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ORIGINAL RESEARCH |
From the Department of Obstetrics and Gynecology and Institute of Clinical Chemistry and Pathobiochemistry, University Hospital, Technical University, Aachen, Germany; and Institute of Pathology, University of Tuübingen, Tübingen, Germany.
Address reprint requests to: Matthias Winkler, MD Frauenklinik fur Gynäkologie und Geburtshilfe Universitätsklinikum der RWTH Pauwelsstr. 30 Aachen, D-52057 Germany E-mail: mwinkler{at}post.klinikum.rwth-aachen.de
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Abstract |
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Methods: We analyzed protein extracts from the lower uterine segments of 38 women who had nonelective cesareans at term. We measured concentrations of intercellular adhesion molecule-1, endothelial leukocyte adhesion molecule-1, vascular cell adhesion molecule-1, and platelet endothelial cell adhesion molecule-1 by enzyme-linked immunosorbent assay. Subjects were grouped according to cervical dilatation (less than 2 cm, n = 10; 2 to less than 4 cm, n = 9; 4–6 cm, n = 9; more than 6 cm, n = 10) and duration of labor (up to 6 hours, n = 14; 6–12 hours, n = 10; 12–24 hours, n = 9; longer than 24 hours, n = 5) at the time of cesarean.
Results: The median concentration of intercellular adhesion molecule-1 increased significantly with increasing dilatation (from 2.24 ng/mg total protein at less than 2 cm to 6.73 ng/mg at 4–6 cm) and increasing duration of labor (from 2.53 ng/mg up to 6 hours to 5.90 ng/mg at 12–24 hours). However, this study did not have adequate statistical power to identify differences in concentrations of the other endothelial adhesion molecules.
Conclusion: The results indicate that parturition at term is associated with expression of intercellular adhesion molecule-1.
According to Liggins,1 cervical ripening at term resembles an inflammatory reaction. Further studies supported that theory, showing that cervical dilatation was accompanied by infiltration of the cervical stroma by neutrophils2,3 and increasing concentrations of proinflammatory cytokines (interleukin-1ß, tumor necrosis factor 
), immunomodulatory cytokines (interleukin-6), chemotactic cytokines (interleukin-8),4–6 and proteases6–8 in the lower uterine segment. Cytokine-mediated expression of adhesion molecules by the capillary endothelium is a prerequisite of extravasation of neutrophils. We have reported the immunohistochemical finding of increased expression of endothelial leukocyte adhesion molecule-1 (also known as E-selectin) and vascular cell adhesion molecule-1 during cervical dilation at term.9 This study was done to verify those results by quantification of concentrations of endothelial leukocyte adhesion molecule-1, vascular cell adhesion molecule-1, intercellular adhesion molecule-1, and platelet endothelial cell adhesion molecule-1.
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Materials and Methods |
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and 2).
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For quantification of cell adhesion molecules, specimens (wet weight approximately 100–200 mg) were homogenized for 1 minute while still frozen (Dismembranator, Braun, Melsungen, Germany), and the protein was extracted overnight at 4C. The powder was suspended in 1 mL triethanolamine-buffered saline (0.02 mol/L Tris-hydrochloric acid pH 8.5, 0.125 sodium chloride) that contained 1 mmol/L diisopropyl flurophosphate, 10 mmol/L ethylenediaminetetra-acetic acid, 5 µmol/L pepstatin, and 50 µmol/L E-64. Aliquots of the clarified extract were investigated for concentrations of intercellular adhesion molecule-1, endothelial leukocyte adhesion molecule-1, vascular cell adhesion molecule-1, and platelet endothelial cell adhesion molecule-1 with commercially available enzyme-linked immunosorbent assays (ELISAs) (R&D Systems, Biermann, Bad Nauheim, Germany), which use polyclonal antibodies, except for platelet endothelial cell adhesion molecule-1, for which the primary antibody is monoclonal. Those assays evaluate the recombinant and native species of the various adhesion molecules. According to the manufacturer, cross-reactions with other adhesion molecules or native human immunoglobulin (Ig) G do not occur. All the assays were done in triplicate, and a zero standard and three standard concentrations were included in each run. The intraassay coefficients of variation ranged from 5–15%. Values were expressed in relation to total protein content, as determined by the pyrogallol method (Dimension AR, Dade, Munich, Germany).
Differences in concentrations of adhesion molecules between various groups, defined according to cervical dilatation and duration of labor, were analyzed by Kruskal-Wallis nonparametric analysis of variance supplemented by nonparametric pairwise comparisons. Probability values of less than 5% were considered significant. A power calculation was done for the non-significant differences in concentration to determine the number of women necessary for significance to be reached with median differences measured, an error of 5%, and a ß error of 10%.
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). The concentration of intercellular adhesion molecule-1 also depended on the duration of labor: the median concentration at 6–12 hours and at 12–24 hours was significantly higher than before 6 hours, but after 24 hours, it fell again and was not significantly higher than before 6 hours (Table 2).
Concentrations of endothelial leukocyte adhesion molecule-1, vascular cell adhesion molecule-1, and platelet endothelial cell adhesion molecule-1 were not related to cervical dilatation or duration of labor (Tables 1 and 2).
Our power calculation showed that the sample would have to be between 21 and more than 1000 patients to detect significant differences in concentrations of those adhesion molecules.
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Discussion |
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, among other substances.12 We reported a progressive increase in interleukin-1ß and tumor necrosis factor in the lower uterine segment during cervical dilation at term,5 which parallels the increase in intercellular adhesion molecule-1 in this tissue. Like intercellular adhesion molecule-1, the concentrations of interleukin-1ß and tumor necrosis factor increase as the cervix dilates up to 4–6 cm, but there is no further increase with further dilation (greater than 6 cm). Those findings suggest that changes in intercellular adhesion molecule-1 concentration in the lower uterine segment might be mediated by proinflammatory cytokines. We interpret the fact that the concentrations of interleukin-1ß, tumor necrosis factor , and intercellular adhesion molecule-1 appear to reach a plateau at 4–6 cm dilatation as an indication that the processes of extravasation of leukocytes are already being curbed from as early as that stage, to prevent severe tissue damage from prolonged action of granulocyte-derived catabolic enzymes, and establish the conditions needed for normal reconstitution of the cervix postpartum. Our in vitro findings that substances such as ß-estradiol and prostaglandin-E2, which induce cervical ripening, also can stimulate significant increases in expression of intercellular adhesion molecule-1 by human umbilical vein endothelial cells13 also suggest that intercellular adhesion molecule-1 might be involved in the process of cervical dilation at term.
Endothelial leukocyte adhesion molecule-1 mediates adhesion of neutrophils, monocytes, and various T-cell subsets to endothelium in the process of acute inflammation. For that to happen, the endothelium has to be activated first, for example by interleukin-1 and tumor necrosis factor .12,14
Vascular cell adhesion molecule-1 also is expressed to only low levels by nonactivated endothelial cells and mediates the adhesion of monocytes, lymphocytes, and basophilic and eosinophilic granulocytes.14 Expression of vascular cell adhesion molecule-1 also can be induced by proinflammatory cytokines (eg, interleukin-1 and tumor necrosis factor ).12
Platelet endothelial cell adhesion molecule-1 mediates endothelial cell-cell contact and interactions between endothelial cells and monocytes, granulocytes, and T cells.15
The quantitative findings of this study on concentrations of intercellular adhesion molecule-1, endothelial leukocyte adhesion molecule-1, vascular cell adhesion molecule-1, and platelet endothelial cell adhesion molecule-1 differ from those of earlier immunohistochemical studies of expression of adhesion molecules in the lower uterine segment by our group9 and Thomson et al.16 In those studies, the numbers of endothelial cells that expressed endothelial leukocyte adhesion molecule-1 and vascular cell adhesion molecule-1 increased as cervical dilation progressed. That almost all the vascular endothelial cells at all stages of cervical dilatation expressed intercellular adhesion molecule-1 and platelet endothelial cell adhesion molecule-1 also was inconsistent with the findings of this study, in that there was no change during labor. The inconsistencies stem most probably from the different methods used. The quantification of expression by ELISA is more accurate than the semiquantitative method used in the immunohistochemical investigation, but with the latter, it is possible to evaluate expression of adhesion molecules by endothelium alone, whereas immunoassay measures expression in tissue as a whole. That is almost certainly relevant in intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and platelet endothelial cell adhesion molecule-1, which all are expressed not only by endothelial cells, but to varying extents by other cells, such as certain leukocyte subsets and fibroblasts.17
The precise functions of the various adhesion molecules during labor remain unclear. We showed that expression of certain endothelial adhesion molecules is increased during labor, indicating that such molecules might be involved in cervical dilation. With the finding that the numbers of leukocytes3,16 and macrophages16 in the lower uterine segment stroma increase during parturition, our results support the hypothesis that cervical dilatation at term resembles an inflammatory reaction.
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Footnotes |
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Received April 7, 1999. Received in revised form August 26, 1999. Accepted September 2, 1999.
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2. Junqueira LC, Zugaib M, Montes GS, Toledo OM, Krisztan RM, Shigihara KM. Morphologic and histochemical evidence for the occurrence of collagenolysis and for the role of neutrophilic polymorphonuclear leukocytes during cervical dilation. Am J Obstet Gynecol 1980;138:273–81.[Medline]
3. Winkler M, Fischer DC, Ruck P, Marx T, Kaiserling E, Rath W. Interleukin-8 associated increase in the number of neutrophilic granulocytes in the cervical stroma as a basic mechanism of cervical dilatation during parturition at term. Geburtsh Frauenheilk 1997;57:300–2.
4. Sennström MK, Brauner A, Lu Y, Granström LM, Malmström AL, Ekman GE. Interleukin-8 is a mediator of the final cervical ripening in humans. Eur J Obstet Gynecol Reprod Biol 1997;74:89–92.[Medline]
5. Winkler M, Fischer DC, Ruck P, Horny HP, Kemp B, Rath W. Cytokine concentration and expression of cell adhesion molecules in the lower uterine segment during parturition at term: Correlations with the stage of cervical dilatation and the duration of labor. Z Geburtshilfe Neonatol 1998;202:172–5.[Medline]
6. Osmers RGW, Blaäser J, Kuhn W, Tschesche H. Interleukin-8 synthesis and the onset of labor. Obstet Gynecol 1995;86:223–9.[Abstract]
7. Rajabi MR, Dean DD, Beydoun SN, Woessner JF Jr. Elevated tissue levels of collagenase during dilation of uterine cervix in human parturition. Am J Obstet Gynecol 1988;159:971–6.[Medline]
8. Winkler M, Oberpichler A, Tschesche H, Ruck P, Fischer DC, Rath W. Collagenolysis in the lower uterine segment during parturition at term: Correlations with stage of cervical dilatation and duration of labor. Am J Obstet Gynecol 1999;181:153–8.[Medline]
9. Winkler M, Ruck P, Horny HP, Wehrmann M, Kemp B, Kaiserling E, et al. Expression of cell adhesion molecules by endothelium in the human lower uterine segment during parturition at term. Am J Obstet Gynecol 1998;178:557–61.[Medline]
10. Winkler M, Fischer DC, Hlubek M, van de Leur E, Haubeck HD, Rath W. Interleukin-1ß and interleukin-8 concentrations in the lower uterine segment during parturition at term. Obstet Gynecol 1998;91:945–9.[Abstract]
11. Rayburn WF, Wapner RJ, Barss VA, Spitzberg E, Molina RD, Mandsager N, et al. An intravaginal controlled-release prostaglandin E2 pessary for cervical ripening and initiation of labor at term. Obstet Gynecol 1992;79:374–9.[Abstract]
12. Carlos TM, Harlan JM. Membrane proteins involved in phagocyte adherence to endothelium. Immunol Rev 1990;114:5–28.[Medline]
13. Winkler M, Kemp B, Hauptmann S, Rath W. Parturition: Steroids, prostaglandin E2, and expression of adhesion molecules by endothelial cells. Obstet Gynecol 1997;89:398–402.[Abstract]
14. Fries JWU, Williams AJ, Atkins RC, Newman W, Lipscomb MF, Tucker C. Expression of VCAM-1 and E-selectin in an in vivo model of endothelial activation. Am J Pathol 1993;143:725–37.[Abstract]
15. Simmons DL, Walker C, Power C, Pigott R. Molecular cloning of CD31, a putative intercellular adhesion molecule closely related to carcinoembryonic antigen. J Exp Med 1990;171:2147–52.
16. Thomson AJ, Telfer JF, Young A, Campbell S, Stewart CJ, Cameron IT, et al. Leukocytes infiltrate the myometrium during human parturition: Further evidence that labour is an inflammatory process. Hum Reprod 1999;14:229–36.
17. Pigott R, Power C. The adhesion molecule facts book. London: Academic Press, Harcourt Brace & Company, 1993.
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  M. A. LEDINGHAM, A. J. THOMSON, F. JORDAN, A. YOUNG, M. CRAWFORD, and J. E. NORMAN Cell Adhesion Molecule Expression in the Cervix and Myometrium During Pregnancy and Parturition Obstet. Gynecol., February 1, 2001; 97(2): 235 - 242. [Abstract] [Full Text] [PDF]
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