testani'" used furylacrylylphenylalanylglycylglycine (FAPGG) in a direct spectrophotometric assay, and Maguire and Price" used the same substrate and developed a continuous monitoring spectrophotometric method for a centrifugal analyser. Highly significant linear correlations between these methods have been reported, although quoted reference ranges differ widely." Commercial kits are available in the UK from Mast Diagnostics, Sigma Diagnostics and BCL. A radioassay kit is produced by Ventrex Laboratories Inc. This very wide range ofpossible assay methods allows any laboratory to measure SACE, but it is neither necessary nor desirable for every laboratory to provide a service for the measurement of SACE since the test result is never urgently required. The local demand for the test is likely to be low in the absence of a specific research or special clinical interest. Hospitals with only an occasional request should send specimens to a reference laboratory which uses an established method. Serum samples may be stored prior to assay. SACE is a relatively stable enzyme and we have found no loss of activity when serum samples have been stored for 20 days at 25 C, 1 month at 4 C or 6 months at - 20 C. Unfrozen specimens may be sent by post to the laboratory undertaking the test. The reagent cost per test using an 'in house' method is likely to be less than 0 20; for kit reagents the cost per patient test is much greater, of the order of 2-3 (1987 prices). SACE in normal healthy subjects SACE levels are surprisingly stable in normal healthy adults and show no significant diurnal and little longitudinal variation when repeat measurements are made." SACE activity is greater in actively growing children and the highest values are found in premature infants." Normal SACE levels may be suppressed by corticosteroid treatment. Gronhagen-Riska et al. 25 demonstrated a significant (P < 0'002) reduction of SACE activity in 11 normal healthy subjects within one week of commencing prednisolone 10 mg/day, A similar response was reported in patients suffering from cryptogenic fibrosing alveolitis and asthma." The mechanism by which normal SACE activity is suppressed by corticosteroids remains unexplained. 'Falsely low' values of SACE will be found in patients on ACE-inhibiting drugs such as captopril (Acepril, Capoten) or enalapril (lnnovace), which may be used in the long-term treatment of hypertension." SACE in sarcoidosis Sarcoidosis is one of a large family of chronic multisystem disorders in which the common feature is the presence ofepithelioid cell granulomas in affected organsand tissues. All partsofthe body can be affected, but it is the lung that is most frequently involved. When suspected, the diagnosis ofsarcoidosis is established by excluding other granulomatous diseases and demonstrating characteristic granulomas on biopsy of the affected tissues. Some combinations of clinical features (i.e. erythema nodosum and lymphadenopathy) and radiographic appearances (i.e. bilateral hilar gland enlargement and infiltration) are so typical as to justify a confident clinical diagnosis ofsarcoidosis provided there are no discordant features. Final confirmation can be obtained by a positive tissue biopsy or K veim-siltzbach test. In less characteristic cases biopsy from several sites may be required before the diagnosis can be accepted." A specific biochemical marker for sarcoidosis would assist the diagnosis, and the chance discovery of elevated SACE in the disease raised hopes that it would provide a simple diagnostic test. ACE is present in sarcoid epithelioid cells, but not macrophages or monocytes, and in significantly greater amounts than are found in other non-sarcoid granulomatous conditions. SACE levels broadly reflect the extent of the sarcoid granulomatous inflammation.p SACE AS A DIAGNOSTIC TEST Only some patients with sarcoidosis have raised SACE activity. In our recent series" SACE was measured by the Friedland and Silverstein spectrofluorimetric method in 134 normal control

3 Serum angiotensin converting enzyme in sarcoidosis 15 subjects and 230 patients with sarcoidosis. The mean ± I SD value for the control group was 34± 9 units/ml, and for the patients with sarcoidosis 79±44 units/ml., with 69% ofthe sarcoidosis patients having values above 52 units/ml, (control mean+2 SD). Making a correction for the non-gaussian distribution and comparing sarcoidosis with other granulomatous disorders including tuberculosis, primary biliary cirrhosis, Crohn's disease and leprosy and with lymphoma and lung fibrosis, the calculated positive and negative predictive values for SACE as a diagnostic test for sarcoidosis was 84% and 74%, respectively. Similar results were obtained in a large collaborative 12 centre study reported by one of us. 22 CXRI CXR J E... ~ c "z:.; 60 ~ 0 w u <[ 50 VI 40 J I, Time in months FIG. I. SACE activity in chronic progressive pulmonary sarcoidosis. Corticosteroid treatment withheld at patient's request. XXXXXX: upperlimit ofnormal range. Arrow I: chest radiograph showed bilateral hilar gland enlargement (see CXR I). Arrow 2: chest radiograph showed pulmonary infiltration and fibrosis (see CXR 2). 48

4 16 Studdy and Bird A 'positive' test with SACE values 2 SD or more above the control mean identified approximately 8 out of 10 active sarcoidosis patients. A negative test is of limited diagnostic value. Extremely high SACE levels (three or four times the upper limit of normal) occur with extensive sarcoidosis when there is combined glandular, hepatic, cutaneous and extensive pulmonary involvement, i.e. when there is a large mass of granulomatous inflammation. SACE is seldom raised in isolated cardiac or central nervous system sarcoidosis. 3D Sequential measurements of SACE in sarcoidosis Sequential SACE measurements show a relationship between changing enzyme activity and clinicalor radiographic changes in sarcoidosis. As the disease remits spontaneously or with corticosteroid therapy, raised SACE levels fall towards normal. Relapse is characterised by rising SACE levels. The severity and extent of the disease is broadly reflected by the extent of the raised SACE activity. The moderately elevated SACE levels characteristic of early sarcoidosis tend to rise as the disease progresses and high levels persist in chronic active widespread disease. Pulmonary disease activity may be assessed by serial chest radiographs, which show a good positive correlation with SACE.29 Two other investigations are used to assess activity," namely radioisotope gallium lung scanning and bronchoalveolar lavage (BAL) T lymphocyte profiles." Both show weak correlations with SACE and stronger correlations with the chest radiograph appearances or other conventional parameters of disease activity. Neither gallium lung scanning nor BAL is widely available or generally applicable whereas measurement ofsace is simple and can be repeated at intervals without hazard to the patient. SACE is the only investigative test that gives a measure of both extrathoracic and pulmonary sarcoidosis activity. The biochemical course of a patient with chronic sarcoidosis is demonstrated in Fig. I. SACE and corticosteroids Corticosteroids are valuable in the treatment of sarcoidosis, and their administration usually results in rapid improvement. Elevated SACE activity falls towards normal after corticosteroid treatment has begun, normal values being attained in 4-10 weeks." There is a strong relationship between corticosteroid dosage and SACE activity: most cases show a marked fall in SACE and satisfactory clinical improvement on prednisolone at 15 mg or more per day. Clinical relapse may occur when corticosteroid treatment is withdrawn and may be heralded by a rising SACE. The reduction in SACE activity in sarcoidosis.is thought to be due to inhibition of J E <,!! u o W uet l/l 10 Prednisolone (mgl 'l o Time in months FIG. 2. Acute pulmonary sarcoidosis with satisfactory clinical and radiographic progress during and after active therapy with prednisolone. XXXXXX: upper limit of normal range.

5 Serum angiotensin converting enzyme in sarcoidosis 17 monocyte and lymphocyte activity in the granulomata." The course of a patient with acute sarcoidosis requiring corticosteroids is shown in Fig. 2. SACE in other conditions Elevated SACE activity is not specific to sarcoidosis. Elevated SACE activity is found in some patients with other granulomatous disorders and in several conditions having chest radiograph abnormalities which may be confused with sarcoidosis. Table 1 lists some important examples. The demonstration ofraised enzyme activity in a number of other conditions underlines the limitation of using SACE as a screening test for sarcoidosis. It is, therefore, important for the clinician suspecting sarcoidosis to exclude tuberculosis, to consider Hodgkin's disease or non Hodgkin's lymphoma and in all but the most typical cases to confirm sarcoidosis histologically. In the southern United States of America it may also be necessary to exclude coccidioidomycosis and histoplasmosis. Silicosis and asbestosis may cause confusion and the clinician will need to enquire specifically about dust exposure at work. TABLE I. Serum ACE in diseases other than sarcoidosis No. (%) of measurements >2 SD of Disease No. controls SACE in diseases which may be confused with sarcoidosis Active tuberculosis (4%) Atypical mycobacteria 39 5 (13%) Miliary tuberculosis 9 8 (89%) Treated tuberculosis (10%) Leprosy III 21 (18%) Histoplasmosis 50 7 (14%) Coccidioidomycosis 18 I (6%) Extensive allergic alveoli tis 67 3 (4%) Berylliosis 25 3 (12%) Primary biliary cirrhosis 55 II (20%) Hodgkin's disease (6%) Lung cancer «1%) Lung fibrosis (5%) Asbestosis 32 6 (19%) Silicosis (45%) SACE in other conditions Alcoholic liver disease (28%) Hyperthyroidism (62%) Diabetes mellitus (18%) Gaucher's disease (80%) Bronchitis and emphysema «1%) Bronchial asthma (1%) Conclusions (1) SACE is best performed at a regional or supra-regional centre, rather than at a nonspecialised district laboratory. (2) SACE is a safe, simple, noninvasive test which can be repeated at intervals. (3) SACE is of limited value in the diagnosis of sarcoidosis, since an elevated SACE activity is not exclusive to sarcoidosis. However, in those situations allowing a confidentclinical or radiographic diagnosis of sarcoidosis, raised SACE provides strong supporting evidence to complement other diagnostic procedures. A low SACE activity does not exclude sarcoidosis. (4) SACE parallels the progress of sarcoidosis and is thus extremely useful in monitoring the disease process and the response to therapy with corticosteroids. In clinical practice the dose of corticosteroids may be titrated against serial SACE measurements to control the disease in the most effective way. Similarly, SACE activity can be used to check patients' compliance to treatment. References Skeggs LT, Marsh KM, Shumay NP. Existence of two forms ofhypertensin. J Exp Med 1954; 99: 275~ Skeggs LT, Khan JR, Shumay NP. The preparation and function ofthe hypertensin converting enzyme. J Exp Med 1965; 103: Erdos EG. Angiotensin converting enzyme. Circ. Res 1975; 36: Bakkhle YS. Converting enzyme; in vitro measurement and properties. Handbook Exp Pharm 1974; 37: Soffer RL. Angiotensin converting enzyme and the regulation ofvasoactive peptides. Ann Rev Biochem 1976; 45: Ng KKF, Vane JR. Conversion of angiotensin I to angiotensin II. Nature 1967; 216: Ryan JW, Ryan US, Schultz DR et al. Sub-cellular localization of pulmonary angiotensin converting enzyme. Biochem J 1975; 146: Favre L, Valloton MB, Muller AF. Relationship between plasma concentrations of angiotensin I, angiotensin II and plasma renin activity during cardiopulmonary bypass in man. Eur J CUn Invest 1974; 4: Dobuler KJ, Catravas JD, Gillis CN. Early detection of oxygen induced lung injury in conscious rabbits. Reduced in vivo activity of angiotensin converting enzyme and removal of 5-hydroxytryptarnine. Am Rev Respir Dis 1982; 126: Lieberman J. Elevation of serum angiotensin converting enzyme (ACE) levels in sarcoidosis. Am J Med 1975; 59: II Studdy PS, Bird R, James DG, Sherlock S. Serum angiotensin converting enzyme (SACE) in sarcoidosis and other granulomatous disorders. Lance/1978; ii:

6 18 Studdy and Bird 12 Studdy PS, James DG. Serum angiotensin converting enzyme (SACE): Experience in ten centres. In: Jones-Williams W, Davies BH, eds. Proc 8th International Conference on Sarcoidosis and Other Granulomatous Diseases. Cardiff: Alpha and Omega Press, 1980; Cushman DW, Cheung HS. Concentration of angiotensin converting enzyme in tissues of the rat. Biochim Biophys Acta 1971; 250: Silverstein E, Friedland J, Pertschuk LP. Sarcoidosis pathogenesis. Mechanisms of angiotensin converting enzyme elevation: epithelioid cell localisation and induction in macrophages and monocytes in culture. In: Jones-Williams W, Davies BH, eds. Proc 8th InternationalConferenceon Sarcoidosisand Other Granulomatous Diseases. Cardiff: Alpha and Omega Press, 1980; Pertshuk LP, Silverstein E, Friedland J. Immunohistologic diagnosis of sarcoidosis; detection of angiotensin converting enzyme in sarcoid granulomas. Am J Clin Patho11981; 75: Silverstein E, Friedland J, Stanek AE et al. Pathogenesis of sarcoidosis; mechanism of angiotensin converting enzyme elevation. T-Iymphocyte modulation of enzyme induction in mononuclear phagocytes; Enzyme properties. In: Chretien J, Marsac J, Saltiel JC, eds. Proc 9th International Conference on Sarcoidosis and Other Granulomatous Diseases. Oxford: Pergamon 1983; Friedland J, Silverstein E. A sensitive ftuorimetric assay for serum angiotensin converting enzyme. Am J Clin Patho11976; 66: Ryan JW, Chung A, Ammons C, Carlton ML. A simple radioassay for angiotensin converting enzyme. Biochem J 1977; 167: Rohrbach MS. Glycine-l-v'C hippuryl-l-histidyl-lleucine: A substrate for the radiochemical assay for angiotensin converting enzyme. Anal Biochem 1978; 84: Ronc- Testani S. Direct spectrophotometric assay for angiotensin converting enzyme screen. Clin Chem 1983; 29: Maguire GA, Price CPo A continuous monitoring spectrophotometric method for the measurement of angiotensin converting enzymes in human sera. Ann Clin Biochem 1985; 22: Studdy PR, James DG. The specificity and sensitivity of serum angiotensin converting enzyme in sarcoidosis and other diseases experienced in twelve centres in six countries. In: Chretien J, Marsac J, Saltiel J, eds. Proc 9th International Conference on Sarcoidosis and Other Granulomatous Diseases. Oxford: Pergamon Press, 1983; Rohatgi PK, Ryan JW, Linderman P. Value of serial measurements of serum angiotensin converting enzyme in the management ofsarcoidosis. Am J Med 1981; 70: Bender JW, Davitt MK, Jose P. Angiotensin converting enzyme in term and premature infants. Bioi Neonate 1978; 34: Gronhagen-Riska C, Selroos 0, Niemisto M. Angiotensin converting enzyme and serum levels as monitors of disease activity in corticosteroid controlled sarcoidosis. Eur J Respir Dis 1980; 61: Turton CWG, Grundy E, Firth G et al. Value of measuring serum angiotensin I converting enzyme and serumlysozyme in the managementofsarcoidosis. Thorax 1979; 34: Roulston JE, MacGregor GA, Bird R. The measurement of angiotensin converting enzyme in subjects receiving captopril. N Engl J Med 1980; 303: Scadding JD, Mitchell DN. Sarcoidosis. 2nd ed. London: Chapman and Hall, 1985; Studdy PRo SACE in sarcoidosis. MD Thesis. University of Newcastle-upon-Tyne, Fleming HA, Baily SM. Sarcoid heart disease. J R Coli Physicians (Lond) 1981; 15: Klech H, Kohn H, Kummer F, Mostbeck A. Value ofdifferent parameters for the assessment ofactivity in sarcoidosis: X-ray, gallium 67 scanning, serum ACE levels and blood lymphocyte sub-populations. In: Chretien J, Marsac J, Saltiel J, eds. Proc 9th International Conference on Sarcoidosis and Other Granulomatous Diseases. Oxford: Pergamon Press. 1983; Klech H, Haslam P, Turner-Warwick M. Worldwide clinical survey in broncho-alveolar lavage (BAL) in sarcoidosis. Experience in 62 centres in 19 countries. Preliminary analysis. In: Chretien J, Marsac J, Saltiel J eds. Proc 9th International Conference on Sarcoidosis and Other Granulomatous Diseases. Oxford: Pergamon Press, 1983; Lieberman J, Nosal A, Schlessner LA, Sastre-Foker A. Serum angiotensin converting enzyme for diagnosis and therapeuticevaluation ofsarcoidosis. Am Rev Respir Dis 1979; 120: Rohrbach MS, Deremee RA. Serum angiotensin converting enzyme activity in sarcoidosis as measured by a simple radiochemical test. Am Rev Respir Dis 1979; 119: Acceptedfor publication 27 June 1988