DEPARTMENT OF WOMEN AND CHILD HEALTH DIVISION OF OBSTETRICS AND GYNAECOLOGY Karolinska Institute, Stockholm, Sweden Mediators of cervical ripening in preterm birth: Experimental and clinical investigations

Mediators of cervical ripening in preterm birth: Experimental and clinical investigations Susanne Abelin Törnblom, MD Department of Women and Child Health, Division of Obstetrics and Gynaecology, Karolinska Institute and Karolinska University Hospital Solna, S-171 76 Stockholm, Sweden Background: Preterm birth (PTB) is by far the leading worldwide cause of infant mortality and morbidity. Despite decades of research, the frequency of PTB has not decreased and the basic mechanisms initiating the onset of labour are still poorly understood. Vaginal preterm birth cannot take place without cervical softening and remodelling. Cervical ripening at term is an inflammatory-like process, in which complex interactions between cytokines, prostaglandins and nitric oxide (NO) are believed to play key roles, with NO acting as the final mediator. An understanding of the mediators regulating this process in connection with spontaneous preterm labour is of equally great importance, e.g., when, due to maternal/foetal complications, induction of premature cervical ripening and onset of labour is desired. Aim: Compare preterm cervical ripening to the analogous process occurring at term. Methods: Transvaginal cervical biopsy specimens were obtained and serum samples taken. Women, exhibiting no clinical signs of infection and undergoing preterm labour with a ripe cervix or preterm birth without labour (unripe cervix) were compared to un-infected women delivering at term, with or without labour. Peripheral white blood cell counts (WBC) and serum levels of C-reactive protein (CRP) were determined by routine procedures in the clinical laboratory. The other parameters monitored and procedures employed were as follows: prostaglandin dehydrogenase, (PGDH), and cyclooxygenase, (COX) proteins (immunohistochemistry (IHC) and dual immunofluorescence) and mRNA ́s (Northern blot). NO synthase; bNOS, eNOS and iNOS proteins (IHC) and mRNA ́s (Real-time RT-PCR) IL-6, IL-8, MCP-1, RANTES and TNF-α proteins (ELISA/ Immulite) and IL-8, MCP-1 and RANTES mRNA (RT-PCR). Induction of cervical ripening and labour was performed using local administration of PgE2 and/or intravenous infusion of oxytocin. Results: The only significant differences between women undergoing preterm and term labour were in the levels of expression of bNOS, eNOS, and iNOS mRNA ́s, which where all higher in the preterm group. Other significant changes were only seen upon comparisons of women undergoing and not undergoing labour, irrespective of the gestational age of the foetus. Thus, cervical levels of IL-8 and MCP-1 mRNA ́s and proteins, and of IL-6 protein and serum WBC counts and levels of CRP were all higher in connection with labour. An indication of increased prostaglandin activity, i.e., reduced cervical expression of PGDH mRNA, was also observed in women in labour. In the clinical investigation induction of cervical ripening and labour with PgE2 applied locally, preterm, term or at postterm was not associated with any differences, in the mode of delivery or neonatal outcome. The fact that postterm pregnancy is a high risk obstetric situation, was emphasized by the four-fold higher frequency of extensive postpartum bleeding, >1000 ml, in this situation compared to labour at term. Conclusions: This study supports the hypothesis that preterm cervical ripening involves an inflammatory process, which may constitute a normal physiological adaptation to the onset of labour. The elevations in WBC count and serum level of CRP are striking indicators of active labour. NO appears to play a crucial role in preterm cervical ripening, consistent with its presumed acting as the final mediator. Local application of PgE2 to induce preterm cervical ripening and labour is effective and safe, for both mother and child. Key words; cervix, preterm, cervical ripening, cytokines, chemokines, cyclooxygenase (COX), prostaglandin dehydrogenase (PGDH), induction, labour, nitric oxide, PgE2-gel, pregnancy, preterm labour LIST OF PUBLICATIONS This thesis is based on the following articles and manuscripts, which will be referred to in the text by their Roman numerals: I. Susanne Abelin Törnblom, Falguni A. Patel, Birgitta Byström, Diana Giannoulias, Anders Malmström, Maria Sennström, Stephen J. Lye, John R.G. Challis, Gunvor Ekman 15-HydroxyProstaglandin Dehydrogenase and Cyclooxygenase 2 Mesenger Ribonucleic Acid Expression and Immunohistochemical Localization in Human Cervical Tissue During Term and Preterm Labor Clinical Journal of Endocrinology & Metabolism, 2004, 89, 2909-15 II. Susanne Abelin Törnblom, Holger Maul, Robert E. Garfield, Birgitta Byström, Anders Malmström, Gunvor Ekman-Ordeberg mRNA Expression and Localization of bNOS, eNOS and iNOS in Human Cervix at Preterm and Term Labour Submitted for publication III. Susanne Abelin Törnblom, Aurelija Klimaviciute, Birgitta Byström, Milan Chromek, Anders Malmström, Annelie Brauner, Gunvor Ekman-Ordeberg Cytokines in Human Cervix in Non-Infected Preterm Birth. Submitted for publication IV. Susanne Abelin Törnblom, Eva Östlund, Lena Granström , Gunvor Ekman Preterm Cervical Ripening and Labor Induction European Journal of Obstetrics & Gynecology and Reproductive Biology. 2002;104:120-3 CONTENTS 1 BACKGROUND..........................................................................................1 1.