Clin Biochern, Vol. 23, pp. 229-232, 1990 Printed in Canada. All righte reserved.
0009-9120/90 $3.00 + .00 Copyright ¢ 1990 The Canadian Society of Clinical Chemists.
Serum Phospholipase A2 Activity in Chronic Pancreatic Diseases DANIELA BASSO, CARLO FABRIS, MARIA PIERA PANOZZO, TAMARA MEGGIATO, GIUSEPPE DEL FAVERO, and REMO NACCARATO Istituto di Medicina Interna, Cattedra di Malattie Apparato Digerente, Universit& degli Studi di Padova, Italy This study was performed to investigate the phospholipase A2 (PLA2) serum activity in patients with chronic pancreatic disease. PLA2, elastase-1, total, and pancreatic isoamylase were evaluated in 40 control subjects, 28 patients with pancreatic cancer, 51 with chronic pancreatitis, and 36 with extrapancreatic diseases, mainly of gastrointestinal origin. Elastase-1, PLA2, and pancreatic isoamylase were increased in 56%, 25%, and 15% of patients with pancreatic cancer, and in 40%, 31%, and 41% of subjects with chronic pancreatitis. All four enzymes gave pathological values in a number of patients with extrapancreatic diseases. We conclude that the diagnostic efficacy of phospholipase A2 in chronic pancreatic disease is similar to that of other well known pancreatic enzymes, with an unsatisfactory sensitivity and specificity.
KEY WORDS: phospholipase A2; elastase-1; amylase; pancreatic isoamylase; pancreatic cancer; chronic pancreatitis. Introduction
p
hospholipase A2 (PLA2), an enzyme hydrolyzing phospholipids, is secreted by the pancreatic acinar cells (1); it plays an important role in the pathogenesis of pancreatic inflammation, due to its noxious activity on cellular membranes (2). An immunological method to determine this enzyme in serum has been described and employed in the attempt to establish both the severity and etiology of acute pancreatitis (3-6). It has been concluded that this assay may be useful in assessing patients with severe alcoholic pancreatitis (7,8). Immunological assays determine both active and inactive forms of enzymes, such as, proenzymes and complexes with inhibitors. An enzymatic method might, therefore, be more effective in the investigation of the pathophysiological role of PLA2 in pancreatic diseases. Furthermore, no comparison has been made between the behavior of this enzyme in
Correspondence: Prof. Remo Naccarato, Istituto di Medicina Interna, Cattedra di Malattie Apparato Digerente, Policlinico Universitario, Via Giustiniani 2, 35100 Padova, Italy. Manuscript received May 5, 1990; revised January 4, 1990; accepted January 8, 1990. CLINICAL BIOCHEMISTRY, VOLUME 23, JUNE 1990
pancreatic and in extrapancreatic diseases. We, therefore, investigated PLA2 serum activity in patients with pancreatic cancer, chronic pancreatitis, and other diseases mainly of gastrointestinal origin. A comparison was made with three other enzymes routinely employed in clinical pancreatology: amylase, pancreatic isoamylase, and elastase-1. Patients
One hundred fifty-five subjects were studied. Forty were control subjects (29 males, 11 females, aged 19-66), healthy members of the medical staff, or blood donors. Twenty-eight subjects had pancreatic cancer of duct cell origin (13 males, 15 females, aged 28-76) confirmed histologically (9); 16 of these patients had liver metastases. Fifty-one patients had chronic pancreatitis (47 males, four females, aged 27-73), diagnosed on the basis of the clinical picture and on positive findings of at least two of the following examinations: plain abdomen X-ray for pancreatic calcifications, pancreatic ultrasonography, CT scanning and endoscopic retrograde pancreatography. Nineteen of these patients were studied during a relapse. Thirty-six patients had extrapancreatic diseases (26 males, 10 females, aged 38-77), mainly of gastrointestinal origin. The diagnoses were based on the clinical picture and on the specific radiological and histological findings: liver cirrhosis (eight cases), gallstones (four cases), chronic hepatitis (two cases), primary liver cell cancer (two cases), main bile duct cancer (two cases), benign stenosis of the papilla of Vater (two cases), duodenal ulcer (one case), irritable colon (one case), gastritits (one case), colorectal carcinoma (two cases), gastric cancer (one case), carcinoma of the esophagus (one case), hiatus hernia (one case), duodenal lymphoma (one case), celiac disease (two cases), Crohn's disease (one case), diverticulosis of the colon (one case), portal vein thrombosis (one case), liver hemangioma (one case), and acute cholecystitis (one case). Methods
Phospholipase A2 was determined in serum by a fluorimetric assay (10). Serum elastase-1 was mea229
BASSO, FABRIS, PANOZZO, ET AL.
sured by an RIA procedure using a commercial kit (Abbott). Pancreatic isoamylase was assessed using a new pancreas specific-amylase antibody assay (11). Total amylase activity was evaluated using Rausctier's method (12). Statistical analysis of the data was made by using analysis of variance (Anova one way), Bonferroni's test for pairwise comparisons (13), Student's t-test and chi-square test.
20OO
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Results
t,ooo O O
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oo 1~0 m
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~
ee
,
t!
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• 0
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t'i,o
Figure 1--Individual values of phospholipase A2 (PLA2) in the patients studied. The continuous line represents the upper normal limit (mean + 2SD of our controls). CS: control subjects; PC: pancreatic cancer; CP: chronic pancreatitis; EPD: extrapancreatic diseases. Triangles: metastatic pancreatic cancers; open dots: chronic pancreatitis studied during a relapse.
Figure 1 shows the individual values of PLA2 in the patients studied. Figure 2 reports the percentage of pathological values and the statistical analysis of the four enzymes in the different groups of subjects. The chi-square tests showed t h a t in the different groups of patients, all the enzymes had pathological values with a higher frequency t h a n in the controls. Table 1 presents means, standard errors, and a statistical evaluation of the data. The analysis of variance showed t h a t the mean enzyme values differed significantly among the four groups of subjects. The pairwise comparisons made using Bonferroni's test indicated significant differences between the groups specified in Table 1. No differences were found between the enzyme values of pancreatic cancer patients with and without liver metastases. Significant correlations were found between PLA2 and elastase-1 (r=0.518, p < 0.001), pancreatic isoamylase (r= 0.317, p < 0.01), and total amylase (r=0.301, p < 0:01).
90
r ~ l CS ill Ill 7 0
ii~
PC
ii
CP
llill E P D
X~15.6
X2:13.7
X~ 2 0 . 3
X~=19.2
p < 0.001
P < 0.005
p < 0.0001
P < 0.0001
ISOAMYLA~
ELASTASE1
USO-
PLA 2
AMYLASE
Figure 2--Percentages of pathological values and a statistical analysis of phospholipase A2 (PLA2), elastase-1, total amylase and pancreatic isoamylase in the four groups of subjects studied. CS: control subjects; PC: pancreatic cancer; CP: chronic pancreatitis; EPD: extrapancreatic diseases.
230
CLINICAL BIOCHEMISTRY, VOLUME 23, JUNE 1990
PHOSPHOLIPASE A2 IN PANCREATIC DISEASES TABLE 1
Mean Values, Standard Errors and Statistical Analysis of Serum Phospholipase A2 (PLA2), Elastase-1, Total, and Pancreatic Isoamylase Cases PLA2 (pmol min-1 mL-1)
Control subjects Pancreatic cancer Chronic pancreatitis Extra-pancreatic diseases Total Analysis of variance
Elastase-1 (ng/dL)
Amylase (U/L) Isoamylase (U/L)
No.
Mean
SEM
Mean
SEM
Mean
SEM
Mean
SEM
40 28 51 36
285.2 389.2 518.3 a 476.0 b
16.8 37.9 58.1 52.3
166.8 518.6 787.2' 415.3
15.6 104.7 158.5 56.5
93.3 101.5 169.3b 166.1
6.1 20.9 23.6 22.4
47.1 69.3 117.0b 88.1
2.9 18.9 23.8 12.8
155 F = 5.03 p < 0.005
F = 6.10 p < 0.001
F = 4.08 p < 0.01
F = 3.09 p < 0.05
Bonferroni's test for pairwise comparisons: a p < 0.005 with respect to control subjects; b p < 0.05 with respect to control subjects.
Discussion Phospholipase A2 is a low molecular weight ( - 1 5 kDa) enzyme (3) which can be liberated from pancreatic acinar cells into the circulation under normal conditions (5-7); it is then probably excreted in the urine (6). High serum levels of this enzyme could be expected in pancreatic inflammatory diseases due to cell necrosis (7,8). A number of our patients with chronic pancreatitis had high circulating PLA2 levels. However the finding was not limited to patients studied during a relapsing phase of the disease. Furthermore, in pancreatic cancer, PLA2 was found to be abnormal at times. Therefore, the diagnostic efficacy of this test in patients with chronic pancreatic diseases seems to be low, and similar to t h a t of other well known enzymes (14). Pancreatic isoamylase was found to be elevated in 11/19 patients with chronic relapsing pancreatitis, a percentage similar to t h a t found for elastase-1 and total amylase. On the other hand, in patients with pancreatic cancer, increased values of elastase-1 were found in a higher percentage t h a n t h a t found for PLA2; both percentages were higher t h a n those found for total and pancreatic isoamylase. This finding indicates t h a t peritumoral cell necrosis may not be the only factor responsible for the increase in PLA2 in pancreatic cancer. Other factors, such as those claimed to play a role in the increase in elastase-1 in this disease, might be considered, such as an imbalance between proteases and antiproteases (15). The need for specificity in pancreatic clinical biochemistry has always been a crucial point in research. With this in mind, a pancreatic isoamylase assay using a monoclonal antibody has been developed (11). An increased sensitivity in detecting acute pancreatitis has been obtained using this test (16); this was not confirmed in our study concerning chronic pancreatic diseases, considering either the
CLINICAL BIOCHEMISTRY, VOLUME 23, JUNE 1990
sensitivity or the specificity of the test. Two other factors may explain the increased isoamylase values in patients with extrapancreatic diseases: 1) the presence of subclinical pancreatic damage in these patients; 2) an impairment in pancreatic isoamylase metabolism. The same considerations could apply to PLA2, and to elastase-1, in view of the similar molecular weight of all these enzymes. Furthermore, it has never been unequivocally demonstrated t h a t circulating elastase-1 and PLA2 are derived only from the pancreas. We conclude t h a t in the assessment of patients with chronic pancreatic diseases, PLA2 determination fails to provide more information t h a n t h a t obtained using the measurement of other well known pancreatic enzymes. The sensitivity and specificity of all these tests in the diagnosis of chronic pancreatic inflammation or pancreatic carcinoma appear to be poor; they are of some help only in detecting clinical relapses of chronic pancreatitis.
Acknowledgements Supported in part by a grant from Centro Regionale di
Alta Specializzazione per lo Studio delle Malattie del Fegato e del Pancreas. Under the auspices of the "R. Farini Association for Gastroenterological Research."
References 1. Grataroli R, De Caro A, Guy O, Amic J, Figarella C. Isolation and properties of prophospholipase A2 from human pancreatic juice. Biochimie 1981; 63: 677-84. 2. Nevalainen TJ. The role of phospholipase A2 in acute pancreatitis. Scand J Gastroenterol 1980; 15: 641-50. 3. Eskola JU, Nevalainen TJ, Aho HJ. Purification and characterization of human pancreatic phospholipase A2. Clin Chem 1983; 29: 1772-6. 4. Eskola JU, Nevalainen TJ, Lovgren TN-E. Timeresolved fluoroimmunoassay of human pancreatic phospholipase A2. Clin Chem 1983; 29: 1777-80.
231
BASSO, FABRIS, PANOZZO, ZT AL. 5. Nishijima J, Okamote M, Ogawa M, Kosaki G, Yamano T. Purification and characterization of human pancreatic phospholipase A2 and development of a radioimmunoassay. J Biochem (Tokyo) 1983; 94: 137-47. 6. Sternby B, Akerstrom B. Immunoreactive pancreatic colipase, lipase, and phospholipase A2 in human plasma and urine from healthy individuals. Biochim Biophys Acta 1984; 789: 164-9. 7. Nevalainen TJ, Eskola JU, Aho AJ, Havia VT, Lovgren TN-E, Nanto V. Immunoreactive phospholipase A2 in serum in acute pancreatitis and pancreatic cancer. Clin Chem 1985; 31: 1116-20. 8. Matsuda Y, Ogawa M, Nishijima J, Miyauchi K, Mori T. Usefulness of determination of serum immunoreactive pancreatic phospholipase A2 content for early identification of severe acute pancreatitis. Hepatogastroenterol 1986; 33: 214-6. 9. Cubilla AL, Fitzgerald PJ. Pancreas cancer. 1. Duct adenocarcinoma. Pathol Annu 1978; 13: 241-89. 10. Thuren T, Virtanen JA, Lalla M, Kinnunen PKJ. Fluorometric assay for phospholipase A2 in serum.
232
Clin Chem 1985; 31: 714-7. 11. Gerber M, Naujoks K, Lenz H, WulffK. A monoclonal antibody that specifically inhibits human salivary c~-amylase. Clin Chem 1987; 33: 1158-62. 12. Rauscher E, Neuman V, Schaich E, Von Bulow S, Wahlefeld AW. Optimized conditions for determining activity concentration of s-amylase in serum, with 1,4-~-D-4-nitrophenylmaltoheptaoside as substrate. Clin Chem 1985; 31: 14-9. 13. Wallenstein S, Zucker CL, Fleiss JL. Some statistical methods useful in circulation research. Circ Res 1980; 47: 1-9. 14. Ventrucci M, Gullo L, Daniele C, et al. Comparative study of serum pancreatic isoamylase, lipase, and trypsin-like immunoreactivity in pancreatic disease. Digestion 1983; 28: 114-21. 15. Travis J, Salvesen GS. H u m a n plasma proteinase inhibitors. Annu Rev Biochem 1983; 52: 655-709. 16. Tietz NW, Burlina A, Gerhardt W, et al. Multicenter evaluation of a specific pancreatic isoamylase assay based on a double monoclonal-antibody technique. Clin Chem 1988; 34: 2096-102.
CLINICAL BIOCHEMISTRY, VOLUME 23, JUNE 1990
0009-9120/90 $3.00 + .00 Copyright ¢ 1990 The Canadian Society of Clinical Chemists.
Serum Phospholipase A2 Activity in Chronic Pancreatic Diseases DANIELA BASSO, CARLO FABRIS, MARIA PIERA PANOZZO, TAMARA MEGGIATO, GIUSEPPE DEL FAVERO, and REMO NACCARATO Istituto di Medicina Interna, Cattedra di Malattie Apparato Digerente, Universit& degli Studi di Padova, Italy This study was performed to investigate the phospholipase A2 (PLA2) serum activity in patients with chronic pancreatic disease. PLA2, elastase-1, total, and pancreatic isoamylase were evaluated in 40 control subjects, 28 patients with pancreatic cancer, 51 with chronic pancreatitis, and 36 with extrapancreatic diseases, mainly of gastrointestinal origin. Elastase-1, PLA2, and pancreatic isoamylase were increased in 56%, 25%, and 15% of patients with pancreatic cancer, and in 40%, 31%, and 41% of subjects with chronic pancreatitis. All four enzymes gave pathological values in a number of patients with extrapancreatic diseases. We conclude that the diagnostic efficacy of phospholipase A2 in chronic pancreatic disease is similar to that of other well known pancreatic enzymes, with an unsatisfactory sensitivity and specificity.
KEY WORDS: phospholipase A2; elastase-1; amylase; pancreatic isoamylase; pancreatic cancer; chronic pancreatitis. Introduction
p
hospholipase A2 (PLA2), an enzyme hydrolyzing phospholipids, is secreted by the pancreatic acinar cells (1); it plays an important role in the pathogenesis of pancreatic inflammation, due to its noxious activity on cellular membranes (2). An immunological method to determine this enzyme in serum has been described and employed in the attempt to establish both the severity and etiology of acute pancreatitis (3-6). It has been concluded that this assay may be useful in assessing patients with severe alcoholic pancreatitis (7,8). Immunological assays determine both active and inactive forms of enzymes, such as, proenzymes and complexes with inhibitors. An enzymatic method might, therefore, be more effective in the investigation of the pathophysiological role of PLA2 in pancreatic diseases. Furthermore, no comparison has been made between the behavior of this enzyme in
Correspondence: Prof. Remo Naccarato, Istituto di Medicina Interna, Cattedra di Malattie Apparato Digerente, Policlinico Universitario, Via Giustiniani 2, 35100 Padova, Italy. Manuscript received May 5, 1990; revised January 4, 1990; accepted January 8, 1990. CLINICAL BIOCHEMISTRY, VOLUME 23, JUNE 1990
pancreatic and in extrapancreatic diseases. We, therefore, investigated PLA2 serum activity in patients with pancreatic cancer, chronic pancreatitis, and other diseases mainly of gastrointestinal origin. A comparison was made with three other enzymes routinely employed in clinical pancreatology: amylase, pancreatic isoamylase, and elastase-1. Patients
One hundred fifty-five subjects were studied. Forty were control subjects (29 males, 11 females, aged 19-66), healthy members of the medical staff, or blood donors. Twenty-eight subjects had pancreatic cancer of duct cell origin (13 males, 15 females, aged 28-76) confirmed histologically (9); 16 of these patients had liver metastases. Fifty-one patients had chronic pancreatitis (47 males, four females, aged 27-73), diagnosed on the basis of the clinical picture and on positive findings of at least two of the following examinations: plain abdomen X-ray for pancreatic calcifications, pancreatic ultrasonography, CT scanning and endoscopic retrograde pancreatography. Nineteen of these patients were studied during a relapse. Thirty-six patients had extrapancreatic diseases (26 males, 10 females, aged 38-77), mainly of gastrointestinal origin. The diagnoses were based on the clinical picture and on the specific radiological and histological findings: liver cirrhosis (eight cases), gallstones (four cases), chronic hepatitis (two cases), primary liver cell cancer (two cases), main bile duct cancer (two cases), benign stenosis of the papilla of Vater (two cases), duodenal ulcer (one case), irritable colon (one case), gastritits (one case), colorectal carcinoma (two cases), gastric cancer (one case), carcinoma of the esophagus (one case), hiatus hernia (one case), duodenal lymphoma (one case), celiac disease (two cases), Crohn's disease (one case), diverticulosis of the colon (one case), portal vein thrombosis (one case), liver hemangioma (one case), and acute cholecystitis (one case). Methods
Phospholipase A2 was determined in serum by a fluorimetric assay (10). Serum elastase-1 was mea229
BASSO, FABRIS, PANOZZO, ET AL.
sured by an RIA procedure using a commercial kit (Abbott). Pancreatic isoamylase was assessed using a new pancreas specific-amylase antibody assay (11). Total amylase activity was evaluated using Rausctier's method (12). Statistical analysis of the data was made by using analysis of variance (Anova one way), Bonferroni's test for pairwise comparisons (13), Student's t-test and chi-square test.
20OO
"T
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i
i¢
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•
1400
•
Results
t,ooo O O
497AI,C
"il"
oo 1~0 m
•
•
8"
""
:';
"::
•
200
~
ee
,
t!
2P"
• 0
cs
Pi:
cP
••
t'i,o
Figure 1--Individual values of phospholipase A2 (PLA2) in the patients studied. The continuous line represents the upper normal limit (mean + 2SD of our controls). CS: control subjects; PC: pancreatic cancer; CP: chronic pancreatitis; EPD: extrapancreatic diseases. Triangles: metastatic pancreatic cancers; open dots: chronic pancreatitis studied during a relapse.
Figure 1 shows the individual values of PLA2 in the patients studied. Figure 2 reports the percentage of pathological values and the statistical analysis of the four enzymes in the different groups of subjects. The chi-square tests showed t h a t in the different groups of patients, all the enzymes had pathological values with a higher frequency t h a n in the controls. Table 1 presents means, standard errors, and a statistical evaluation of the data. The analysis of variance showed t h a t the mean enzyme values differed significantly among the four groups of subjects. The pairwise comparisons made using Bonferroni's test indicated significant differences between the groups specified in Table 1. No differences were found between the enzyme values of pancreatic cancer patients with and without liver metastases. Significant correlations were found between PLA2 and elastase-1 (r=0.518, p < 0.001), pancreatic isoamylase (r= 0.317, p < 0.01), and total amylase (r=0.301, p < 0:01).
90
r ~ l CS ill Ill 7 0
ii~
PC
ii
CP
llill E P D
X~15.6
X2:13.7
X~ 2 0 . 3
X~=19.2
p < 0.001
P < 0.005
p < 0.0001
P < 0.0001
ISOAMYLA~
ELASTASE1
USO-
PLA 2
AMYLASE
Figure 2--Percentages of pathological values and a statistical analysis of phospholipase A2 (PLA2), elastase-1, total amylase and pancreatic isoamylase in the four groups of subjects studied. CS: control subjects; PC: pancreatic cancer; CP: chronic pancreatitis; EPD: extrapancreatic diseases.
230
CLINICAL BIOCHEMISTRY, VOLUME 23, JUNE 1990
PHOSPHOLIPASE A2 IN PANCREATIC DISEASES TABLE 1
Mean Values, Standard Errors and Statistical Analysis of Serum Phospholipase A2 (PLA2), Elastase-1, Total, and Pancreatic Isoamylase Cases PLA2 (pmol min-1 mL-1)
Control subjects Pancreatic cancer Chronic pancreatitis Extra-pancreatic diseases Total Analysis of variance
Elastase-1 (ng/dL)
Amylase (U/L) Isoamylase (U/L)
No.
Mean
SEM
Mean
SEM
Mean
SEM
Mean
SEM
40 28 51 36
285.2 389.2 518.3 a 476.0 b
16.8 37.9 58.1 52.3
166.8 518.6 787.2' 415.3
15.6 104.7 158.5 56.5
93.3 101.5 169.3b 166.1
6.1 20.9 23.6 22.4
47.1 69.3 117.0b 88.1
2.9 18.9 23.8 12.8
155 F = 5.03 p < 0.005
F = 6.10 p < 0.001
F = 4.08 p < 0.01
F = 3.09 p < 0.05
Bonferroni's test for pairwise comparisons: a p < 0.005 with respect to control subjects; b p < 0.05 with respect to control subjects.
Discussion Phospholipase A2 is a low molecular weight ( - 1 5 kDa) enzyme (3) which can be liberated from pancreatic acinar cells into the circulation under normal conditions (5-7); it is then probably excreted in the urine (6). High serum levels of this enzyme could be expected in pancreatic inflammatory diseases due to cell necrosis (7,8). A number of our patients with chronic pancreatitis had high circulating PLA2 levels. However the finding was not limited to patients studied during a relapsing phase of the disease. Furthermore, in pancreatic cancer, PLA2 was found to be abnormal at times. Therefore, the diagnostic efficacy of this test in patients with chronic pancreatic diseases seems to be low, and similar to t h a t of other well known enzymes (14). Pancreatic isoamylase was found to be elevated in 11/19 patients with chronic relapsing pancreatitis, a percentage similar to t h a t found for elastase-1 and total amylase. On the other hand, in patients with pancreatic cancer, increased values of elastase-1 were found in a higher percentage t h a n t h a t found for PLA2; both percentages were higher t h a n those found for total and pancreatic isoamylase. This finding indicates t h a t peritumoral cell necrosis may not be the only factor responsible for the increase in PLA2 in pancreatic cancer. Other factors, such as those claimed to play a role in the increase in elastase-1 in this disease, might be considered, such as an imbalance between proteases and antiproteases (15). The need for specificity in pancreatic clinical biochemistry has always been a crucial point in research. With this in mind, a pancreatic isoamylase assay using a monoclonal antibody has been developed (11). An increased sensitivity in detecting acute pancreatitis has been obtained using this test (16); this was not confirmed in our study concerning chronic pancreatic diseases, considering either the
CLINICAL BIOCHEMISTRY, VOLUME 23, JUNE 1990
sensitivity or the specificity of the test. Two other factors may explain the increased isoamylase values in patients with extrapancreatic diseases: 1) the presence of subclinical pancreatic damage in these patients; 2) an impairment in pancreatic isoamylase metabolism. The same considerations could apply to PLA2, and to elastase-1, in view of the similar molecular weight of all these enzymes. Furthermore, it has never been unequivocally demonstrated t h a t circulating elastase-1 and PLA2 are derived only from the pancreas. We conclude t h a t in the assessment of patients with chronic pancreatic diseases, PLA2 determination fails to provide more information t h a n t h a t obtained using the measurement of other well known pancreatic enzymes. The sensitivity and specificity of all these tests in the diagnosis of chronic pancreatic inflammation or pancreatic carcinoma appear to be poor; they are of some help only in detecting clinical relapses of chronic pancreatitis.
Acknowledgements Supported in part by a grant from Centro Regionale di
Alta Specializzazione per lo Studio delle Malattie del Fegato e del Pancreas. Under the auspices of the "R. Farini Association for Gastroenterological Research."
References 1. Grataroli R, De Caro A, Guy O, Amic J, Figarella C. Isolation and properties of prophospholipase A2 from human pancreatic juice. Biochimie 1981; 63: 677-84. 2. Nevalainen TJ. The role of phospholipase A2 in acute pancreatitis. Scand J Gastroenterol 1980; 15: 641-50. 3. Eskola JU, Nevalainen TJ, Aho HJ. Purification and characterization of human pancreatic phospholipase A2. Clin Chem 1983; 29: 1772-6. 4. Eskola JU, Nevalainen TJ, Lovgren TN-E. Timeresolved fluoroimmunoassay of human pancreatic phospholipase A2. Clin Chem 1983; 29: 1777-80.
231
BASSO, FABRIS, PANOZZO, ZT AL. 5. Nishijima J, Okamote M, Ogawa M, Kosaki G, Yamano T. Purification and characterization of human pancreatic phospholipase A2 and development of a radioimmunoassay. J Biochem (Tokyo) 1983; 94: 137-47. 6. Sternby B, Akerstrom B. Immunoreactive pancreatic colipase, lipase, and phospholipase A2 in human plasma and urine from healthy individuals. Biochim Biophys Acta 1984; 789: 164-9. 7. Nevalainen TJ, Eskola JU, Aho AJ, Havia VT, Lovgren TN-E, Nanto V. Immunoreactive phospholipase A2 in serum in acute pancreatitis and pancreatic cancer. Clin Chem 1985; 31: 1116-20. 8. Matsuda Y, Ogawa M, Nishijima J, Miyauchi K, Mori T. Usefulness of determination of serum immunoreactive pancreatic phospholipase A2 content for early identification of severe acute pancreatitis. Hepatogastroenterol 1986; 33: 214-6. 9. Cubilla AL, Fitzgerald PJ. Pancreas cancer. 1. Duct adenocarcinoma. Pathol Annu 1978; 13: 241-89. 10. Thuren T, Virtanen JA, Lalla M, Kinnunen PKJ. Fluorometric assay for phospholipase A2 in serum.
232
Clin Chem 1985; 31: 714-7. 11. Gerber M, Naujoks K, Lenz H, WulffK. A monoclonal antibody that specifically inhibits human salivary c~-amylase. Clin Chem 1987; 33: 1158-62. 12. Rauscher E, Neuman V, Schaich E, Von Bulow S, Wahlefeld AW. Optimized conditions for determining activity concentration of s-amylase in serum, with 1,4-~-D-4-nitrophenylmaltoheptaoside as substrate. Clin Chem 1985; 31: 14-9. 13. Wallenstein S, Zucker CL, Fleiss JL. Some statistical methods useful in circulation research. Circ Res 1980; 47: 1-9. 14. Ventrucci M, Gullo L, Daniele C, et al. Comparative study of serum pancreatic isoamylase, lipase, and trypsin-like immunoreactivity in pancreatic disease. Digestion 1983; 28: 114-21. 15. Travis J, Salvesen GS. H u m a n plasma proteinase inhibitors. Annu Rev Biochem 1983; 52: 655-709. 16. Tietz NW, Burlina A, Gerhardt W, et al. Multicenter evaluation of a specific pancreatic isoamylase assay based on a double monoclonal-antibody technique. Clin Chem 1988; 34: 2096-102.
CLINICAL BIOCHEMISTRY, VOLUME 23, JUNE 1990
