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Articles |
Department of Clinical Pathology, University of Ulsan College of Medicine and Asan Medical Center, 388–1 PoongNap-Dong SongPa-Gu, Seoul 138–736 Korea.
wkmin{at}amc.ulsan.ac.kr
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Transient increases in concentrations of Lp(a) noted in tumor or postoperative patients have led some, but not all, researchers to conclude that Lp(a) is an acute-phase reactant (APR). Kawade et al., who observed a transient increase in serum Lp(a) concentrations among postoperative patients, reported that Lp(a) was one of the APRs (15). Maeda et al. reported a transient twofold increase in concentrations among acute myocardial infarction patients and postoperative patients (16). Noma et al. observed that relative Lp(a) values (relative to the basal values) averaged 4 times higher among acute myocardial infarction patients and 2.5 times higher among postoperative patients (17). By contrast, Slunga et al., in a study of 32 acute myocardial infarction patients, found no clear evidence of Lp(a) as an APR (18).
The question of whether Lp(a) is an APR is particularly important with regard to Lp(a) as a CHD risk factor in patients with an acute-phase response (referred to as APR patients). If Lp(a) is an APR, then serum Lp(a) concentrations in APR patients can be expected to be increased and may reach an amount that would confound risk analysis.
The purpose of this study was to investigate whether Lp(a) is an APR that can cause substantial bias in risk factor analysis for CHD among APR patients. To determine if serum Lp(a) concentrations increased concurrently with an acute-phase response, we compared the serum Lp(a) concentrations and apo(a) phenotypes in 100 controls and in a random sampling of 100 APR patients.
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Materials and Methods |
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To serve as controls, 100 age- and sex-matched subjects were randomly selected from among healthy blood donors without findings of acute inflammatory reaction. All in the control group were free of the clinical, biochemical, or hematological manifestations of cardiovascular, hepatic, renal, or endocrinologic disorders.
Among the controls, the male-to-female ratio was 1:0.85; the age distribution was 45.9 ± 14.4 (mean ± SD) years. Among the APR patients, the male-to-female ratio was also 1:0.85; age distribution was 46.4 ± 15.3 years.
We obtained all specimens from our laboratory after all requested diagnostic testing had been completed. Clinical specimens were used in accordance with the policies of the Institutional Review Board of Asan Medical Center.
analytical methods
Serum Lp(a) was determined with a commercially available one-step
sandwich Immunozym Lp(a) kit (Immuno, Vienna, Austria). All steps in
the test procedure were performed exactly as prescribed by the
manufacturer. Because the minimal detectable concentration of Lp(a) was
0.01 g/L, results below that were recorded as 0.01 g/L. The mean
intraassay CV was 3.2%; interassay, 6.3%.
Apo(a) isoforms were separated with a commercial 4–15% gradient PhastGel and a PhastSystem (both from Pharmacia Biotech). Immunodetection was performed with a Lp(a) phenotype kit (Immuno). First, 10 µL of the sample was mixed with 5 µL of mercaptoethanol and 85 µL of reducing agent and reduced for 5 min at room temperature. Next, 1 µL of pretreated sample was applied to a gel and electrophoresed for 70 min at Vmax = 250 V, Imax = 10 mA, P = 3 W, and 15 °C. The separated proteins were blotted from the gel to a nitrocellulose membrane by diffusion at 70 °C for 1 h. The membrane was allowed to react overnight at room temperature with 1:500-diluted polyclonal anti-human Lp(a) in Tris-buffered saline (TBS) containing skim milk, 10 g/L, after blocking with TBS containing skim milk at 30 g/L. The membrane was washed twice in TBS containing 10 g/L skim milk and incubated with 1:500-diluted rabbit anti-sheep IgG–alkaline phosphatase conjugated at room temperature for 1 h. After one washing with 10 g/L skim milk in TBS, the nitrocellulose membrane was equilibrated in 0.1 mol/L Tris buffer (pH 9.5) and immersed in alkaline phosphatase developing solution until the bands became clearly visible. After washing in distilled water, the membranes were dried and kept for documentation.
The apo(a) phenotypes are designated as F, B, S1, S2, S3, S4, and S5; their respective numbers of kringle IV repeats are 11–13, 14–16, 17–19, 20–22, 23–25, 26–28, and 29–42 (19). Isoforms were interpreted by comparison with an apo(a) phenotype standard with known kringle IV number, which was included in an Lp(a) phenotype kit. The apo(a) phenotype standard consisted of B, S1, S3, S4, and S5 isoforms, with kringle IV numbers of 14, 19, 23, 27, and 35, respectively.
ESR was measured by a modified Westergren method.
Statistical analyses were performed with the Statistical Analysis System (SAS Institute). More than two groups were compared by the Kruskal–Wallis one-way ANOVA test. The chi-square test of association was performed to elucidate an independent relationship between the variables.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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). Moreover, the medians and
interquartile ranges of serum Lp(a) in the APR patients were shifted to
a higher value than that for controls (Table 1
). The mean serum Lp(a) concentration (±SD) of APR patients was
0.300 ± 0.284 g/L, whereas that of the controls was 0.118 ±
0.193 g/L (P <0.0001). Among the groups of APR patients,
the Lp(a) concentration was more than double that of the controls: The
mean serum Lp(a) concentration was 0.262 ± 0.266 g/L
(P <0.001) in patients with infections; 0.288 ± 0.266
g/L (P <0.001) in postoperative patients; 0.359 ±
0.288 g/L (P <0.0001) in patients with tumors; and
0.274 ± 0.431 g/L (P <0.05) in patients with other
diseases.
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Among the 100 APR patients, 41% had an Lp(a) concentration >0.30 g/L, whereas among the 100 controls only 11% had concentrations that great.
distribution of apo(a) phenotypes and serum lp(a) concentrations
We found no significant difference between the controls and the
APR patients regarding the distribution of apo(a) phenotypes
(P >0.05) (Table 2
). The most frequently occurring phenotype in the two groups was
S5 (41.0%), followed by S4S5 (20.0%), S5S5 (14.0%), S4 (8.5%), and
the remaining phenotypes (16.5%).
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However, significant differences between controls and APR patients were
found for the medians and interquartile ranges of serum Lp(a) among the
frequently observed apo(a) phenotypes S5, S4S5, S5S5, and S4 (Table 3
). Among these phenotypes, the mean Lp(a) concentration was 3.1
to 4.8 times higher in APR patients than in the controls. The mean
serum Lp(a) concentration (±SD) in controls with the S5 phenotype was
0.050 ± 0.097 g/L, whereas that among APR patients was 0.240
± 0.282 g/L (P <0.001). The mean serum Lp(a)
concentrations (±SD) in S4S5, S5S5, and S4 phenotypes among the
controls were 0.148 ± 0.184, 0.062 ± 0.053, and 0.064
± 0.094 g/L, respectively, whereas those of APR patients were
0.462 ± 0.319 (P <0.001), 0.265 ± 0.202
(P <0.01), and 0.284 ± 0.252 g/L (P
<0.05), respectively.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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To confirm the increase of Lp(a) values among APR patients, we randomly selected 24 samples from the APR patients and analyzed them with a second assay, TintElize Lp(a) (biopool, Umea, Sweden). The mean serum Lp(a) concentration (±SD) of these 24 samples by the Immunozym Lp(a) kit (x) was 0.331 ± 0.347 g/L; that by the TintElize Lp(a) kit (y) was 0.353 ± 0.436 g/L. The regression equation for the two methods was y = 1.202x - 0.04 (r = 0.94; Sy|x =0.13).
We also found that the frequency distribution of Lp(a) concentration in the APR patients was shifted substantially to the right of that of the controls. Whereas only 11% of the controls had serum Lp(a) concentrations >0.30 g/L (the value generally accepted as an independent risk factor for CHD (10)(19)(20)), 41% of the APR patients did. If the Lp(a) samples were limited to those for which the ESR was >50 mm/h, the positive risk rate for CHD could be falsely increased by as much as 3.7-fold (41%/11%).
In most previous studies of serum concentrations of Lp(a) as a CHD risk factor, the clinical conditions of the patients were not described. However, the patients who are likely to have their CHD risk analyzed by an Lp(a) test are those who have had acute myocardial infarction, have undergone bypass surgery, or have infection. In these cases (i.e., for patients in acute-phase states), Lp(a) should not be used in a risk analysis because of positive bias.
Maeda et al. (16) compared various APRs in acute
myocardial infarction patients and postoperative patients. The serum
concentration of C-reactive protein reached its peak 2–3 days after
surgery; 
1-antitrypsin, 4 to 5 days;
1-acid glycoprotein, 5 to 6 days; haptoglobin, 6 to 9
days. But the serum concentration of Lp(a) in acute myocardial
infarction patients reached its peak after 10.4 ± 1.9 days and in
postoperative patients 6.5 ± 2.3 days after surgery.
As a preliminary study, we observed for 10 days after surgery the Lp(a), C-reactive protein, and ESR results of 8 patients who had undergone gastrectomy. C-reactive protein reached a peak concentration on the 2nd day after surgery, ESR on the 4th day, and Lp(a) on the 6th day, paralleling the findings of Maeda et al. (16). C-reactive protein had normalized by the 10th day, but ESR and Lp(a) were still above normal on the 10th day.
Because in this preliminary study the increment patterns of ESR and
Lp(a) were similar, we chose an ESR value of 50 mm/h, twice the upper
limit in healthy subjects, as an arbitrary cutoff value for acute
inflammatory state. In a previous study, the response patterns of APRs
and Lp(a) were found to differ, with no or weak correlations
(21). In this study, however, we observed no significant
correlation between serum Lp(a) concentration and ESR
(r = 0.02), but this might be due in part to the high
ESR values in our chosen group. Ledue et al. (21) have
suggested complement component 4 and 1-acid glycoprotein
as candidate criteria for excluding an acute inflammatory reaction
before measuring Lp(a). Because Lp(a) concentrations have shown
persistent increases in APR patients for as long as 20 days
(17) and no, or weak, correlation with other APRs, further
study is needed to elucidate the relationship of Lp(a) to other APRs.
The transient increase in serum Lp(a) concentrations during acute
inflammatory states is thought to be caused by increased Lp(a)
synthesis or reduced removal of Lp(a) or by a change in the
distribution of Lp(a) particles between the intravascular and
extravascular compartments. Maeda et al. (16) suggested
that during acute inflammatory states the synthesis of Lp(a), whose
sialic acid content is 6 times that of LDL, increases in the liver with
a concurrent increase of 1-acid glycoprotein,
1-antitrypsin, haptoglobin, and fibrinogen, which also
have high contents of sialic acid. Kawade et al. (15)
reported that patients whose Lp(a) concentration reached a peak on the
5th to 10th day after surgery and then returned to the initial value in
1 week had a good prognosis, whereas those who did not experience the
transient increase of Lp(a) had a poor prognosis. These findings could
be interpreted to mean that Lp(a) played an important role in the
patients' recovery from the injuries of surgery. The idea that
increased serum Lp(a) plays an important role in tissue recovery from
injury, especially angiogenesis, was prompted by Noma et al.
(17), who proved immunohistochemically that anti-apo(a)
antibody stained along the capillaries in the specimens from skins of
psoriasis vulgaris patients, in the healing area of gastric ulcer, and
in the peritumorous area of gastric cancer. It has also been reported
that Lp(a) rendered a lipid pool saved from LDL receptor degradation to
rapid cell regeneration, active membrane biogenesis, or an acute
inflammatory process (22), but the role of increased Lp(a)
in relation to acute inflammatory states was not fully documented.
On the basis of our findings of increases in serum Lp(a) concentrations among APR patients, we recommend avoiding the use of Lp(a) as a measurement of risk for CHD in such patients. Only after excluding the possible effects of the acute inflammatory state should Lp(a) concentrations be used to calculate risk for CHD.
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Acknowledgments |
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Footnotes |
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The following articles in journals at HighWire Press have cited this article:
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  C. H. Saely, T. Marte, H. Drexel, C. P. Kovesdy, A. A. Ariyo, R. P. Tracy, and C. Thach Lp(a) Lipoprotein, Vascular Disease, and Mortality in the Elderly N. Engl. J. Med., March 11, 2004; 350(11): 1150 - 1152. [Full Text] [PDF]
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 D J Byrne, I A Jagroop, H E Montgomery, M Thomas, D P Mikhailidis, N G Milton, and A F Winder Lipoprotein (a) does not participate in the early acute phase response to training or extreme physical activity and is unlikely to enhance any associated immediate cardiovascular risk J. Clin. Pathol., April 1, 2002; 55(4): 280 - 285. [Abstract] [Full Text] [PDF]
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 C. A. Glader, B. Stegmayr, J. Boman, H. Stenlund, L. Weinehall, G. Hallmans, and G. H. Dahlen Chlamydia pneumoniae Antibodies and High Lipoprotein(a) Levels Do Not Predict Ischemic Cerebral Infarctions : Results From a Nested Case-Control Study in Northern Sweden Stroke, October 1, 1999; 30(10): 2013 - 2018. [Abstract] [Full Text] [PDF]
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