Inflammatory Cytokines and Spontaneous Preterm Birth in Asymptomatic Women

A Systematic Review
Shu-Qin Wei,
MD, PhD,

William Fraser,

MD, MSc,

and Zhong-Cheng Luo,

MD, PhD

OBJECTIVE: To estimate the association between inflammatory cytokines and the risk of spontaneous preterm birth in asymptomatic women. DATA SOURCES: We searched electronic databases of the human literature in PubMed, EMBASE, and the Cochrane Library up to February 2010 using the following key words: preterm/pre-term (birth/delivery) and cytokine or inflammation/inflammatory marker/ biomarker. METHODS OF STUDY SELECTION: We included observational studies that reported the association between common inflammatory cytokines and spontaneous preterm birth as an outcome in asymptomatic women. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated using fixed and random effects models. TABULATION, INTEGRATION, AND RESULTS: Seventeen primary studies comprising 6,270 participants met the inclusion criteria. Spontaneous preterm birth was strongly associated with increased levels of interleukin-6 (IL-6) in midtrimester cervicovaginal fluid (OR 3.05, 95% CI 2.00 4.67) (number needed to treat 7 for identifying an additional preterm delivery) and amniotic fluid (OR 4.52, 95% CI 2.677.65) (number needed to treat 7), but
From the Department of Obstetrics and Gynecology, Sainte-Justine Hospital, University of Montreal, Quebec, Canada. Supported by a grant from the Canadian Institutes of Health Research (CIHR grant 154107 to Z.-C.L.). S.-Q.L. is supported by a Scholarship from the CIHR Strategic Training Initiative in Research in Reproductive Health Sciences (STIRRHS). Z.-C.L. is supported by a Junior Scholar award from the Quebec Foundation of Health Research (FRSQ) and a CIHR New Investigator award and W.D.F. by a CIHR research chair award. Presented at the 30th Annual Meeting of the Society for MaternalFetal Medicine (SMFM), Chicago, Illinois, February 5, 2010. Corresponding author: Zhong-Cheng Luo, MD, PhD, Department of Obstetrics & Gynecology, Bureau 4986B, CHU Sainte-Justine Universite de Montreal, 3175 Cote Sainte-Catherine, Montreal, Quebec, Canada H3T 1C5; e-mail: zhong-cheng.luo@recherche-ste-justine.qc.ca. Financial Disclosure The authors did not report any potential conflicts of interest. 2010 by The American College of Obstetricians and Gynecologists. Published by Lippincott Williams & Wilkins. ISSN: 0029-7844/10

there was no association in plasma specimen (OR 1.5, 95% CI 0.73.0). Spontaneous preterm birth was strongly associated with increased C-reactive protein (CRP) levels in midtrimester amniotic fluid (OR 7.85, 95% CI 3.88 15.87) (number needed to treat 3), but the association was weak in plasma specimen (OR 1.53, 95% CI 1.221.90). There were insufficient data (fewer than three studies) for meta-analysis in other inflammatory cytokines. CONCLUSION: Inflammatory cytokine IL-6 in cervicovaginal fluid and IL-6 and CRP in amniotic fluid but not in plasma are strongly associated with spontaneous preterm birth in asymptomatic women, suggesting that inflammation at the maternalfetal interface, rather than systemic inflammation, may play a major role in the etiology of such spontaneous preterm births.
(Obstet Gynecol 2010;116:393401)

reterm birth is defined as delivery before 37 complete weeks of gestation and is the event most responsible for poor perinatal outcomes in developed countries.1 It accounts for approximately three fourths of all neonatal deaths and approximately half of long-term neurologic disability.2,3 There remains no effective means to prevent preterm birth. An important public health concern is the increase in preterm delivery rates over the last two decades in many developed countries.4 Therefore, one of the most urgent problems in perinatology is to identify women at high risk for preterm birth and develop appropriate preventive strategies.3 Further reductions of perinatal mortality rates, the costs of neonatal care, and subsequent physical and neurological impairments among preterm infant survivors depend on reducing preterm delivery rates.5 The etiology of preterm birth is complex. Accumulated research data indicate that infection and inflammation (most often subclinical) might be an important underlying cause. A growing body of evidence suggests a role for cytokines in the mechanisms

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responsible for preterm birth.6 Improved understanding of the link between cytokines and spontaneous preterm birth may provide an exciting potential for the development of sensitive new markers to identify women at risk as well as effective interventions. There remains a lack of systematic reviews on the roles of inflammatory cytokines in the etiology of spontaneous preterm birth among asymptomatic women. The current systematic review and meta-analysis was aimed to summarize the evidence on common inflammatory cytokines in various biologic fluids in relation to the risk of spontaneous preterm birth in asymptomatic women.

Box 1. Quality Assessment of Observational Studies (total 10 points)*

Selection of participants (1/0) Cohort studies (1/0) Selected cohort was representative of the general population (population-based studies) or target catchment population (hospital-based studies) (1) Cohort was a selected unrepresentative group (0) Case control studies (1/0) Cases and control subjects drawn from the same population (1) Cases and control subjects drawn from different sources or the selection of groups (0) Comparability of groups (2/0) No significant differences between the groups reported in terms of age, plurality, smoking, preterm birth history, or preexisting medical conditions were explicitly reported, unless it was one of these variables under investigation or these differences were adjusted for (2). Differences between groups were not examined (1). Groups differed and no adjustment results provided (0) Definition of outcomes (2/0) Definition of outcomes (preterm birth) Explicit specified commonly accepted definition (preterm birth less than 37 completed weeks of gestation, with or without very preterm substrata), how gestational age at delivery was determined is clearly described (2) Explicit modified definition, but the method for determining the gestational age at delivery was unspecified (1) Unspecified or unacceptable definition (0) Ascertainment of outcomes (2/0) How the diagnosis was made Prospectively diagnosed or review of notes or hospital discharge records (2) Retrospective chart review or database coding (1) Process not described (0) Sample size (1/0) Two hundred or more participants in a cohort study; 50 or more participants in either group (case control) (1) One hundred participants or more, less than 200 in a cohort; 15 participants or more, less than 50 in either group (case control) (0.5) Participants less than 100 in a cohort; participants, less than 15 in either group (case control) (0) (continued)

SOURCES
We followed the guidelines for Meta-analysis Of Observational Studies in Epidemiology (MOOSE).7 Our electronic searches targeted studies on inflammatory cytokines and the risk of spontaneous preterm birth. We searched the human literature in PubMed, EMBASE, and the Cochrane Library up to February 2010 for articles on levels of inflammatory cytokines in various biologic fluids (plasma, cervicovaginal fluid, amniotic fluid) and the risk of spontaneous preterm birth in asymptomatic women. The search key words were preterm/pre-term (birth/ delivery) and cytokine or inflammation/inflammatory marker/biomarker. Relevant studies were further sought manually in the reference lists of primary papers and reviews.

STUDY SELECTION
Full-length articles of studies evaluating inflammatory cytokines and the risk of preterm birth were scrutinized and subsequently selected if they fulfilled the following inclusion criteria: 1) maternal biologic fluid samples were taken for assays of inflammatory cytokines before 37 weeks of gestation and before delivery; 2) spontaneous preterm birth as an outcome; 3) study participants: asymptomatic womenthose who were not at risk of immediate preterm delivery (no clinical indications of labor and rupture of membranes) at the time of recruitment and specimen collection for the cytokine assays; and 4) meet the predefined methodological quality assessment criteria for nonrandomized observational studies (Box 1).8 Studies on medically indicated preterm delivery with incomplete data or data only on mean or median values of cytokine levels but no data on their associations with the risk of spontaneous preterm birth were excluded. Studies were selected in a two-stage process. Two reviewers (S.Q.W. and Z.C.L.) independently scruti-

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Box 1. Quality Assessment of Observational Studies (total 10 points)* (continued)

Study design (2/0) Prospective cohort or nested case control within a prospective cohort (2) Cross-sectional, case control or retrospective cohort (1) Not described or poorly designed (0) Exclusion: score zero in any item (1 to 6) or a total score less than 7 out of 10 maximal points
* A score based quality assessment criteria for nonrandomized observational studies. Modified from Duckitt K, Harrington D. Risk factors for preeclampsia at antenatal booking: systematic review of controlled studies. BMJ 2005; 330:565.

nized the electronic searches and obtained full-length articles of all citations that likely met the predefined selection criteria. Final inclusion or exclusion decisions were then made after we read these articles. In cases of duplicate publications, we selected the most complete version. We resolved any disagreements through consensus or arbitration by a third reviewer (W.D.F.). We evaluated the methodological quality of each study based on the study design, selection of participants, comparability of groups, definition of outcomes, ascertainment of outcomes. and sample size using the assessment criteria for nonrandomized observational studies adapted from Duckitt and Harrington.8 We excluded any study with a score of zero

in any item (1 to 6) or a total score less than 7 out of 10 maximal points. Quality scores of all selected studies are summarized in Table 1. Data were extracted from all selected articles to construct 2 2 tables of positive and negative inflammatory cytokines results (above or below the reported cutoff, respectively) compared with presence or absence of spontaneous preterm birth. Adjusted odds ratios (ORs) were also recorded if available along with the factors adjusted for. We calculated pooled OR with 95% confidence intervals (95% CIs) by cytokine and specimen. We consider ORs greater than 3 strong associations. Data on dichotomous outcomes were combined using the Mantel-Haenszel method. The heterogeneity across studies was assessed graphically using forest plots and statistically using 2 test. Each forest plot shows a point estimate for each study (with 95% CIs) with a diamond at the bottom representing the pooled point estimate with 95% CIs for each cytokine. Pooled ORs were calculated using the fixed effects or random effects models. The presence of significant heterogeneity was explored by I2 statistics.9 In cases in which I2 exceeded 50%, we pooled results using random effects models. Meta-analyses were performed using the Review Manager (RevMan) 5.0 (Copenhagen, Denmark: The Nordic Cochrane Centre, The Cochrane Collaboration; 2008).

RESULTS
Figure 1 summarizes the process of literature search and selection of studies. We identified 1,594 studies,

Table 1. Quality Scores of Included Studies on Inflammatory Cytokines and Spontaneous Preterm Birth in Asymptomatic Women
Study
Ghezzi 200210 Ghidini 199711 Goepfert 200112 Goldenberg 200113 Hvilsom 200214 Karinen 200515 Kramer 201016 Lockwood 199417 Lohsoonthorn 200718 Massaro 200919 Ozer 200520 Paternoster 200221 Pitiphat 200522 Rosa 200023 Tarim 200524 Wenstrom 199825 Yoon 200126

Selection of Participants
1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1

Comparability of Groups
2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2

Outcomes Definition
2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2

Ascertainment
2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 1

Sample Size
1 1 1 1 1 1 1 0.5 1 0.5 0.5 1 1 0.5 1 1 0.5

Study Design
2 1 2 2 2 2 2 2 1 1 2 2 2 1 2 1 1

Total Score
10 9 10 10 10 10 10 9.5 10 9.5 9.5 10 10 8.5 10 9 7.5

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Fig. 1. Flow chart of study selection process in a systematic review of inflammatory cytokines and spontaneous preterm birth in asymptomatic women.
Wei. Inflammatory Cytokines and Preterm Birth. Obstet Gynecol 2010.

and after screening the abstracts, we read 187 articles. Seventeen primary studies comprising 6,230 participants met the inclusion criteria.10 26 Table 1 presents the quality scores of the 17 included studies. The characteristics of the included studies are summarized in Table 2. No obvious publication biases were observed in a funnel plot (available on request). Figure 2 presents the results for interleukin 6 (IL-6) and spontaneous preterm birth. There were nine studies including 1,711 participants. Their overTable 2. Characteristics of the Included Studies
Study
Ghezzi 200210 Ghidini 199711 Goepfert 200112 Goldenberg 200113 Hvilsom 200214 Karinen 200515 Kramer 201016 Lockwood 199417 Lohsoonthorn 200718 Massaro 200919 Ozer 200520 Paternoster 200221 Pitiphat 200522 Rosa 200023 Tarim 200524 Wenstrom 199825 Yoon 200126

all summary crude OR (95% CI) was 2.97 (2.20 4.01). We conducted subgroup analysis by the type of specimen. The summary crude OR (95% CI) among studies of blood specimen, cervicovaginal fluid, and amniotic fluid were increasingly larger: 1.46 (0.72 2.97), 3.05 (2.00 4.67), and 4.52 (2.677.65), respectively. The overall absolute risk increase in spontaneous preterm birth was 14% for elevated IL-6 levels in both cervicovaginal and amniotic fluids. The number needed to treat was approximately seven for identify-

Country
Italy United States United States United States Denmark Finland Canada United States Thailand Italy Turkey Italy United States Spain Turkey United States Korea

Study Design
Prospective cohort Prospective cohort Nested casecontrol Nested casecontrol Nested casecontrol Nested casecontrol Nested casecontrol Prospective cohort Prospective cohort Prospective cohort Prospective cohort Prospective cohort Nested casecontrol Cross-section Prospective cohort Nested casecontrol Casecontrol

Sample Size (n)

306 179 250 214 484 506 648 161 1769 100 141 225 234 54 216 414 114

Specimen GA (wk)
1518 1420 24 2428 1418 10.4 (mean) 2426 2436 13 (mean) 1620 1520 24 519 34 or less 17.2 2.0 16.3 1.9 15.422.3

Specimen Type
AF AF CVF CVF Serum Serum Plasma CVF Serum AF AF CVF Plasma Serum AF AF AF

GA, gestational age at the time of specimen sampling; SPB, spontaneous preterm birth; AF, amniotic fluid; CRP, C-reactive protein; ELISA, enzyme-linked immunosorbent assay; IL-6, interleukin-6; CVF, cervicovaginal fluid; CLIA, chemiluminescent immunometric assay.

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ing one additional preterm delivery than that would have been expected without testing the biomarker. There was significant heterogeneity in ORs among the studies of amniotic fluid ( 2 7.09, P .07; I2 58%). Figure 3 summarizes the ORs for C-reactive protein (CRP) and spontaneous preterm birth. There were eight studies including 4,190 participants. The overall summary crude OR (95% CI) was 1.77 (1.44 2.17). There was significant statistical heterogeneity of ORs across the studies ( 2 21.72, P .001; I2 68%). There was significant statistical heterogeneity in ORs between studies of amniotic fluid ( 2 6.72, P .03; I2 70%) but no significant heterogeneity among studies of plasma CRP levels. The summary crude OR (95% CI) in studies of blood specimen and amniotic fluid were 1.53 (1.221.90) and 7.85 (3.88 15.87), respectively. The overall absolute risk increase in spontaneous preterm birth was 32% for elevated CRP levels in amniotic fluid; the number needed to treat was approximately three for identifying one additional preterm delivery. There were only a few studies on other common inflammatory cytokines (IL-1,23 IL-2,23 IL-8,21,23 IL10,13 tumor necrosis factor- [TNF- ]21) and spontaneous preterm birth in asymptomatic women. There was no evidence that these cytokines were associated

with spontaneous preterm birth. There were insufficient data (less than three studies) for the summary estimates of these cytokines for spontaneous preterm birth among asymptomatic women. Two studies18,22 provided the adjusted ORs, and both were for the association between CRP levels and spontaneous preterm birth: controlling for maternal age, race, parity, prepregnancy body mass index, and gestational age at blood sampling in Lohsoonthorn et al18 and controlling for maternal age, race, cigarette smoking, pregnancy mass index, and gestational age at blood sampling in Pitiphat et al.22 The adjustments did not affect the association in the study by Lohsoonthron et al but substantially attenuated the association in the study by Pitiphat et al.

CONCLUSION Main Findings

Our results suggest that elevated inflammatory cytokines IL-6 and CRP in amniotic fluid but not plasma are strongly associated with increased risk of spontaneous preterm birth among asymptomatic women, indicating that inflammation at the maternalfetal interface, rather than systemic inflammation, may play a major role in the etiology of such spontaneous preterm births. Elevated IL-6 levels in cervicovaginal

Storage
Test within 1 h 20C 70C 70C 80C 25C 70C 20C 80C 40C Test within 1 h 20C 80C 30C 70C 70C

Method
Turbidimetric assay ELISA ELISA ELISA ELISA Immunoenzymometric assay Cytometric assay Immunoassay Immunoassay ELISA ELISA CLIA Immunoturbidimetric assay ELISA Behring Nephelometer ELISA ELISA

Exposure
CRP more than 110 ng/mL IL6 more than 1,740 pg/mL IL6 more than 305 pg/mL IL6 more than 90th CRP 9.9 mg/L or greater CRP more than 4.3 mg/L CRP more than 5 g/L IL-6 more than 125 pg/mL CRP 7.5 mg/L or greater (75th or greater) IL-6 more than 1.2 ng/dL CRP more than 0.65 mg/L IL-6 more than 202.5 pg/L (90th or greater) CRP more than 7.96 mg/L (90th or greater) IL more than 10 pg/mL CRP 5 mg/L or greater IL-6 more than 2.9 ng/mL (more than 95th) More than 0.6 ng/mL

SPB Outcome (wk)

34 or less 34 or less 34 or less 34 or less Less than 37 Less than 37 Less than 37 Less than 37 Less than 37 Less than 37 Less than 37 Less than 37 Less than 37 34 or less Less than 37 34 or less Less than 32

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Fig. 2. Forest plot of summary crude odds ratios of the association between spontaneous preterm birth and interleukin-6 (IL-6) levels in asymptomatic women. M-H, Mantel-Haenszel test; CI, confidence interval.
Wei. Inflammatory Cytokines and Preterm Birth. Obstet Gynecol 2010.

fluid were also strongly associated spontaneous preterm birth. However, there was insufficient evidence for inflammatory cytokines IL-1, IL-2, IL-8, IL-10, and TNF- in spontaneous preterm birth among asymptomatic women.

unspecified or unacceptable definition of outcomes, or study design not described or poorly designed were not applicable to the studies we reviewed.

Mechanisms of the Cytokine-Associated Preterm Birth

Preterm birth may be precipitated by different pathophysiological events, including immune response, intrauterine infection, and stress.29 In the last two decades, much evidence has indicated that immunologic mechanisms play a key role in the success or failure of pregnancy.30 In light of multiple influences of cytokines and immune interactions at the fetomaternal interface, we may view pregnancy as a situation of altered immune status.31 Evidence points to the importance of maternal immune tolerance in successful pregnancy.32 Immune maladaptations may involve inflammatory responses at the maternalfetal interface and could be involved in a number of adverse pregnancy outcomes.33 Studies in animals have shown that an increase in proinflammatory cytokines IL-1, IL-2, IL-6, IL-8, and TNF- precedes uterine contractions and is capable of inducing the production of

Quality of Review
In contrast to previous reviews that focused on a single cytokine,27,28 we conducted a quantitatively systematic review to assess multiple common proinflammatory (IL-1, IL-2, IL-6, IL-8, TNF- , CRP) and antiinflammatory cytokines (IL-10) in various biospecimens (amniotic fluid, cervicovaginal fluid, and blood) in relation to spontaneous preterm birth. The quality of systematic reviews is dependent on the quality of studies included; we scrutinized the selected studies for their quality to exclude the poor-methodological-quality primary studies. We used strict selection criteria8 to ensure the quality of included studies. Methodological issues that may affect study quality such as if cohort was a selected unrepresentative group, significant differences between the groups reported in terms of age, plurality, smoking, preterm birth history, preexisting medical conditions,

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Fig. 3. Forest plot of summary crude odds ratios of the association between spontaneous preterm birth and C-reactive protein (CRP) levels in asymptomatic women. M-H, Mantel-Haenszel test; CI, confidence interval.
Wei. Inflammatory Cytokines and Preterm Birth. Obstet Gynecol 2010.

prostaglandins associated with labor.34 There is clear evidence demonstrating a role of inflammatory cytokines in the sequence of events leading to parturition associated with intrauterine infection, which may result in the secretion of inflammatory cytokines, including IL-1 , IL-2, IL-6, IL-8, and TNF- .35 Subclinical intrauterine infection has been implicated in preterm birth and is associated with the increased production of various cytokines in the amniotic fluid, particularly the proinflammatory cytokines IL-1, IL-2, IL-6, IL-8, and TNF- with subsequent activation and production of prostaglandins, which induce uterine contractions.36 Our meta-analysis found that certain inflammatory cytokines at the maternalfetal interface may play a major role in the events leading to spontaneous preterm birth among asymptomatic women. However, systematic inflammation seems to be not present in the etiology of spontaneous preterm birth in asymptomatic women. Studies have suggested that maternal age, parity, race and ethnicity, or prepregnancy body mass index may be associated with the risk of spontaneous preterm birth.37,38 However, ORs adjusting for these factors were unavailable in most included studies. Results from the two studies with adjusted ORs provide a mixed signal; one showed no evidence of confounding effect,18 whereas the other indicated some confounding effect.22 There are suggestions that increased uterine cytokine release is also a condition in cases of preterm

birth that are not the result of infection. Intrauterine inflammatory response syndrome may account for cases of preterm birth in which no infectious organisms can be identified and that the production of inflammatory cytokines may be a mechanism in the pathophysiological basis of this association.39 The results from this meta-analysis suggested that IL-6 and CRP may well be involved in the sequence of events leading to spontaneous preterm birth among asymptomatic women. These mediators have not yet been entirely elucidated, but it is likely that inflammatory cytokines bring about parturition through increasing prostaglandin levels by manipulating prostaglandin production, metabolism, or both. The elucidation of how these cytokines mediate such effects and whether their effects can be mitigated by appropriate treatments merit further research.

Limitations
The findings of the current meta-analysis must be interpreted cautiously. First, there are wide variations in the reported value of cytokines. There are also several assay techniques to measure cytokines, including radioactive assay, chemiluminescent immunometric assay, turbidimetric assay, immunoturbidimetric assay, and enzyme-linked immunosorbent assay. Second, different test thresholds were chosen arbitrarily in different studies. However, these variations are unlikely to be of clinical significance in comparing the

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highest exposed with the remaining reference group among various studies. Third, we reviewed only published studies. Unpublished studies may contain valid data that conflict with the results. Finally, there were some relatively small sample sizes with wide CIs in some studies.

prenatal care. These promising approaches in combination with other screening tests (eg, cervical length, fibronectin) may offer the potential to identify patients at risk of spontaneous preterm birth and improve perinatal outcomes. REFERENCES
1. Goldenberg RL, Culhane JF, Iams JD, Romero R. Epidemiology and causes of preterm birth. Lancet 2008;371:75 84. 2. McCormick MC. The contribution of low birth weight to infant mortality and childhood morbidity. N Engl J Med 1985; 312:8290. 3. Saigal S, Doyle LW. An overview of mortality and sequelae of preterm birth from infancy to adulthood. Lancet 2008;371: 2619. 4. Slattery MM, Morrison JJ. Preterm delivery. Lancet 2002;360: 1489 97. 5. El-Bastawissi AY, Williams MA, Riley DE, Hitti J, Krieger JN. Amniotic fluid interleukin-6 and preterm delivery: a review. Obstet Gynecol 2000;95:1056 64. 6. Velez DR, Fortunato SJ, Morgan N, Edwards TL, Lombardi SJ, Williams SM, et al. Patterns of cytokine profiles differ with pregnancy outcome and ethnicity. Hum Reprod 2008;23: 19029. 7. Stroup DF, Berlin JA, Morton SC, Olkin I, Williamson GD, Rennie D, et al. Meta-analysis of observational studies in epidemiology: a proposal for reporting. Meta-analysis Of Observational Studies in Epidemiology (MOOSE) group. JAMA 2000;283:2008 12. 8. Duckitt K, Harrington D. Risk factors for pre-eclampsia at antenatal booking: systematic review of controlled studies. BMJ 2005;330:565. 9. Higgins JP, Thompson SG. Quantifying heterogeneity in a meta-analysis. Stat Med 2002;21:1539 58. 10. Ghezzi F, Franchi M, Raio L, Di Naro E, Bossi G, DEril GV, et al. Elevated amniotic fluid C-reactive protein at the time of genetic amniocentesis is a marker for preterm delivery. Am J Obstet Gynecol 2002;186:268 73. 11. Ghidini A, Jenkins CB, Spong CY, Pezzullo JC, Salafia CM, Eglinton GS. Elevated amniotic fluid interleukin-6 levels during the early second trimester are associated with greater risk of subsequent preterm delivery. Am J Reprod Immunol 1997;37: 22731. 12. Goepfert AR, Goldenberg RL, Andrews WW, Hauth JC, Mercer B, Iams J, et al. The Preterm Prediction Study: association between cervical interleukin 6 concentration and spontaneous preterm birth. National Institute of Child Health and Human Development MaternalFetal Medicine Units Network. Am J Obstet Gynecol 2001;184:483 8. 13. Goldenberg RL, Iams JD, Mercer BM, Meis PJ, Moawad A, Das A, et al. The Preterm Prediction Study: toward a multiplemarker test for spontaneous preterm birth. Am J Obstet Gynecol 2001;185:64351. 14. Hvilsom GB, Thorsen P, Jeune B, Bakketeig LS. C-reactive protein: a serological marker for preterm delivery? Acta Obstet Gynecol Scand 2002;81:424 9. 15. Karinen L, Pouta A, Bloigu A, Koskela P, Paldanius M, Leinonen M, et al. Serum C-reactive protein and Chlamydia trachomatis antibodies in preterm delivery. Obstet Gynecol 2005;106:73 80. 16. Kramer MS, Kahn SR, Platt RW, Genest J, Chen MF, Goulet L, et al. Mid-trimester maternal plasma cytokines and CRP as

Clinical Application
The evidence from this systematic review indicates the association between elevated levels of certain proinflammatory cytokines and spontaneous preterm birth. We conclude that elevated IL-6 and CRP in asymptomatic women is consistently associated with spontaneous preterm birth among asymptomatic women. Elevated levels of IL-6 in amniotic fluid or cervicovaginal fluid and of CRP in amniotic fluid may indicate preterm delivery. In clinical practice, tests for inflammatory cytokines in amniotic fluid requiring invasive amniocentesis procedures are not justifiable for asymptomatic women. In contrast, inflammatory cytokines in cervicovaginal fluid may have clinical use because the required specimen collection procedures pose virtually no risks to the mother and fetus. The strong association between elevated IL-6 levels in cervicovaginal fluid and spontaneous preterm birth in asymptomatic women indicates the need for more studies on other inflammatory cytokines in cervicovaginal fluid. The wide ranges in reported cutoffs in cervicovaginal fluid IL-6 levels indicate the need for standardized assay methods and protocols in more clinical studies to better define the cutoffs and clinical use for this promising biomarker. Elevated IL-6 levels in vaginal fluid may alert obstetricians to consider more frequent follow-up of such asymptomatic patients and additional other tests (eg, fetal fibronectin) in subsequent follow-up visits to evaluate the risk of preterm delivery. The demonstration of a possible association between proinflammatory cytokines and spontaneous preterm birth raises the question as to whether the evaluation of inflammatory cytokine reactivity may be valuable even in the clinical management of asymptomatic patients. The risk of preterm birth, and hence the therapeutic benefits, depends on the posttest probabilities of preterm birth associated with cytokines IL-6 and CRP testing. Our results may inform clinicians to make a more rational approach to decision-making regarding inpatient admission and administration of antenatal steroids in women with elevated inflammatory cytokines IL-6 or CRP levels. Future research should focus on clinical application of standardized immunoassays to detect elevated IL-6 and CRP levels during

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