Glucose deprivation promotes chemoresistance to gemcitabine through an Akt-dependent mechanism Michael Abramson MD, Edward W Swanson BAS, Edward E Whang MD, FACS Brigham and Women’s Hospital, Boston, MA INTRODUCTION: Pancreatic cancer is characterized by profound chemoresistance. The role of microenvironmental conditions in me- diating chemoresistance is poorly understood. We hypothesized that nutrient deprivation, a feature of the cancer microenvironment, ac- tivates signaling mechanisms that contribute to chemoresistance in pancreatic cancer. METHODS: MiaPaCa2 pancreatic cancer cells were cultured in glucose-containing media (control) or glucose-deprived media (450mg/dL versus 0 mg/dL glucose concentration). At 24 hours, cells were assayed for total Akt and phospho-Akt (serine 473) expres- sion (Western blotting). In chemosensitivity studies, MiaPaCa2 cells were cultured in 1) glucose-containing media, 2) glucose-deprived media, or 3) glucose-deprived media containing the PI3-kinase in- hibitor, LY 294002 (25 M) for 24 hours. Cells were then treated with gemcitabine (100 ng/ml) in glucose-containing media for 48 hours. Cell viability was assayed using the MTS assay. RESULTS: Glucose deprivation was associated with Akt activation, with a greater than 4-fold upregulation of Akt phosphorylation ver- sus control (p0.01). Total Akt expression level did not vary with glucose concentration. Exposure to glucose deprivation resulted in a 78% reduction in cell death following gemcitabine administration (p0.05). This glucose deprivation-induced increase in resistance to gemcitabine was abrogated by addition of the PI3-kinase inhibitor LY 294002. CONCLUSIONS: Our results suggest that glucose-deprivation, a feature of the cancer microenvironment, promotes chemoresistance to gemcitabine through activation of PI3-kinase/Akt signaling. Our findings also highlight the importance of studying cancer chemo- therapy efficacy under physiological conditions. Failure to account for these conditions may contribute to our failure to have made clinically meaningful advances in therapy for this deadly cancer. Inhibition of p38 MAP kinase as a target for prophylaxis of postoperative ileus SvenWehner PhD, Stefan Straesser, Mariola Lysson, Andreas Hirner MD, Joerg C Kalff MD, FACS University of Bonn, Bonn, Germany INTRODUCTION: Postoperative ileus is mediated by a severe in- flammation of the tunica muscularis. Resident muscularis macro- phages are major players in the initiation of this inflammation. We investigated if p38 MAP Kinase activation is involved in genesis of postoperative ileus and if its inhibition by the macrophage specific inhibitor semapimod prevents intestinal dysmotility. METHODS: Postoperative ileus was induced by manipulation of the small bowel. p38 MAPK activation was assessed by immunoblotting of muscularis externa preparations. Proinflammatory gene expres- sion was quantified by real-time PCR. Myeloperoxidase histochem- istry for neutrophils was performed in jejunal segments. Nitric oxide production was measured by Griess reaction in smooth muscle organ culture supernatants. Jejunal contractility was assessed within an or- gan bath setup. Intestinal motility was analyzed by gastrointestinal transit measurement. RESULTS: Strong p38MAPK phosphorylation was observed im- mediately after intestinal manipulation. Semapimod treatment led to a significant decrease of (1) p38MAPK phosphorylation, (2) proin- flammatory gene expression of MIP-1alpha, IL-6, MCP-1 and ICAM-1, (3) neutrophil infiltration and (4) nitric oxide production within the tunica muscularis. Subsequently, semapimod prevented the suppression of smooth muscle contractility and intestinal motil- ity after intestinal manipulation. CONCLUSIONS: A single preoperative semapimod administration prevents intestinal macrophage activation by abdominal surgery and subsequent gastrointestinal dysmotility. Semapimod acts both, via inhibition of p38MAPK and NO production in macrophages and seems to be a promising strategy for prophylaxis of postoperative ileus. Rapid upregulation of the Na/glucose cotransporter SGLT1 in response to an unexpected glucose load Adam T Stearns MD, Anita Balakrishnan MD, David B Rhoads PhD, Stanley W Ashley MD, FACS, Ali Tavakkolizadeh MD Brigham and Women’s Hospital, Boston, MA INTRODUCTION: The intestinal Na+/glucose cotransporter SGLT1 performs all active transport of glucose from the bowel lu- men. As such, SGLT1 is a potential therapeutic target for obesity and associated diabetes. Long-term dietary regulation and anticipatory diurnal regulation by nutrient intake rhythms are well characterized. We now describe a third, short-term regulatory pathway in rodents. METHODS: Male Sprague-Dawley rats (200g, n=12) were accli- matized to a 12-hour light-dark cycle (lights-on 7am). Rats were anesthetized at 11am, laparotomized, and randomly assigned to re- ceive duodenal infusion of either 250mM D-glucose or water (5mL/h, 3 h) via a polyethylene cannula inserted though a gastric incision. Jejunal gene expression was assessed in mucosal scrapings. SGLT1 and facilitated glucose transporter GLUT2 proteins were quantified by immunoblotting of whole-cell protein extracts and densitometry. SGLT1, GLUT2 and SGLT1 regulatory subunit RS1 mRNA abundances were quantified by reverse transcription of ex- tracted RNA and qPCR, normalized to actin. RESULTS: Total mucosal SGLT1 protein content increased signif- icantly following glucose infusion (2.9-fold, p0.02) with no change in SGLT1 mRNA abundance (0.9-fold, p0.3). In contrast, glucose infusion had no effect on GLUT2 protein (1.1-fold, p0.2) or mRNA (0.85 fold, p0.6). RS1 mRNA abundance was also un- changed. CONCLUSIONS: Our results document the existence of a third regulatory pathway for SGLT1, characterized by a rapid increase in S12 Surgical Forum Abstracts J Am Coll Surg