Confluent monolayers of the prostatic cell line, JHU-4 were infected with a range inocula (1,5-150 IFU/cell) using described protocols. Immunofluorescence analysis of Cm infected PEC were performed using a monoclonal antibody against Chlamydial LPS-FITC. Supernatants were collected at different times until 72 h post infection (p.i) and assayed for nitric oxide (NO) production by the Griess reaction. Total cellular RNA was extracted from infected and non-infected PEC and qualitative RT-PCR for iNOS, IL-8, MCP-1 and â-actin were performed. Flow cytometry analysis of PEC ICAM-1 and MHC class I expression was assayed using specific monoclonal antibodies.
Immunofluorescence assays using antibodies against Chlamydial LPS showed that Cm can infect and complete its biologic cycle in PEC efficiently. Flow cytometry analysis demonstrated that infection of PEC with Cm increased ICAM-1 and MHC class I expression on PEC. Increased levels of NO in culture supernatants were first detected at 24 h p.i, and reached a maximun at 72 h p.i. Infection of PEC with Cm increased the levels of iNOS, IL-8 and MCP-1 mRNA studied at 24 and 48 h p.i.
These results show that Cm is able to infect in vitro PEC, which respond inducing early changes such as an increase of ICAM-1, MHC class I and iNOS mRNA expression, and also an induction of chemokines mRNA expression that suggest a pivotal role for PEC in the defence against chlamydial infection.
