437. A Light-Producing Model of Infection-Related Preterm Birth

Abstract

Background:Preterm birth is increasing in incidence and current therapies are relatively ineffective. It is responsible for >1million neonatal deaths per annum worldwide and long term complications in survivors. Approximately 50% of PTBs are preceded by microbial invasion of the intrauterine space; current clinical management centres on diagnosis of intrauterine bacterial presence by identifying the resultant inflammatory response. To investigate the relationship between intrauterine bacterial presence and inflammation we developed two separate gene technology approaches:1. Intravaginal bioluminescent bacteria to measure bacterial ascent into the uterus which mimics the ascending vaginal infection seen in preterm birth.2. Lentiviral gene transfer of an NFKB activated luciferase reporter construct to allow bioluminescent imaging of the subsequent systemic NFKB response.Methods:An NFkB response element was cloned into a lentivirus vector upstream of the genes encoding a codon-optimised firefly luciferase. High titred virus was injected intravenously at birth to neonatal female C57BL/6 J-Tyrc-2J mice to achieve luciferase expression predominantly in the liver (to monitor systemic inflammatory response). These mice received Escherichia coli (non-pathogenic K-12, MG1655 with integrated luxABCDE operon) intra-vaginally once reaching adulthood and intraperitoneal lipopolysaccharide (LPS) three weeks later. Luciferase expression was monitored by whole body bioluminescence imaging. Local inflammation was determined using H&E, ICAM-1 (Intracellular adhesion molecule 1) and Ly6g immunohistochemistry and enzyme-linked immunosorbent assays for serum and uterine TNF-α and IL1-β cytokines.Results:Bioluminescent imaging revealed that C57BL/6 J-Tyrc-2J mice were the most susceptible mice breeds for modelling of ascending vaginal infection with E. coli luxABCDE operon. Intraperitoneal LPS induced an NF-KB response in the liver by biosensing (p<0.01), however intravaginal E. coli administration induced no response. There was evidence of uterine inflammation with an upregulation of ICAM-1 and neutrophils.Conclusion:Although it is possible to detect LPS-induced NFKB inflammation in the liver by biosensing, ascending vaginal infection induced no response. This highlights the clinical challenge of identifying bacterial presence, confined to the uterus, using systemic markers. This model can be used to test new treatments for the prevention of PTB.View Large Image | Download PowerPoint Slide