OBJECTIVE-An important mechanism in the pathogenesis of type 2 diabetes in obese individuals is elevation of plasma free fatty acids (FFAs), which induce insulin resistance and chronically decrease ␤-cell function and mass. Our objective was to investigate the role of oxidative stress in FFA-induced decrease in ␤-cell function. RESEARCH DESIGN AND METHODS-We used an in vivo model of 48-h intravenous oleate infusion in Wistar rats followed by hyperglycemic clamps or islet secretion studies ex vivo and in vitro models of 48-h exposure to oleate in islets and MIN6 cells.RESULTS-Forty-eight-hour infusion of oleate decreased the insulin and C-peptide responses to a hyperglycemic clamp (P Ͻ 0.01), an effect prevented by coinfusion of the antioxidants N-acetylcysteine (NAC) and taurine. Similar to the findings in vivo, 48-h infusion of oleate decreased glucose-stimulated insulin secretion ex vivo (P Ͻ 0.01) and induced oxidative stress (P Ͻ 0.001) in isolated islets, effects prevented by coinfusion of the antioxidants NAC, taurine, or tempol (4-hydroxy-2,2,6,6-tetramethyl-piperidine-1-oxyl). Forty-eight-hour infusion of olive oil induced oxidative stress (P Ͻ 0.001) and decreased the insulin response of isolated islets similar to oleate (P Ͻ 0.01). Islets exposed to oleate or palmitate and MIN6 cells exposed to oleate showed a decreased insulin response to high glucose and increased levels of oxidative stress (both P Ͻ 0.001), effects prevented by taurine. Real-time RT-PCR showed increased mRNA levels of antioxidant genes in MIN6 cells after oleate exposure, an effect partially prevented by taurine. T ype 2 diabetes is characterized by both insulin resistance and defective insulin secretion (1). Obesity is the major predisposing factor for type 2 diabetes and is associated with excessive release of fatty acids from the expanded adipose tissue mass, leading to elevated plasma free fatty acids (FFAs), which are known to induce insulin resistance (2,3). Acute FFA exposure stimulates insulin secretion (4), but studies in vitro and in situ have shown that prolonged FFA exposure decreases glucose-stimulated insulin secretion (GSIS) (5). The effect of prolonged FFA elevation on ␤-cell function in vivo has been more controversial, as absolute GSIS was found to be increased (6 -8), unchanged (9,10), or decreased (11-13) by FFA. However, in most of these studies ␤-cell function was inadequate to compensate for FFA-induced insulin resistance (9 -13), at least in predisposed individuals (14). Although the mechanisms behind FFA-induced decrease in ␤-cell function are unclear, one possibility points toward oxidative stress. Pancreatic ␤-cells have low antioxidant defenses (15) and are thus susceptible to reactive oxygen species (ROS)-induced decrease in function and viability (16,17). Oxidative stress has been implicated in the decrease in GSIS induced by prolonged exposure to glucose (18,19), which is in many respects similar to that induced by prolonged exposure to FFA (4,20). However, whether oxidative stress plays a role in FFA-...

β-Cell lipotoxicity is thought to play an important role in the development of type 2 diabetes. However, no study has examined its role in type 1 diabetes, which could be clinically relevant for slow-onset type 1 diabetes. Reports of enhanced cytokine toxicity in fat-laden islets are consistent with the hypothesis that lipid and cytokine toxicity may be synergistic. Thus, β-cell lipotoxicity could be enhanced in models of autoimmune diabetes. To determine this, we examined the effects of prolonged free fatty acids elevation on β-cell secretory function in the prediabetic diabetes-prone BioBreeding (dp-BB) rat, its diabetes-resistant BioBreeding (dr-BB) control, and normal Wistar-Furth (WF) rats. Rats received a 48-h iv infusion of saline or Intralipid plus heparin (IH) (to elevate free fatty acid levels ~2-fold) followed by hyperglycemic clamp or islet secretion studies ex vivo. IH significantly decreased β-cell function, assessed both by the disposition index (insulin secretion corrected for IH-induced insulin resistance) and in isolated islets, in dp-BB, but not in dr-BB or WF, rats, and the effect of IH was inhibited by the antioxidant N-acetylcysteine. Furthermore, IH significantly increased islet cytokine mRNA and plasma cytokine levels (monocyte chemoattractant protein-1 and IL-10) in dp-BB, but not in dr-BB or WF, rats. All dp-BB rats had mononuclear infiltration of islets, which was absent in dr-BB and WF rats. In conclusion, the presence of insulitis was permissive for IH-induced β-cell dysfunction in the BB rat, which suggests a link between β-cell lipotoxicity and islet inflammation.

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4207 Introduction: Multiple diagnostic tests are available to evaluate patients with suspected venous thromboembolism (VTE). The diagnosis of VTE involves combining a clinical pre-test probability using validated scoring systems such as the Wells score (Wells, 2000) with laboratory testing and/or diagnostic imaging. The D-dimer assay is a highly sensitive screening test for VTE that has been shown to safely exclude VTE in patients with low pre-test probability (Kearon, 2006). This project was aimed at determining if the SimpliRed D-dimer assay is used appropriately at our institution. More specifically, we wanted to determine if it is being used in patients with low clinical probability of pulmonary embolism (PE) or deep vein thrombosis (DVT) based on the Wells score and in the absence of other variables which could falsely elevate the D-dimer as determined by expert opinion. Methods: A retrospective review of 199 charts from St. Joseph's Hospital in Hamilton, Ontario from March 2007 to April 2007 was performed. Information was obtained on a standardized data collection form. Duplicate data extraction of the first 20 charts was undertaken to ensure concordance. Results: Of the 199 SimpliRed D-dimer assays ordered during this time period, 84 (42%) were inappropriate. Forty (48%) were considered inappropriate because they were ordered in patients with a moderate to high pre-test probability for VTE while 44 (52%) were ordered in patients with a co-existing medical condition that should have precluded the use of the assay including sepsis, pregnancy, active malignancy, and recent major surgery. The D-dimer assay was positive in 51 (61%) of the 84 inappropriate tests and 31 of these patients underwent further testing. Twelve patients were eventually diagnosed with VTE. The remaining 20 patients with a positive D-dimer ordered in an inappropriate context did not undergo further testing to investigate the positive result. Thirty-three (41%) patients with an inappropriately ordered D-dimer had a negative result and did not undergo further investigation. For patients with an appropriately ordered D-dimer assay (115 out of 199 or 58%), 96 had a negative D-dimer result. Of the 96, 28 patients went on to further testing including Doppler ultrasounds, ventilation/perfusion scans, or CT scans. One of the 28 patients had VTE. Conclusions: This retrospective review revealed that 42% of the SimpliRed D-dimer assays ordered at our institution are inappropriate. Thirty three patients identified in this review who had a non-low pretest probability and negative D-dimer did not undergo further testing. A negative D-dimer result in these patients is not sufficient to rule out VTE – the lack of further appropriate testing could have led to missed VTE events. D-dimer assays, including the one used at our institution, have been shown to reliably rule out both DVT and PE in patients with a low pre-test probability (Kearon, 2006). At our institution 28 patients with an appropriately ordered, negative D-dimer underwent further testing that would not be recommended based on current diagnostic algorithms. These investigations are costly and expose the patient to possible unnecessary harm (such as radiation exposure from CT scans or use of anticoagulants until testing can be arranged). Of the patients with a negative D-dimer who went on to have further investigations, 1 was found to have VTE. This patient may have had a higher clinical pre-test probability than was assigned retrospectively from limited chart records. If the pre-test score assigned was accurate, this suggests that the failure rate of the D-dimer for ruling out VTE in a patient with a low pre-test probability is 3.6% (0.1% to 18.3%) in our series, higher than previously reported (Kearon, 2006). Disclosures: Crowther: Pfizer: Consultancy, Research Funding; Leo Pharma: Consultancy, Research Funding; Bayer: Consultancy; CSL: Consultancy; BI: Consultancy, Research Funding; Behring: Consultancy; Octapharma: Consultancy; Artisan: Consultancy.