The overall objective of the work proposed in this application and accomplished during the previous 16 years the grant has been active is to define and understand the mechanisms regulating GI adaptation. During the past 5 years, we have concentrated our studies on the adaptations that occur during normal development and in response to mucosal damage, specifically the process of healing. These studies have shown that polyamines are involved in both processes and that they are required for the healing of mucosal stress erosions. The current proposal describes studies directed towards elucidating the role of polyamines in the process of mucosal healing. The work is divided into 8 specific aims that involve studies utilizing the rat stress ulcer model or the IEC-6 culture system. Using the rat stress ulcer model, we will 1) determine the cellular and intracellular localization of ornithine decarboxylase (ODC) before and after damage; 2) determine whether polyamines are essential to healing by virtue of their ability to take part in protein cross-linking; and 3) determine whether the activity of transglutaminase (TGase) which catalyzes protein cross-linking is altered during injury or healing. Using both the stress ulcer model and IEC-6 cells in vitro, we will 4) determine the role of polyamines in the regulation of TGase activity, and 5) elucidate whether the control of TGase is at the transcriptional level. We then plan to use the IEC-6 cells to 6) establish a methodology for measuring cell migration in vitro, and 7) determine the role of polyamines, protein cross-linking and TGase in cell migration. finally, we will 8) use a variety of synthetically modified polyamine analogues to determine the structure- function relationships that are important for the polyamine involvement in healing, TGase activity and cell migration. This last specific aim will provide significant insight into the mechanism of action of polyamines at the molecular level. We know that mucosal healing involves both an early phase of cell migration and then cell division to replace lost cells. Our data indicate that polyamines are essential to both. The actual mechanisms involved in these processes, and especially in cell migration in the injured mucosa are virtually unstudied. Investigation of the role of polyamines in these processes should provide important information about this clinically significant event.
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