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Title: DNA-mediated inflammation in pregnancy
Author: van Boeckel, Sara Raquel
ISNI:       0000 0004 9355 8635
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2020
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During pregnancy, the maternal immune system must adapt to allow for the growth of a semiallogenic fetus whilst maintaining the ability to respond to pathogens. Maternal immune system adaptions are imperfect. Pregnant women are more susceptible to infections and have a higher morbidity from infections than non-pregnant women. An inappropriate inflammatory response to the developing fetus can result in miscarriage or preterm birth. The maternal immune system is exposed to circulating cell-free fetal DNA (cff-DNA) in pregnancy, produced by the cell death of placental syncytiotrophoblasts. Circulating DNA is pro-inflammatory through various cellular pathways, including through intracellular toll-like receptor 9 (TLR9). Fetal DNA can elicit inflammation both in vivo, resulting in fetal resorption in mouse models, and in vitro in peripheral blood mononuclear cells (PBMCs) from pregnant women. As such, cff-DNA has potential to be involved in the pathogenesis of preterm birth. When compared to microbial DNA, it is more difficult for mammalian DNA (such as cff-DNA) to reach intracellular TLR9. Cathelidicin, a circulating host defence peptide, facilitates DNA entry to cells, and can enhance TLR9-mediated inflammation. It is not known whether cathelicidin can mediate inflammation in pregnancy. Indeed, little is known about microbial and cff-DNA sensing mechanisms in pregnancy. This thesis was based on the following hypotheses: (1) cff-DNA is pro-inflammatory in pregnancy; (2) cathelicidin modulates cff-DNA induced inflammatory response in pregnancy; and (3) the pathways of DNA-induced inflammation are altered in pregnancy. To investigate these hypotheses, I aimed to: (1) assess the pro-inflammatory potential of cff-DNA (2) explore the role of cathelicidin in cff-DNA-mediated inflammation in pregnancy, (3) to determine if PBMC responses to DNA-stimuli differ between pregnant and non-pregnant women. To assess the pro-inflammatory potential of cff-DNA, cff-DNA was generated using human placental explants. cff-DNA was not pro-inflammatory to PBMCs from pregnant women and in an in vivo model of preterm birth, injection of mouse placental DNA did not decrease time to delivery. These data suggested that cff-DNA and placental DNA alone are not proinflammatory, possibly due to the inability of cff-DNA to reach intra-cellular TLR9. When cathelicidin was added, cff-DNA successfully induced inflammation in the form of chemokine interferon-γ inducible protein ϭϬ ;CXCLϭϬͿ production from PBMCs from pregnant women. In a lipopolysaccharide (LPS) ʹ induced inflammatory preterm birth model, cathelicidin deficient mice had decreased rates of preterm birth compared to wildtype mice. Cathelicidin deficient mice also had significantly less circulating interleukin 6, suggesting that cathelicidin contributes to inflammation in LPS-induced inflammatory preterm birth. To examine whether pregnancy results in an altered inflammatory response to DNA-stimuli, the inflammatory responses of PBMCs from pregnant and non-pregnant women to CpG-ODN (a synthetic DNA TLR9 agonist) were compared. PBMCs from pregnant women produced a significantly smaller type 1 interferon-driven response (at protein level) and an overall lesser inflammatory response (at transcriptional level) compared to PBMCs from non-pregnant women. PBMCs from pregnant women also had a lower proportion of TLR9-positive plasmacytoid dendritic cells (pDCs), providing a possible mechanism for the reduced inflammatory response to DNA-stimuli in pregnancy. The placenta produces large amounts of progesterone in pregnancy, and progesterone can affect TLR9 responsiveness. Therefore, the ability for progesterone to alter TLR9 expression and function on PBMCs from non-pregnant women was explored. Progesterone pre-treatment significantly decreased the CXCL10 response of PBMCs from non-pregnant women to CpG-ODN, and decreased TLR9 expression in pDCs from non-pregnant women. Together, this suggested that DNA-mediated inflammation from PBMCs is altered during pregnancy, potentially due to progesterone. The altered inflammatory response to DNA-stimuli in pregnancy described in this thesis is a novel finding. It represents a mechanism that contributes to the altered immune response in pregnancy. This mechanism may be a protective physiological response of the maternal immune system against increasing cff-DNA levels during pregnancy. It may, however, also contribute to the known vulnerability of pregnant women to infections. Together, the findings of this thesis provide an avenue for the future investigation of the altered maternal immune system and of the ability of cathelicidin to modulate its response to DNA-stimuli.
Supervisor: Stock, Sarah ; Davidson, Donald ; Norman, Jane Sponsor: Medical Research Council (MRC)
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
Keywords: pregnancy ; cell-free fetal DNA ; DNA ; inflammation ; cathelicidin ; antimicrobial peptides ; preterm birth ; maternal infection ; maternal immune modulation ; parturition ; toll-like receptor 9