ovalbumin and lead-chloride

Lead (Pb) is known to preferentially suppress the activation and development of type-1 CD4+ helper T cell (Th1) responses, whereas it enhances the development of type-2 CD4+ helper T cell (Th2) responses. The inhibition of interferon-gamma (IFNgamma) production has been demonstrated in vitro with a Th1 clone and DO11.10 ovalbumin-transgenic (OVA-tg) CD4+ T cells, and in vivo with wild-type and OVA-tg BALB/c mice; however, the mechanisms responsible for the Pb-induced downregulation of IFNgamma have not been reported. Here, we assessed the modulation of IFNgamma production at the mRNA and protein levels. Pb did not significantly affect IFNgamma mRNA expression by a Th1 clone or activated splenocytes, as measured by reverse transcriptase-polymerase chain reaction (RT-PCR), ribonuclease protection, and real-time RT-PCR. However, Pb did significantly lower the amount of IFNgamma protein in supernatants and cell lysates of antigen-activated T cells in comparison to stimulated controls, suggesting that the lower amounts of IFNgamma released into culture supernatants were not due to a blockage of secretion that gave rise to a cytoplasmic accumulation of IFNgamma. Pb inhibition also was not prevented by addition of zinc or iron. Pb did not enhance protein degradation of IFNgamma, in that lactacystin, an effective blocker of proteosomal proteolysis, did not prevent loss of IFNgamma; additionally, Pb did not accelerate loss of IFNgamma after cycloheximide treatment. Pb did, however, significantly suppress IFNgamma biosynthesis, as investigated using 35S-incorporation in pulse/chase experiments, although it did not suppress total protein synthesis, indicating that Pb selectively inhibits IFNgamma biosynthesis. Thus, Pb appears to selectively interfere with the translation of certain proteins, such as IFNgamma. IL-12 blocked Pb's preferential promotion of Th2 cells, but absence of STAT6 did not prevent the Pb skewing. Thus, Pb may modulate unique regulatory pathways.

Topics: Animals; Cell Line; Chlorides; Down-Regulation; Ferrous Compounds; Immunity, Cellular; Immunity, Innate; Interferon-gamma; Interleukin-12; Interleukin-4; Lead; Male; Mice; Mice, Inbred BALB C; Mice, Transgenic; Ovalbumin; Proteasome Endopeptidase Complex; Protein Biosynthesis; RNA Processing, Post-Transcriptional; RNA, Messenger; Spleen; STAT4 Transcription Factor; STAT6 Transcription Factor; Th1 Cells; Th2 Cells; Time Factors; Zinc Compounds

The hypothesis that lead disturbs gut immune functions upon oral ingestion was tested. Long-term exposure to oral PbCl(2)for 10 days caused persistent downregulation of TGF-beta mRNA levels in intestinal tissue. PbCl(2) also disturbed oral tolerance induction to the dietary antigen ovalbumin. Upon challenge with an immunizing dose of ovalbumin and rechallenge of draining lymph node cells in vitro, tolerance induction was partially suppressed in animals exposed to oral PbCl(2). This was shown by increased proliferation to antigenic stimulus, increased production of IFN-gamma and decreased secretion of TGF-beta. In conclusion, we show for the first time that oral exposure to PbCl(2)has a significant effect on the gut immune system, demonstrated by a bias of the cytokine pattern towards Th(1)and by disturbed oral tolerance mechanisms.

Topics: Administration, Oral; Animals; Cells, Cultured; DNA Primers; Down-Regulation; Enzyme-Linked Immunosorbent Assay; Female; Immunity; Interferon-gamma; Interleukin-10; Interleukin-12; Intestine, Small; Lead; Lymph Nodes; Mice; Mice, Inbred NOD; Mucous Membrane; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Th1 Cells; Th2 Cells; Time Factors; Transforming Growth Factor beta