Faculty Bibliography

PROBLEM/OBJECTIVE:Placental infection induces increased levels of pro-inflammatory cytokines, which have been implicated in the pathogenesis of preterm labor. Endotoxin tolerance is a phenomenon in which exposure to a dose of endotoxin makes tissue less responsive to subsequent exposures. The objective of our study is to determine if repeated exposure to endotoxin will induce a tolerant phenotype in normal human second trimester placental tissue. METHODS OF STUDY/METHODS:Human second trimester placental explants from elective termination of pregnancy were cultured and exposed to endotoxin (LPS). After 24 hours, the media was collected for analysis, and the explants were re-exposed to LPS after adding fresh media for another 24 hours. This process was repeated for a total of 4 LPS doses. The media was collected from each day and analyzed for cytokine levels. RESULTS:The first LPS treatment stimulated the secretion of the pro-inflammatory cytokines IL-1β, TNF-α. However, their production was significantly diminished with repeated LPS doses. Production of anti-inflammatory cytokines, IL-1ra and IL-10, was also stimulated by the first LPS treatment, but secretion was more gradually and moderately decreased with repeated LPS doses compared to the pro-inflammatory cytokines. The ratios of the anti-inflammatory/pro-inflammatory mediators (IL-1ra/IL-1β and IL-10/ TNF-α) indicate a progressively more anti-inflammatory milieu with repeated LPS doses. CONCLUSIONS:Repeated LPS exposure of human second trimester placental tissues induced endotoxin tolerance. We speculate that endotoxin tolerance at the maternal-fetal interface will protect the fetus from exaggerated inflammatory responses after repeated infectious exposure.

BACKGROUND: Intrauterine infection and inflammation during pregnancy, which leads to up-regulation of inflammatory cytokines and prostaglandin synthesis, has been implicated in the pathogenesis of preterm delivery and other pregnancy complications. Effective preventive and therapeutic strategies to reduce these outcomes are lacking to date. Pentoxifylline (PTX) is a non-specific phosphodiesterase inhibitor which raises intracellular cyclic adenosine monophosphate and decreases production of pro-inflammatory mediators while enhancing anti-inflammatory cytokines. We hypothesized that pentoxifylline will decrease lipopolysaccharide (LPS)-induced pro-inflammatory cytokines production in human placental explants. METHODS: Placental explants derived from normal second trimester human placentas were treated with PTX, stimulated with LPS and cultured at 37 degrees C in 5% CO2. Conditioned media were assayed for pro- and anti-inflammatory mediators with multiplex immunoassays or ELISA, and explant tissues for mRNA with real time PCR. Means of PTX-treated and untreated samples were compared using paired t tests and Wilcoxon-signed rank tests. RESULTS: PTX preferentially inhibited placental expression and production of LPS-induced pro-inflammatory cytokines including TNF-alpha (25461 vs. 1908 pg/ml, p < 0.001), IL-1beta (2921 vs. 1067 pg/ml, p < 0.001) and IFN-gamma (2190 vs 427 pg/ml, p < 0.001) with relative preservation of anti-inflammatory mediators. The suppressive effects on LPS-induced placental inflammation were independent of the timing of PTX administration in relation to LPS-induced stimulation. CONCLUSION: Our study suggests that PTX attenuates the LPS-induced pro-inflammatory milieu in human placental explants. We speculate that PTX may have utility as a candidate anti-inflammatory agent for prophylaxis and/or treatment of human placental inflammation.