Regulation of gene expression lies at the very heart of the immune response. Delineating the basic mechanisms of how specific genes become upregulated improves our understanding of the immune response to injury and infection. The chemokines, including IL-8, have become the focus of intense research because of their wide-ranging biological properties. Our previous work has demonstrated that the IL-8 gene is closely regulated by intracellular reactive oxygen intermediates (R01). In this competing renewal we will test the hypothesis that the CXC chemokines, including IL-8, are induced by R01 in multiple different primary cultures. The IL-8 gene has unique sequences, which allow its upregulation by intracellular R01. Our long-term objective is to define the R01 interactions, which lead to induction of the chemokines. In the first specific aim we will extend our previous observations that anti-oxidants inhibit IL-8 by determining if other chemokines show similar properties. We will specifically evaluate both CXC and CC chemokines. The second specific aim will determine if oxidant regulation occurs in primary cultures of T cells, B cells, NK cells, monocytes and neutrophils. This will determine if there is a direct effect of anti-oxidants on chemokine production or if it is occurring through complex cell-cell interactions. We have previously observed that the IL-8 mRNA may be extremely stable under certain conditions. This has been confirmed in disease states where there are elevated levels of IL-8 for prolonged periods. Therefore, our third specific aim will define the stimuli which account for the production of this stable mRNA. The fourth specific aim will closely evaluate the gene regulatory sequences in the 5' end of the IL-8 gene to identify the oxygen responsive elements. Since antioxidants decrease IL-8 but have no effect on or increase IL-6, we will use both 5' promoter regions to identify the cis gene regulatory sequences that account for this differential gene regulation. Successful completion of our specific aims will provide substantial insight into the regulation of chemokine gene expression and improve our understanding of basic mechanisms of disease.
