During the bloodstage of malaria infection, the parasite internalizes and degrades massive amounts of hemoglobin from the host red blood cell. Using serial thin-section electron microscopy and threedimensional reconstruction, we demonstrate four independent, but partially overlapping, hemoglobin-uptake processes distinguishable temporally, morphologically, and pharmacologically. Early ring-stage parasites undergo a profound morphological transformation in which they fold, like a cup, onto themselves and in so doing take a large first gulp of host cell cytoplasm. This event, which we term the ''Big Gulp,'' appears to be independent of actin polymerization and marks the first step in biogenesis of the parasite's lysosomal compartment-the food vacuole. A second, previously identified uptake process, uses the cytostome, a well characterized and morphologically distinct structure at the surface of the parasite. This process is more akin to classical endocytosis, giving rise to small (<0.004 fl) vesicles that are marked by the early endosomal regulatory protein Rab5a. A third process, also arising from cytostomes, creates long thin tubes previously termed cytostomal tubes in an actin-dependent manner. The fourth pathway, which we term phagotrophy, is similar to the Big Gulp in that it more closely resembles phagocytosis, except that phagotrophy does not require actin polymerization. Each of these four processes has aspects that are unique to Plasmodium, thus opening avenues to antimalarial therapy.actin ͉ endocytosis ͉ food vacuole ͉ cytosome ͉ transport

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Objective: Develop a test methodology for evaluating the reason for infusion set (IS) failure in persons with diabetes using insulin pump therapy (Continuous Sub-cutaneous Insulin Infusion, CSII). Insulin stability (physical, chemical, and microbiological) and/or the loss of preservatives in the infusate are thought to play significant roles in shortening IS wear time. In-vitro and in-vivo methods were used to assess the cause(s) site inflammation and the loss of preservatives. Method: Fast-acting insulin (insulin aspart and lispro) were pumped through different infusion sets (&gt;4) under simulated-use conditions. The pumped insulin and non-pumped insulin controls were characterized for physical, chemical, and microbiological properties. The pumped insulin, combined with fibrinogen (an initiator for material foreign body responses), were tested in murine macrophage cell culture for inflammatory responses. The corresponding infusion sets were tested in vivo in a diabetic porcine model to determine IS wear-time. Result: Pumped insulins in ≥8 insulin/IS combinations met corresponding criteria in USP insulin monographs, indicating no significant chemical changes. Lower preservative content was found to increase insulin aggregates in-vitro/in-vivo. Pro-inflammatory cytokines (MIP-1α, and MCP-1) increased significantly in the inflammatory in-vitro model. The IS wear-time was shortened from 4.9±0.3 days (control) to 1.7±0.3 days (p&lt;0.0001) with lower preservative content (&lt;50% per label claim). Conclusion: Preliminary findings from in-vitro and in-vivo test model development indicate that macrophage number, the inflammatory response, and device wear time were significantly impacted by loss of preservative and trace aggregates/particles. The new test methodology can further our understanding as to how to improve IS wear time. Disclosure S. Chattaraj: Employee; Self; Medtronic. G. Zhang: None. E. Anselmo: Employee; Self; Medtronic. J.C. Fusselman: None.

This study was funded by Medtronic. Declaration of Financial / Other RelationshipsSC, TK, KS, KW and GZ are employees of Medtronic.JME peer reviewers on this manuscript have received an honorarium from JME for their review work, but have no other relevant financial relationships to disclose.The Medtronic Extended-Wear Infusion Set (branded as Medtronic Extended) is a CE marked device in EU, and an investigational device in the US; it is currently not approved for use in the US.