Decidual hemorrhage/abruption results in intense local thrombin generation and is associated with both preterm premature rupture of the membrane (PPROM) and chorioamnionitis (CAM). Prior studies indicate that thrombin, acting via protease activated receptors (PARs), induces an intense decidual cytokine and proteolytic response. The link between abruption and CAM suggests an impaired immune response. Tolllike receptors (TLRs) initiate the host innate immune response. By promoting the innate immune response, TLRs provide the first line of defense against an array of microbial pathogens. We now demonstrate that decidual cells express TLRs and their intermediate signaling proteins. Importantly, we demonstrate that thrombin down-regulates decidual cell TLR expression and signaling. Finally, we demonstrate that abruption-associated PTD with or without CAM are accompanied by decreased decidual TLR expression. We postulate that decidual hemorrhage leads to intense local thrombin generation which paradoxically induces a local aseptic inflammatory reaction and promotes ascending genital tract infections by downregulating TLR expression and function. We propose four Specific Aims to assess the interactions between thrombin and TLRs expressed by all decidual cells. 1) Immunohistochemistry, immunofluorescence and microdissection coupled with quantitative RT-PCR will be utilized to quantify the association between altered TLR expression and abruption-associated PTD with and without related CAM. 2) To determine the mechanism(s) by which thrombin down-regulates decidual cell-expressed TLR levels and function. Studies will dissect out the role of PARs in the expression of TLRs and their downstream signaling intermediates as well as cytokine and NFkB expression. These studies will use agonists, antagonists and small interference RNA (siRNAs) in cell culture. 3) To determine the functional effects of thrombin on TLR-ligand interactions. Cultured decidual cells will be treated with thrombin vs. control and then exposed to TLR-2, 3 and 4 ligands. Endpoints of these studies will be assessed by microarray analysis, RT-PCR and immunoassays as well as assessment of the NFkB components by western blotting. 4) Lastly, in coordination with Projects I and III of this PO1, we will utilize a murine model to study the effects of thrombin on TLR expression and function as well as the susceptibility to bacterial infection. These studies will provide unique insights into the fundamental mechanisms underlying abruption associated PTD and dissect out the role of innate immune dysfunction in this major public health problem.

Public Health Relevance

Decidual hemorrhage is a major cause of perinatal morbidity and mortality associated with aseptic inflammation, increased CAM and PTD. Abruption results in intense decidual thrombin generation and PPROM. We show that the excess thrombin alters TLR expression and TLR-signaling in the decidua. This proposal ascertains the relationship between the decidual innate immune system and mechanisms underlying abruption-associated PTD and ascending genital tract infections in this major health problem.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Program Projects (P01)
Project #
5P01HD054713-05
Application #
8473081
Study Section
Special Emphasis Panel (ZHD1-MCHG-B)
Project Start
Project End
Budget Start
2013-06-01
Budget End
2014-05-31
Support Year
5
Fiscal Year
2013
Total Cost
$206,627
Indirect Cost
$81,245
Name
Yale University
Department
Type
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
06520
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Racicot, Karen; Cardenas, Ingrid; Wunsche, Vera et al. (2013) Viral infection of the pregnant cervix predisposes to ascending bacterial infection. J Immunol 191:934-41
Krikun, Graciela; Potter, Julie A; Abrahams, Vikki M (2013) Human endometrial endothelial cells generate distinct inflammatory and antiviral responses to the TLR3 agonist, Poly(I:C) and the TLR8 agonist, viral ssRNA. Am J Reprod Immunol 70:190-8

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