Term trophoblasts are known to both produce and respond to a variety of cytokines in response to infection. However, comparatively little is known about the capacity of first trimester trophoblasts to respond to an infection during the early gestation of the embryo and what role if any, these cells play in coordinating an effective immune response. This study was undertaken to better characterize the in vitro expression of the proinflammatory cytokines G-CSF and RANTES in response to lipopolysaccharide (LPS). Confluent cultures of the first trimester trophoblast cell line (HTR-8/SVneo) were exposed to E. coli serotype 055.B5 LPS (1 μg/ml) for 0, 2, 4, 6, 8 and 24h in DMEM-Ham's F-12 media supplemented with 10% fetal bovine serum. Culture supernatants were collected and rendered free from cellular debris by centrifugation. G-CSF and RANTES protein levels were measured using an ELISA validated for culture supernatant (R&D Systems, Minneapolis, MN). Sensitivities of the assays were 7.2 pg/ml and 2.5 pg/ml respectively. All standards and test samples were assayed in duplicate and the results averaged. All culture media was tested for the presence of contaminating endotoxin prior to LPS exposure using a Limulus assay with a sensitivity of 0.06 to 0.10 ng/ml (Endotect t m , ICN Biomedicals, Inc., Auora, Ohio). Levels of immunoreactive G-CSF protein showed a steady time-dependent increase in concentration to 24 hours in LPS treated cells with a greater than 600-fold increase in protein levels ovver non-LPS exposed cells at 24h (p < 0.005). Conversely, there was essentially no constitutive or induced G-CSF expression in non-LPS treated cells. Levels of immunoreactive RANTES protein in LPS induced cells increased at a low time-dependent steady state level to 8h in comparison with non-LPS induced cells. There was a nearly 12-fold increase in protein concentration by 24h in LPS treated cells over non-treated controls (p < 0.05). Both induced and non-induced cells showed a low level of constitutive RANTES expression. The results of this study demonstrate the LPS induced expression of the proinflammatory cytokines G-CSF and RANTES, and further support the contention that first trimester trophoblasts participate in cytokine based immune signaling in response to infection.