Evaluating the efficacy of subcutaneous C1-esterase inhibitor administration for use in rat models of inflammatory diseases.

CONTEXT: C1-esterase inhibitor (C1-inh) therapy is currently administered to patients with C1-inh deficiency through intravenous injections. The possibility of subcutaneous administration is currently being explored since this would alleviate need for hospitalization and increase mobility and well-being of patients. Recently, it was observed in pigs that C1-inh indeed can effectively be applied by subcutaneous injection. For studies on the effectiveness of C1-inh therapy for other indications than acquired and hereditary angioedema, rats are commonly used as model animal. For rats, however, subcutaneous C1-inh administration has never been investigated. OBJECTIVE: To evaluate the efficacy of subcutaneous C1-inh administration in rats. MATERIALS AND METHODS: Three boli of 100 U/kg human plasma-derived C1-inh were administered to Wistar rats on three consecutive days through subcutaneous injection or intravenous injection. Blood samples were collected from the tail veins 3, 4.5 or 6 h after C1-inh administration for measurement of C1-inh plasma levels. Antigen and activity levels of C1-inh of each plasma sample were determined by means of a specific ELISA. RESULTS: For both C1-inh antigen and C1-inh activity, 21- to 119-fold higher plasma levels were measured after intravenous administration compared with subcutaneous administration. Subcutaneous administration also resulted in C1-inh plasma levels that were less stable and with decreased relative activity. CONCLUSION: These combined results indicate that in rats, subcutaneous injections in the present formulation are not effective as alternative administration route for C1-inh.

Secondary bjerknes forces deform targeted microbubbles.

In this study, we investigated the effect of secondary Bjerknes forces on targeted microbubbles using high-speed optical imaging. We observed that targeted microbubbles attached to an underlying surface and subject to secondary Bjerknes forces deform in the direction of their neighboring bubble, thereby tending toward a prolate shape. The deformation induces an elastic restoring force, causing the bubbles to recoil back to their equilibrium position; typically within 100 µs after low-intensity ultrasound application. The temporal dynamics of the recoil was modeled as a simple mass-spring system, from which a value for the effective spring constant k of the order 10(-3) Nm(-1) was obtained. Moreover, the translational dynamics of interacting targeted microbubbles was predicted by a hydrodynamic point particle model, including a value of the spring stiffness k of the very same order as derived experimentally from the recoiling curves. For higher acoustic pressures, secondary Bjerknes forces rupture the molecular adhesion of the bubbles to the surface. We used this mutual attraction to quantify the binding force between a single biotinylated microbubble and an avidin-coated surface, which was found to be between 0.9 and 2 nanonewtons (nN). The observation of patches of lipids left at the initial binding site suggests that lipid anchors are pulled out of the microbubble shell, rather than biotin molecules unbinding from avidin. Understanding the effect of ultrasound application on targeted microbubbles is crucial for further advances in the realm of molecular imaging.

Lack of complex N-glycans on HIV-1 envelope glycoproteins preserves protein conformation and entry function.

The HIV-1 envelope glycoprotein complex (Env) is the focus of vaccine development aimed at eliciting humoral immunity. Env's extensive and heterogeneous N-linked glycosylation affects folding, binding to lectin receptors, antigenicity and immunogenicity. We characterized recombinant Env proteins and virus particles produced in mammalian cells that lack N-acetylglucosaminyltransferase I (GnTI), an enzyme necessary for the conversion of oligomannose N-glycans to complex N-glycans. Carbohydrate analyses revealed that trimeric Env produced in GnTI(-/-) cells contained exclusively oligomannose N-glycans, with incompletely trimmed oligomannose glycans predominating. The folding and conformation of Env proteins was little affected by the manipulation of the glycosylation. Viruses produced in GnTI(-/-) cells were infectious, indicating that the conversion to complex glycans is not necessary for Env entry function, although virus binding to the C-type lectin DC-SIGN was enhanced. Manipulating Env's N-glycosylation may be useful for structural and functional studies and for vaccine design.