Development of postoperative peritoneal adhesions phenomenologically involves a complex array of interacting cytokines and growth factors that are produced as a result of differential gene expression induced by hypoxia. Reactive radicals are produced during hypoxia and remain even after normoxia is restored. Among them is nitric oxide (NO), which is generated by NO synthases. Our long-term goal is to prevent or selectively limit the development of postoperative peritoneal adhesions. The objective of this application is to identify the mechanisms that underlie their development. The central hypothesis is that hypoxia modulates inducible nitric oxide synthase (iNOS), further resulting in reduced production of NO, which contributes to pro-inflammatory signaling, thus leading to the development of postoperative adhesions. The hypothesis has been formulated on the basis of strong preliminary and published data, which suggest that hypoxia causes cells to acquire an adhesion phenotype that is pathogenetically central to the process. There will be four specific aims, which will determine: 1) the biological effects of nitric oxide on the formation and development of the peritoneal adhesion phenotype;2) the mechanisms by which hypoxia decreases nitric oxide production in peritoneal cells;3) the in vivo effects of nitric oxide on postoperative adhesion development;and 4) the causal mechanism responsible for induction of, and protection against, postoperative adhesions. Both in vitro (human and rat) and in vivo (rat cecal abrasion model) studies will be conducted. Gene expression, iNOS activity, cellular apoptosis, production of extracellular matrix, tissue remodeling and cellular proliferation will be assayed as they relate to the production of adhesions. The applicants are particularly well prepared to undertake the proposed research because of extensive experience with the systems that will be employed, coupled with the fact that all necessary reagents and technologies needed are already in place. The proposed work is innovative, because of the combined cellular and molecular biological approaches that will be applied quantitatively, both in vitro and in vivo, thereby, reducing complexity and fostering identification of the relevant pathogenetic mechanisms. The outcomes are expected to collectively identify decreased production of NO as a result of hypoxia as the critical pathologic change, resulting in decreased apoptosis and tissue remodeling in concert with increased production of extracellular matrix, thereby, promoting the development of severe adhesions. These results are expected to be significant, because, once the molecular mechanisms are known, specific pathways can then be targeted selectively targeted to limit the underlying adhesion development processes. This, in turn, is expected to lead to innovative, new approaches to the prevention and treatment of peritoneal adhesions.
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