Lactate Dehydrogenase-1 Research Articles

Analytical Evaluation and Comparison of Dupont aca® Lactate Dehydrogenase-1 (LD1) Isoenzyme Assay Diagnostic Efficiency for Acute Myocardial Infarction Detection with Other LD1 Methods and aca@ CK-MB

The purpose of this study was to examine the analytical characteristics of a rapid new assay for lactate dehydrogenase 1 (LD1) isoenzyme on the Dupont aca analyzer and the diagnostic efficiency of LD1 for detection of acute myocardial infarction (AMI) when used alone and with creatine kinase MB (CKMB). Total aca LD1 assay precision, with percent coefficients of variation (%CVs) of less than 3.2% from 56 U/L to 469 U/L LD1, across fifty assay days was excellent. Linearity was confirmed from 0 to 332 U/L and no detectable calibration drift was noted from 5 to 489 U/L over a ninety-day period. The aca LD1 results compared well with Roche Isomune-LD1, Abbott Spectrum A-Gent, Helena electrophoresis, and Beckman electrophoresis LD1 methods, giving r's from 0.987 to 0.994, slopes from 0.94 to 1.65, and y-intercepts from -2.95 to 6.94 U/L. Examination of 450 ambulatory subjects, about equally distributed by sex, yielded a 43 +/- 14 U/L aca LD1 patient reference interval. Serum samples from 159 consecutive patients at the University of Tennessee Memorial Hospital and 96 patients at Allentown Hospital, submitted for AMI detection assistance, were assayed in a single-blind study for LD1 and ion-exchange CKMB by Dupont aca methods, which provide automated results in ten minutes whenever needed. The aca LD1 assay yielded a clinical sensitivity of 89% and specificity of 95% for AMI with a decision threshold of 120 U/L. The diagnostic efficiency of the aca LD1 assay was 94% at 120 U/L, which equaled or exceeded that of the three comparative LD1 methods. The predictive value of positive (PV+) and negative (PV-) results on the first sample collected were 80% and 85% for aca LD1, 65% and 90% for aca ion-exchange CKMB, and 83% and 90% when both tests were combined. Significantly, the PV+ and PV- results when two or more samples were assayed was improved to 88% and 95% for aca LD1, relatively constant at 65% and 97% for aca ion-exchange CKMB, and dramatically improved to 95% and 100% when both CKMB and LD1 tests were combined. The results from these two medical centers show that the aca LD1 assay provides useful clinical information for AMI diagnosis when employed alone or in combination with aca CKMB. These results also suggest that LD1 should be included in biochemical evaluations for AMI to attain optimal predictive values of results.

Testicular cancer

The human genome project has revolutionized technology for the study of DNA. Several of this year's papers have applied these techniques to the study of testicular cancer, especially the use of double-fluorescence in situ hybridization to identify the germ cell tumor marker isochrome 12p in tissue sections, and loss-of-heterozygosity studies to demonstrate a candidate suppressor gene on the long arm of the same chromosome that may be a ligand for the c-kit protooncogene. The report of a solitary case of a patient with a tumor (in a solitary testis) who was treated by partial orchiectomy and then fathered two children emphasizes the need for more information on fertility of patients with carcinoma in situ before the highly effective low-dose radiation to the testis can be accepted. The confirmation of the occurrence of acute myeloid leukemia as a late effect of etoposide and stomach cancer as a late effect of radiotherapy for stage I seminoma has drawn attention to the need to reduce treatment in good-risk patients. Results of trials substituting cisplatin with carboplatin and a trial eliminating bleomycin are particularly disappointing, and the 10% lower cures with the experimental regimen in these studies is an object lesson of the risks involved in such studies. Two other issues that continue to be debated include the increasing recognition of the value of lactate dehydrogenase-1 for identifying poor-risk patients, and the benefits of referral to a unit specializing in testicular cancer. There is an increasing trend to use high-dose chemotherapy for previously untreated patients who have poor risk factors. who have poor risk factors.

High plasma concentration of myeloperoxidase in cirrhosis: A possible marker of hypersplenism

Plasma myeloperoxidase levels in patients with cirrhosis were compared with those in patients with chronic hepatitis and healthy controls by means of a specific radioimmunoassay for myeloperoxidase. The mean concentration of plasma myeloperoxidase in cirrhotic patients (309.1 +/- 17.2 ng/ml, n = 41) was markedly higher than that in chronic hepatitis patients (222.6 +/- 17.2 ng/ml, n = 21) (p < 0.01) and normal controls (219.5 +/- 5.7 ng/ml, n = 50) (p < 0.01). Plasma myeloperoxidase showed good negative correlations with neutrocyte count (r = -0.32, p < 0.01), thrombocyte count (r = -0.40, p < 0.01), red blood cell count (r = -0.32, p < 0.01), serum albumin (r = -0.35, p < 0.01), and cholinesterase (r = -0.32, p < 0.02) and positive correlations with serum alkaline phosphatase (r = 0.49; p < 0.01) and lactate dehydrogenase (r = 0.31, p < 0.01) in patients with cirrhosis or chronic hepatitis. Among lactate dehydrogenase isozymes, a good positive correlation was seen between plasma myeloperoxidase and lactate dehydrogenase-2 (r = 0.40, p < 0.01) and lactate dehydrogenase-1 (r = 0.03, p < 0.02). Plasma myeloperoxidase was significantly higher in the cirrhotic and chronic hepatitis patients with splenomegaly (341.1 +/- 19.4 ng/ml, n = 31) than in those without splenomegaly (217.4 +/- 12.2 ng/ml, n = 29) (p < 0.01). We also examined the difference between plasma levels of myeloperoxidase in the portal and peripheral blood.(ABSTRACT TRUNCATED AT 250 WORDS)

High plasma concentration of myeloperoxidase in cirrhosis: A possible marker of hypersplenism

Plasma myeloperoxidase levels in patients with cirrhosis were compared with those in patients with chronic hepatitis and healthy controls by means of a specific radioimmunoassay for myeloperoxidase. The mean concentration of plasma myeloperoxidase in cirrhotic patients (309.1 ± 17.2 ng/ml, n = 41) was markedly higher than that in chronic hepatitis patients (222.6 ± 17.2 ng/ml, n = 21) (p < 0.01) and normal controls (219.5 ± 5.7 ng/ml, n = 50) (p < 0.01). Plasma myeloperoxidase showed good negative correlations with neutrocyte count (r = −0.32, p < 0.01), thrombocyte count (r = −0.40, p < 0.01), red blood cell count (r = −0.32, p < 0.01), serum albumin (r = −0.35, p < 0.01), and cholinesterase (r = −0.32, p < 0.02) and positive correlations with serum alkaline phosphatase (r = 0.49; p < 0.01) and lactate dehydrogenase (r = 0.31, p < 0.01) in patients with cirrhosis or chronic hepatitis. Among lactate dehydrogenase isozymes, a good positive correlation was seen between plasma myeloperoxidase and lactate dehydrogenase-2 (r = 0.40, p < 0.01) and lactate dehydrogenase-1 (r = 0.03, p < 0.02). Plasma myeloperoxidase was significantly higher in the cirrhotic and chronic hepatitis patients with splenomegaly (341.1 ± 19.4 ng/ml, n = 31) than in those without splenomegaly (217.4 ± 12.2 ng/ml, n = 29) (p < 0.01). We also examined the difference between plasma levels of myeloperoxidase in the portal and peripheral blood. In the patients with nonhepatic disease, we found a significant portalsystemic difference (portal myeloperoxidase, 221.5 ± 19.7 ng/ml, vs. systemic myeloperoxidase, 179.1 ± 10.8 ng/ml; n = 15) (p < 0.01), whereas there was no such difference in the cirrhotic patients (portal myeloperoxidase, 281.5 ± 15.2 ng/ml vs. systemic myeloperoxidase, 268.0 ± 28.2 ng/ml; n = 7). A significantly high concentration of plasma myeloperoxidase was evident in the case of esophageal varices, which is a sign of portalsystemic shunting in human beings. The results of our study imply that the release of myeloperoxidase from destroyed neutrocytes, in addition to its decreased hepatic clearance, are responsible for the high concentrations of plasma myeloperoxidase in cirrhotic patients. Therefore measurement of plasma myeloperoxidase may be useful in assessing hypersplenic function in liver disease. (HEPATOLOGY 1993;18:1377–1383.)

Characterization and mathematical correction of hemolysis interference in selected Hitachi 717 assays

The effect of hemolysis on several assays performed with the Hitachi 717 was quantified by relating the amount of error to the concentration of hemoglobin. Hemolysis interference was judged clinically significant when analyte concentration varied by > 10% from the initial value. Hemolysis interference was significant for alkaline phosphatase, aspartate aminotransferase, alpha-amylase, bilirubin, creatine kinase, gamma-glutamyltransferase, lactate dehydrogenase, lactate dehydrogenase-1, potassium, and theophylline assays. Error (expressed in absolute terms) was linearly dependent on hemoglobin concentration and independent of the initial analyte concentration in each case, except for bilirubin and theophylline, where multiple regression analysis was required to quantify the effect. Relative error was dependent on the initial analyte concentration in all cases. Correction formulas were calculated from linear regression of absolute error vs hemoglobin concentration. Clinical application of correction formulas and mechanisms of hemolysis interference for each assay are discussed.

Testicular cancer

Despite the overall cure rate now in excess of 90%, innovation in management of germ cell tumors continues. The report that 80% of patients with extragonadal germ cell tumors have either carcinoma in situ or atrophic tubules as evidence of tumor rejection emphasizes the need to investigate the testis in patients with undiagnosed primary cancer because even today treatment delay is worsening prognosis. The evidence that testicular atrophy is a precursor of malignancy may explain why testis cancer has increased while normal sperm count has fallen over the past 50 years and why there is an association between exposure during service in Vietnam to agents that damage spermatogenesis and development of testis cancer. The improved prognostication from analysis of large databases and salvage with high-dose chemotherapy and bone marrow rescue are giving confidence to explore new innovations, eg, carboplatin instead of cisplatin. In addition, as the database on patients with stage I disease on surveillance enlarges, so does interest in adjuvant chemotherapy, encouraging the search for better markers to predict poor response. Linkage between overproduction of the tumor marker lactate dehydrogenase-1 and the increased copy number of the isochrome 12p in the tumor may be of use in this respect. Reports that germ cell tumor patients exposed to etoposide, eg, leukemic, lung, and ovarian cancer patients, can develop an acute myeloid leukemia with a marker on chromosome 11 are tempering enthusiasm for its use in adjuvant therapy. However, the observation that radiotherapy or chemotherapy may reduce second testis tumor incidence more effectively than surgery does encourages more detailed exploration of the results of adjuvant treatment.

Electrolytes, protein, and lactate dehydrogenase isozymes of parotid saliva in mineralocorticoid-treated rats.

Small quantities of parotid saliva were obtained from 7 of 11 rats in which hypokalemia was induced by treatment with deoxycorticosterone acetate (DOCA). Thirst in the treated rats was indicated by an increase in water consumption. A decrease of 28% in plasma potassium levels indicated that the DOCA-treated rats had become hypokalemic; chloride decreased 6-7% while sodium was unchanged. Saliva showed significant increases in potassium and protein and a decrease in sodium; lactate dehydrogenase-2 and -4 bands appeared darker on the electrophoretic profile. These changes are relevant to the clinical manifestations and diagnosis of hypokalemia.

Dynamic QRS-complex and ST-segment monitoring in acute myocardial infarction during recombinant tissue-type plasminogen activator therapy

Changes of the QRS complex are the electrocardiographic expression of irreversible injury of the myocardium. In humans, the process of infarction occurs over several hours. A more rapid development of QRS changes has been reported in patients treated with thrombolytic agents. Patients with strongly suspected acute myocardial infarction (AMI) included in a placebo-controlled trial of 100 mg of recombinant tissue-type plasminogen activator (rt-PA) were monitored for 24 hours with continuous, on-line vectorcardiography. The magnitude of the QRS vector changes correlated with infarct size estimated by the maximal value of lactate dehydrogenase-1 (r = 0.69, p < 0.001) as well as with left ventricular ejection fraction 30 days after randomization (r = 0.49, p < 0.001). Treatment with intravenous rt-PA limited total QRS vector change but the QRS vector changes observed occurred more rapidly and reached a plateau 131 minutes earlier in patients treated with rt-PA than in those receiving placebo (p < 0.01). A certain pattern of highly variable ST vector magnitude was identified and was associated with higher maximal lactate dehydrogenase-1 values (23 ± 13 vs 14 ± 10 μkat/liter, p < 0.001) and a tendency to higher 1-year mortality (24 vs 9%, p = 0.08) than in patients without this pattern. In patients with this pattern, rt-PA did not affect maximal lactate dehydrogenase-1, time to maximal creatine kinase and final magnitude of QRS vector change.

The predictive value of serum enzymes for perioperative myocardial infarction after cardiac operations: An autopsy study

We evaluated the utility of serum enzyme and isoenzyme activities for detecting autopsy-proved perioperative myocardial infarction in patients who died after cardiac operations. We studied 79 patients who had autopsies performed after coronary artery bypass grafting or valve replacement, or both. Thirty-seven had histologic evidence of a perioperative myocardial infarction. We found statistically significant differences between the group of patients with infarction and the group without infarction when we compared the mean activities of creatine kinase, creatine kinase MB, aspartate aminotransferase, and the lactate dehydrogenase-1/lactate dehydrogenase-2 ratio. The postoperative changes in serum enzymes were analyzed by logistic regression for their relation to perioperative myocardial infarction. Creatine kinase MB exhibited the best diagnostic association with the presence of perioperative myocardial infarction. The lactate dehydrogenase-1/lactate dehydrogenase-2 ratio correlated to a lesser extent with infarction. Adjustment of the diagnostic cutoff to 133 U/L for creatine kinase-MB measured 15 hours after operation yielded a sensitivity of 0.60 and a specificity of 1.0. This study demonstrates that no combination of enzyme activity changes after operation can completely discriminate all patients with perioperative myocardial infarction from those without. Nonetheless, measurement of creatine kinase MB activity provide 96% accuracy for diagnosing infarction at a prevalence of 10%.

Immune response gene control of the mouse antibody responses to human creatine kinase-MM and the lactate dehydrogenase-1 enzymes

We therefore investigated the genetic regulation of the immune response to human CK•MM and LDH-1, using intra-H-2 recombinant mice

Eight independent Ldh-1 mutations of the mouse recovered in mutagenicity experiments: biochemical characteristics and chromosomal localization.

Eight mouse mutants with altered charge or activity of lactate dehydrogenase-1 have been detected in offspring derived from mutagen-treated spermatogonia. Using two chromosome-7 marker genes pooled recombination frequencies are estimated as c-14.4 +/- 0.8-p-6.9 +/- 0.6-Ldh-1.

Quantification of lactate dehydrogenase-1 in serum with use of an M-subunit-specific monoclonal antibody.

We have developed a rapid, one-step assay for measuring lactate dehydrogenase-1 (LD-1) activity in serum after extraction of LD-2, LD-3, LD-4, and LD-5 isoenzymes by an immobilized M-subunit-specific monoclonal antibody (D.8.1). In the assay, 100 microL of serum is mixed with 50 microL of a suspension of 0.8-micron-diameter latex particles coated with 30 micrograms of the monoclonal antibody D.8.1, then incubated at room temperature for 5 min. The latex particles, to which LD-2 through LD-5 are bound, are pelleted by centrifugation for 2 min at 12,000 X g, and the LD-1 activity is measured kinetically in the supernatant fluid. We optimized the assay for antibody immobilization, antibody concentration, and time and temperature of incubation. Serum bilirubin concentrations up to 0.33 g/L (0.56 mmol/L) did not interfere in the assay. Hemolysis interfered solely through LD-1 released from erythrocytes. The within-assay CV for low-concentration quality-control material (LD-1 33 U/L) was 3.5% (n = 9) and for high-concentration material (LD-1 185 U/L) was 1.9% (n = 8); the between-assay CVs for the two materials were 6.1% (n = 9) and 2.5% (n = 10), respectively. The LD-1 activity measured in 98 samples by our assay compared well with a two-step polyclonal antibody-based assay (Isomune-LD, Roche Diagnostic Systems; r = 0.998) and with an electrophoretic method (Paragon, Beckman Instruments; r = 0.956).

Catabolism of circulating enzymes: plasma clearance, endocytosis, and breakdown of lactate dehydrogenase-1 in rabbits.

Lactate dehydrogenase-1 (EC 1.1.1.27), intravenously injected into rabbits, was cleared with first-order kinetics (half-life 27 min), until at least 80% of the injected activity had disappeared from plasma. Radioactivity from injected 125I-labeled enzyme disappeared at this same rate. Trichloroacetic-acid-soluble breakdown products started to appear in the circulation shortly after injection of the labeled enzyme. Body scans of the rabbits for 80 min after injection of 131I-labeled enzyme revealed rapid accumulation of label in the liver, peaking 10-20 min after injection. Subsequently, activity in the liver declined and radioactivity (probably labeled breakdown products of low molecular mass) steadily accumulated in the bladder. Tissue fractionation of liver, 19 min after injection of labeled enzyme, indicated that the radioactivity was present both in endosomes and in lysosomes, suggesting uptake by endocytosis, followed by breakdown in the lysosomes. Measurements of radioactivity in liver and plasma suggest that the liver is responsible for the breakdown of at least 75% of the injected enzyme. Radioautography of tissue sections of liver and spleen showed accumulated radioactivity in sinusoidal liver cells and red pulpa, respectively. These results are very similar to those for lactate dehydrogenase-5, creatine kinase MM, and several other enzymes that we have previously studied in rats.

Analysis of recombination in the centromere region of mouse chromosome 7 using ovarian teratoma and backcross methods.

Recombination near the centromere of mouse chromosome 7 was studied using data obtained from ovarian teratomas and backcrosses. The recombination percentage for the centromere-Gpi-1 (glucose phosphate isomerase-1) interval was 13.4 +/- 2.6 using the ovarian teratoma mapping method. In a backcross using the Robertsonian translocation Rb(7.18)9Lub (Rb9) as the centromeric marker, the centromere-Gpi-1 recombination percentage was 4.5 +/- 1.3, demonstrating that Rb9 suppresses recombination near the centromere of chromosome 7. The recombination percentage for the Gpi-1-Ldh-1 (lactate dehydrogenase-1) interval was estimated on the LT/Sv mouse genetic background to be 19.0 +/- 2.9 using the ovarian teratoma mapping method, a value comparable to the 15.5 +/- 4.8 reported earlier. On the same genetic background in a backcross segregating for Rb9, the Gpi-1-Ldh-1 recombination percentage was 7.1 +/- 1.6. Another backcross, without the Rb9 translocation but utilizing a different genetic background, produced a recombination percentage for the Gpi-1-Ldh-1 interval of 10.7 +/- 1.5, a value similar to that obtained in the Rb-containing cross. These results suggest that either the recombination suppression in the centromere area caused by Rb9 does not extend to the Gpi-1-Ldh-1 genetic region or, if it does, that the differing genetic backgrounds of these two crosses influence recombination. No recombinants were detected among 410 offspring produced from a backcross mating segregating for Ldh-1 and ru-2 (ruby-eye-2). Thus, the gene order of Ldh-1 and ru-2 on chromosome 7 remains uncertain.

Information induction for predicting acute myocardial infarction.

We show how to make an unsupervised discrimination of disease and nondisease states by measuring information and using newer notions of inductive reason. We also present a new theory of group-based reference values that is based on measuring information uncertainty. We use data on the isoenzymes creatine kinase-MB (CK-MB) and lactate dehydrogenase-1 (LD1) and on the percentage of LD1 from 101 patients with acute myocardial infarction (AMI) and from 41 patients with suspected, but unfounded, infarction (non-AMI). Calculating the Shannon entropy, a concept from information theory, of the data base allows determination of a difference in entropy values ("effective information"), which determines decision cutoff values that produce binary-base patterns yielding the fewest classification errors. Redundancy in testing is important because it provides the information to approach a goal of errorless discrimination by coding the test results and meeting the conditions of the "Noisy Channel Theorem" of information theory. This redundancy improves the predictive value of diagnosis by isolating the area of equivocation to evident patterns. Results for CK-MB and LD1 are 99% correct in assigning cases to AMI and non-AMI categories; adding %LD1 increases the proportion of errorless binary patterns from 25% to 90%.

Determination, by radioimmunoassay, of the mass of lactate dehydrogenase isoenzyme 1 in human serum and of its rate of removal from serum after a myocardial infarction.

In this radioimmunoassay of lactate dehydrogenase-1 (LD-1; EC 1.1.1.27) in human serum we use a commercial LD-1-selective assay system and a goat antiserum. We have determined the fractional rate of disappearance from serum and the half-life of LD-1, in terms of both enzyme activity and enzyme mass, in 21 myocardial infarction patients. Our evidence suggests that this isoenzyme is inactivated in serum. Furthermore, our data suggest that the conventionally accepted half-life of about 110 h for serum LD-1 activity may grossly overestimate the actual LD-1 half-life in many post-myocardial infarction patients.

Quantitation of serum lactate dehydrogenase-5 with monoclonal antibodies

Spleen cells from BALB/cJ mice which had been immunized with human lactate dehydrogenase-1 (LDH-1) were fused with SP2/0-Ag14. Two hybridomas were produced which recognized the antigen. Competitive RIA revealed that one antibody (‘Smit-LDH’) recognized the H subunit of LDH while the other (‘Hem-LDH’) recognized both H and M subunits of LDH. With the use of these antibodies we developed an assay for LDH-5 activity in which serum is incubated for 30 min at room temperature with the two antibodies (‘Smit-LDH’ and ‘Hem-LDH’ in the ratio 64:1.3, μg/ml) immobilized on latex beads to extract LDH-1 through LDH-4. After centrifugation, the LDH activity of the supernatant is measured and represents LDH-5 activity. Latex beads coated with bovine serum albumin were used as control. The LDH-5 activity as determined by our assay correlated well ( r = 0.98) with the values obtained by an electrophoresis method. There was no interference due to LDH-1 through LDH-3 up to 3000 U/l and LDH-4 up to 350 U/l. Serum samples with total LDH activity above 1000 U/l were appropriately diluted in order to avoid interference by LDH-4. Use of these monoclonal antibodies allows precise, rapid and direct measurement of LDH-5 activity in serum.

Genetic analyses of electrophoretic enzyme variants, mating type, and hermaphroditism in Pyricularia oryzae Cavara

Pyricularia oryzae (teleomorph: Magnaporthe grisea) parasitizes a variety of gramineous hosts and causes the rice blast disease worldwide. Through matings among P. oryzae isolates from rice, finger millet, and weeping lovegrass the inheritance of electrophoretic variants of six enzymes, phosphoglucomutase (PGM), phosphoglucose isomerase (PGI), glycerate-2-dehydrogenase (G2DH), malate dehydrogenase-3 (MDH-3), lactate dehydrogenase-1 (LDH-1), and lactate dehydrogenase-3 (LDH-3) was determined. All six variants were under single gene control as determined by tetrad and random spore analysis. However, at Ldh-3 and Mdh-3, there were consistent excesses of variant alleles among ascospore segregants. Preliminary data on the genetic control of hermaphroditism suggested that maleness in two Japanese rice isolates might be due to a single gene mutation. Linkage analyses among the six electrophoretic markers, mating type, and hermaphroditism suggested loose linkage between Pgm and G2dh with a recombination frequencies of 43.0%.Key words: linkage, Magnaporthe grisea, rice blast fungus.

Linkage of lactate dehydrogenase-2, Ldh-2, in the mouse.

An electrophoretically detectable variant of lactate dehydrogenase-2 in Mus musculus has been found and used to locate the structural gene, Ldh-2, on chromosome 6. Gene order and recombination frequencies are estimated as Sig--36.0 +/- 4.8--Lc 21.0 +/- 4.1--Miwh--20.0 +/- 4.0--Ldh-2.

Enzyme inactivation and inhibition by gossypol.

Three lactate dehydrogenase isozymes and malate dehydrogenase purified from mouse tissues were inactivated with time by low concentration of gossypol. The degree of enzyme inactivation is both gossypol- and enzyme-concentration-dependent. Under the same experimental conditions, lactate dehydrogenase-X and lactate dehydrogenase-5 were inactivated faster than lactate dehydrogenase-1. NADH was shown to partially protect the enzymes against inactivation by gossypol. The results of this study suggest that the enzymes are inactivated by the minor components in gossypol preparations. Isozymes of glutathione S-transferases were reversibly inhibited by gossypol. The inhibition of transferases by gossypol was shown to be competitive with respect to the 1-chloro-2,4-dinitrobenzene. It is proposed that the male antifertility effect of gossypol may be related to the selective inactivation of sperm-specific lactate dehydrogenase-X.