Evaluation of Platelet Dysfunction In Children: Improved Detection and Efficiency

Abstract 1446 Currently, there is no single assay that will detect platelet function abnormalities in all individuals. We prospectively studied 369 patients with a strong history of bruising and mucosal membrane bleeding for possible platelet dysfunction following documentation of normal Von Willebrand antigen and activity levels. In an effort to evaluate platelet function under more diverse conditions we chose to simultaneously evaluate 1) aggregation using platelet rich plasma and light transmission aggregometry (LTA), 2) adhesion under high shear using the Platelet Function Analyzer (PFA-100, ADP-collagen, and Epinephrine-collagen cartridges) and 3) platelet initiated clot formation using heparinized whole blood and the two standard mapping agonists, arachadonic acid (AA) and ADP (Hemascope Thromboelastograph Analyzer). Of the 369 platelet evaluations performed, 87 patients (24%) were found to be normal in all three assays with all agonists. On repeated assays with increased attention to medication and food history, an additional 152 patients were felt to have a transient or acquired dysfunction which improved or normalized on further testing. Leaving 130 patients with persistent bleeding and documented abnormalities on one or more of the assays used. There were no patients with only Collagen (CN) or arachadonic acid (AA) aggregation alone on LTA. Using LTA, there was one patient with combined CN and ADP aggregation defect only, another with AA and EPI absent aggregation only, and one with only AA and Thrombin receptor agonist peptide (TRAP) aggregation abnormalities. A summary of the remaining 127 patients are displayed in the table below:Abnormal AssayLTA EPILTA ADPLTA ADP + EPILTA ADP, EPI CNLTA ADP, EPI, AALTA ADP, EPI CN, AAPFA and PLT Mapping ONLYNumber201240841825M/F3M/17F6M/6F16M/24F1M/7F1M/3F8M/10F12M/13FPFA1M/2F4M7M/4F01M01M/3FPlt Mapping2M/2F1M/2F3F1F009M/7FBoth PFA and Mapping1M0001F2M/9F2M/3F Summary: By using a combination of three assays, we were able to identify 25 additional patients with significant platelet dysfunction detected with abnormal platelet mapping or PFA-100 despite normal light transmission aggregometry. Patients with the most abnormalities on aggregation, also demonstrated abnormal adhesion and platelet initiated clot formation. However, the use of PFA-100 and/or platelet mapping alone would miss the majority of patients with aggregation defects. In the future, the unique combination of platelet function defects as measured by these assays, and future technologies, will not only improve detection, but also facilitate phenotype to genotype associations and expedite mutational analysis. Disclosures: Off Label Use: Rituximab to treat ITP.

Bacterial Lipopolysaccharide Induces an Endocrine Switch from Prostaglandin F2α to Prostaglandin E2 in Bovine Endometrium

Escherichia coli infection of the endometrium causes uterine disease after parturition and is associated with prolonged luteal phases of the ovarian cycle in cattle. Termination of the luteal phase is initiated by prostaglandin F2α (PGF) from oxytocin-stimulated endometrial epithelial cells. Compared with normal animals, the peripheral plasma of animals with E. coli infection of the endometrium had higher concentrations of lipopolysaccharide (LPS) and prostaglandin E2 (PGE) but not PGF. Endometrial explants accumulated predominantly PGE in the culture medium in response to LPS, and this effect was not reversed by oxytocin. Endometrial cells expressed the Toll-like receptor 4/CD14/MD-2 receptor complex necessary to detect LPS. Epithelial and stromal cells treated with LPS had higher steady-state media concentrations of PGE rather than PGF. Arachadonic acid is liberated from cell membranes by phospholipase 2 (PLA2) enzymes and converted to prostaglandins by synthase enzymes. Treatment of epithelial and stromal cells with LPS did not change the levels of PGE or PGF synthase enzymes. However, LPS stimulated increased levels of PLA2 group VI but not PLA2 group IV C immunoreactive protein in epithelial cells. Endometrial cells expressed the E prostanoid 2 and E prostanoid 4 receptors necessary to respond to PGE, which regulates inflammation as well as being luteotropic. In conclusion, LPS detection by endometrial cells stimulated the accumulation of PGE rather than PGF, providing a mechanism to explain prolonged luteal phases in animals with uterine disease, and this PGE may also be important for regulating inflammatory responses in the endometrium.