Intestinal crypt cell survival plays a crucial role in conditions that affect cell turnover as a consequence of their function as the proliferative unit for the intestinal epithelium. Understanding the specific mechanisms that these cells use to inhibit programmed cell death, or apoptosis, will aid in the treatment of diseases in which there is enterocyte loss, such as the short gut syndrome. Regulation of the processes that determine whether a crypt cell undergoes apoptosis or survives relies, in part, on extracellular signals that activate second messengers such as adenosine 3', 5'-cyclic monophosphate (cAMP). Preliminary studies have shown that in the undifferentiated T84 intestinal cell line, cAMP inhibits apoptosis. In addition, we have demonstrated that cAMP activates a known survival pathway in these cells, the extracellular-signal regulate kinase (ERK1/2) cascade. This application will address the hypothesis that cAMP inhibits apoptosis in undifferentiated intestinal epithelial cells through the activation of the ERK1/2 signaling cascade. We propose the following specific aims: 1.) elucidation of the signal transduction pathway involved in cAMP-mediated initiation of ERK1/2 signaling, 2.) establishment of the ERK1/2 cascade as a mechanism for cAMP-induced inhibition of apoptosis, and 3.) the in vivo demonstration of cAMP-induced inhibition of intestinal apoptosis. In vitro studies will use a combination of pharmacological agents and molecular inhibitors to dissect two critical aspects of cAMP-mediated ERK1/2 activation, namely the activation of a cAMP responsive binding protein and the phosphorylation of a Raf kinase isoform at the entrance point of the ERK1/2 cascade. In addition, we will determine the specific survival pathways induced by cAMP to inhibit apoptosis. The in vivo studies will use a mechanistic approach to reverse irradiation induced crypt cell apoptosis by increasing intestinal levels of cAMP and demonstrate the role of cAMP in the protective effects of prostaglandin E2 signaling. These studies will lay the ground work for utilizing cAMP-mediated signaling to enhance intestinal proliferation at the level of the crypt cell and to enhance the adaptive response in short gut syndrome and in other disorders of enterocyte loss.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Clinical Investigator Award (CIA) (K08)
Project #
5K08DK066297-02
Application #
6846545
Study Section
Diabetes, Endocrinology and Metabolic Diseases B Subcommittee (DDK)
Program Officer
Podskalny, Judith M,
Project Start
2004-02-01
Project End
2008-12-31
Budget Start
2005-01-01
Budget End
2005-12-31
Support Year
2
Fiscal Year
2005
Total Cost
$126,091
Indirect Cost
Name
Children's Hospital Med Ctr (Cincinnati)
Department
Type
DUNS #
071284913
City
Cincinnati
State
OH
Country
United States
Zip Code
45229
Rudolph, Jeffrey A; Pratt, Julia; Mourya, Reena et al. (2007) Novel mechanism of cyclic AMP mediated extracellular signal regulated kinase activation in an intestinal cell line. Cell Signal 19:1221-8