Obstetrics & Gynecology:
Seroprevalence of Antibodies to Chlamydia pneumoniae in Women With Preeclampsia
Heine, R. Phillip MD; Ness, Roberta B. MD, MPH; Roberts, James M. MD
Duke University Medical Center, Durham, North Carolina; and University of Pittsburgh, and Magee-Womens Research Institute, Pittsburgh, Pennsylvania.
Address reprint requests to: Roberta B. Ness, MD, MPH, University of Pittsburgh, Graduate School of Public Health, Room 517, Parran Hall, 130 DeSoto Street, Pittsburgh, PA 15261; E-mail: email@example.com.
Funding source: P01 HD30367 from the National Institute of Child Health and Human Development.
Received May 20, 2002. Received in revised form August 9, 2002. Accepted August 15, 2002.
OBJECTIVE: Preeclampsia shares many risk factors and pathophysiologic features with coronary heart disease. We studied whether, like atherosclerosis, preeclampsia is related to seroprevalence of immunoglobulin (Ig) G antibodies to Chlamydia pneumoniae.
METHODS: Cross-sectional comparisons were made for 37 women with preeclampsia and 37 women with normal pregnancies at term. In these two groups, antibody titers for IgG, IgM, and IgA seroprevalence to C pneumoniae and IgG to Chlamydia trachomatis and Chlamydia psittaci were compared.
RESULTS: Immunoglobulin G antibodies to C pneumoniae at a titer of at least 1:16 were more common in women with preeclampsia (25 of 37) than in women without (15 of 37) (odds ratio 3.1; 95% confidence interval 1.2, 7.9). There were no significant differences in the seroprevalence of IgA or IgM antibodies to C pneumoniae. Women with preeclampsia were also no more likely to have IgG antibodies to C trachomatis or C psittaci.
CONCLUSION: Women with preeclampsia had an increased IgG seroprevalence to C pneumoniae but not to C trachomatis or C psittaci. These preliminary data suggest a specific association between infection with C pneumoniae and preeclampsia.
Preeclampsia and coronary heart disease share many common pathophysiologic features and risk factors. One common feature is endothelial dysfunction, which may derive from inflammation. In atherosclerosis, injury-induced mononuclear cell accumulation, migration and proliferation of smooth muscles, and formation of fibrous tissue ultimately lead to plaque formation and vessel obstruction.1 These pathologic features, plus the finding that an elevation of inflammatory markers precedes atherosclerosis,2 have suggested an inflammatory origin to the altered endothelial dysfunction seen in atherosclerosis. In preeclampsia, too, heightened intra-vascular inflammation has been suggested as the origin of the abnormal endothelial function.3,4
Accumulating evidence suggests that infection by Chlamydia pneumoniae, a common pathogen causing upper and lower respiratory tract disease, may be involved in the endothelial injury that marks atherosclerosis. Chlamydia pneumoniae is prevalent in atherosclerotic lesions but generally absent in normal coronary arteries.5 It induces atherosclerotic lesions in mice and rabbits,6–8 particularly in association with hypercholesterolemia. Further, in some studies, but not all, C pneumoniae antibodies are more common in individuals with myocardial infarction and coronary heart disease death.9–18 Finally, recent studies among patients with coronary artery syndromes have shown that, shortly after their initiation, macrolides reduce the risk of subsequent adverse cardiac events.16–18
In this study we explored the relationship between C pneumoniae seroprevalence and preeclampsia. We assessed the occurrence of immunoglobulin (Ig) G, IgM, and IgA antibodies to C pneumoniae and IgG antibodies to Chlamydia trachomatis and Chlamydia psittaci among women with preeclampsia and pregnant controls.
MATERIALS AND METHODS
Serum samples were obtained as part of an ongoing study of preeclampsia that has been approved by the Magee-Womens Hospital Institutional Review Board.19 Samples were obtained at the time of admission for labor and delivery, processed within 2 hours, and stored at −70C. Samples were randomly selected from 37 nulliparous pregnant women with preeclampsia and from 37 nulliparous pregnant women with uncomplicated, term pregnancies.
Preeclampsia was defined using the criteria of hypertension, proteinuria, hyperuricemia, and resolution of hypertension and proteinuria after pregnancy. Hypertension was defined as an increase of 30 mm Hg systolic or 15 mm Hg diastolic blood pressure as compared with values obtained before 20 weeks' gestation, or an absolute blood pressure of 140/90 mm Hg or higher after 20 weeks' gestation if earlier blood pressure values were unknown.20 Among the 27 preeclamptic patients, all had a blood pressure value greater than 140 mm Hg systolic or 90 mm Hg diastolic. Proteinuria was defined as 300 mg per 24-hour collection, or more than 2+ on a voided or 1+ on a catheterized random urine specimen. Hyperuricemia was defined as more than one standard deviation above the usual for the gestational age at which the sample was obtained (5.5 mmol/L at term). All abnormalities had to return to normal by 12 weeks postpartum for the patient to be considered preeclamptic.
Sera from preeclamptic and control women were sent in batches to MRL Reference Laboratory (Cypress, CA) for the detection of antibodies to C pneumoniae using an established microimmunofluorescence technique for detection of antibodies.21 Briefly, antigen was fixed to a slide and probed with patient sera. Antibody specific to IgG, IgM, and IgA was fluoroscein labeled and used as a secondary probe. Positive samples were diluted to determine the absolute antibody titer. To rule out the presence of cross-reactive antibody, testing was also performed for C trachomatis and C psittaci. For all organisms a titer of at least 1:16 was considered positive for IgG and IgA, where as a titer of at least 1:10 was considered positive for IgM.
Covariate information was obtained by review of the maternal and neonatal medical records at the time of labor and delivery. Information was obtained on maternal age, race (white versus nonwhite), current smoking, gestational age at delivery, and infant birth weight.
We evaluated data on demographic and clinical features and serologies among the 37 preeclamptic women and 37 controls. Sample size analyses suggested that with 17–45 cases and the same number of controls we could detect a difference between a prevalence of 10–20% in controls and 50–60% in cases, the magnitude of difference initially observed for seroprevalence of C pneumoniae among subjects with and without coronary artery disease.9 This calculation and logistical considerations led us to our sample size.
Odds ratios were the main measure used for comparison of preeclamptic patients and controls. Cross-sectional differences in covariates were analyzed. The proportions of women with a positive serum test at a titer of at least 1:16 (for IgG and IgA) and at least 1:10 (for IgM) were then compared, first using the Fisher exact test and then odds ratios. Subsequently, a series of interval antibody titers to C pneumoniae were used to compare the seroprevalence rates between cases and controls, using odds ratios as the measure of comparison. Logistic regression analyses were used to adjust for age and then for gestational age, considering preeclamptic versus control status as the main dependent variable and antibody presence as the main independent variable.
The 37 women with pregnancies complicated by preeclampsia were not significantly different from the 37 with unaffected term pregnancies with respect to age, race, or smoking status (Table 1). As expected, women with preeclampsia were significantly more likely to deliver preterm.
The seroprevalence of IgG to C pneumoniae at a titer of at least 1:16 was more common in women with preeclampsia (25 of 37) than in controls (15 of 37) at a significance level of P < .05 (Figure 1). Positive titers for C pneumoniae IgM and IgA were not significantly more likely among preeclamptic women. Women with preeclampsia, as compared with controls, also had no greater seroprevalence of IgG to C trachomatis or C psittaci (Figure 2).
We further explored the possibility that higher titers of IgG antibodies to C pneumoniae might reflect higher risk (Table 1). At a cutoff of 1:16, the risk of seropositivity to C pneumoniae was 3.1 (95% confidence interval 1.2, 7.9). Odds ratios for antibody titers of 1:16 or 1:32, 1:64, and at least 1:128 were 4.6, 3.1, and 2.6, respectively, indicating a lack of higher risk with higher titers. Given the smaller sample sizes involved in these subset analyses, none of the associations between specific antibody titer cutoffs and preeclampsia were significant. Age adjustment did not substantially alter these results (data not shown). Gestational age adjustment also had little impact on the odds ratios but widened the confidence intervals because gestational age data were missing for a third of study women.
We found that IgG seroprevalence to C pneumoniae was more common among women with preeclampsia than among women with unaffected term pregnancies. The fact that we did not find an association to preeclampsia for IgG seroprevalence to C trachomatis or C psittaci suggests a specific association between preeclampsia and the particular species of Chlamydia that has been associated with atherosclerosis. This finding is consistent with the known lack of cross-reactivity of the immunofluorescent assay used.21 Altogether, it is unlikely that cross-reactivity to other pathogens might explain the relationship found between preeclampsia and C pneumoniae.22
An association to preeclampsia was not found for IgA or IgM seroprevalence to C pneumoniae. Neither did we find a dose-response relationship between IgG titer and risk of preeclampsia. Both of these findings suggest that it is past, persistent, or chronic active infection and not acute infection or reinfection with C pneumoniae that is associated with preeclampsia. Acute infection or reinfection would cause an elevation in IgM titers. Further, studies of paired sera obtained from patients with pneumonia indicate that acute infection generally causes high titers (1:512 or 1:1024).21 In animal models, the induction of atherosclerosis has required persistent or chronic active infection, rather than a single past exposure to C pneumoniae. Repeated respiratory infection with C pneumoniae, and not a single, acute infection, is needed to promote the growth of atherosclerotic plaque in rabbits. Antibiotic treatment can prevent or reduce atherosclerotic changes in animals being reinfected with C pneumoniae.23 Human serological data suggest that recent reinfection also may be important, at least in relation to myocardial infarction or coronary heart disease death. Although prospective observational studies are more mixed than case-control studies24 in showing an association between C pneumoniae and coronary heart disease,10–15 two prospective observational studies that did show an association related recent antibody positivity and high titers of C pneumoniae to newly manifest coronary heart disease.12,14 There is some, limited evidence that antibiotic use in pregnancy may reduce the risk of preeclampsia.25
How might C pneumoniae increase the risk of preeclampsia? Preeclampsia can be considered a two-stage disorder, with the first stage involving abnormal placental implantation and the second being the abnormal maternal adaptation to the pregnancy that results in systemic dysfunction.26 In healthy pregnancies, the maternal spiral arteries undergo extensive remodeling.27 This remodeling is incomplete in preeclampsia. Reduced placental perfusion, the outcome of abnormal spiral artery remodeling, is considered to be the root cause of preeclampsia. Further, in preeclamptic spiral arteries a frequent histological finding thought to contribute to reduced placental perfusion is atherosis, a pathologic lesion involving the same lipid-laden foam cells observed in atherosclerosis. Chlamydia pneumoniae may contribute to atherosclerosis by bringing monocytes to areas of arterial damage to form foam cells, leading to plaque, and thrombosis.28 Similarly, past, persistent, or recurrent infection with C pneumoniae may contribute to abnormal vascular function, atherosis, and the abnormal placental perfusion that marks the first stage of preeclampsia.
Chlamydia pneumoniae may also contribute to the second stage of preeclampsia through endothelial dysfunction.4 Overweight, diabetes, hyperlipidemia, and hypertension are all established risk factors for coronary heart disease and, when present in women entering pregnancy, greatly enhance their likelihood of developing preeclampsia.29 The pathophysiology shared by all of these risk factors is endothelial dysfunction. Inflammation also increases endothelial dysfunction by enhancing oxidative stress.30 Markers of inflammation independently predict incident coronary artery disease, and a predisposition to a muted inflammatory response reduces the risk of coronary artery disease.2,31 Such direct temporal relationships have yet to be demonstrated for preeclampsia, but inflammatory mediators are elevated in it.3 Chlamydia pneumoniae may, in both coronary heart disease and preeclampsia, be an initiator of the inflammatory process. Alternatively, C pneumoniae may simply reside on monocytes activated during inflammation.24 If so, the association between C pneumoniae and coronary heart disease or preeclampsia may be an effect rather than a cause.
We recognize that weaknesses of our study were its small size and cross-sectional design. Further, although race did not significantly differ between cases and controls, nonwhite race was more common in patients and may have confounded our results. Our study also had strengths. Because our case and control selection was embedded in a large study of preeclampsia, we were able to approach a large proportion of eligible women. Thus, selection of cases and controls was relatively unbiased. That the prevalence of C pneumoniae in this study is similar to that seen in young adults in Europe reassures us as to the generalizability of our finding.32 Further, we used a rigorous case definition for preeclampsia and well-established methods for the serological detection of C pneumoniae. However, based on this single report in a modest-sized sample, it would be premature to consider our observed link between C pneumoniae and preeclampsia to be other than an association in need of replication.
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