Inherently Radiopaque Narrow-Size-Calibrated Microspheres: Proof of Principle in a Pig Embolization Model.

PURPOSE: To investigate radiopacity, size and size calibration, morphology, and vascular distribution of inherently radiopaque microspheres in vitro and in a pig embolization model. MATERIALS AND METHODS: We compared three types of microspheres: DCBead™ (size 100-300 µm) and Embozene™ (250 µm) as clinically established microspheres, and the prototype Visible (250 µm) that contains additional radiopaque material. Size and size calibration of microspheres were examined by laser diffraction. Pulmonary artery embolization was performed in 12 pigs, and radiopacity was examined by in vitro micro-computed tomography (CT), in vivo cone-beam CT, and ex vivo micro-CT after killing. Morphology and vascular distribution of microspheres were microscopically examined. RESULTS: In in vitro and ex vivo micro-CT, radiopacity of Visible was higher than that of Embozene™, whereas DCBead™ showed no radiopacity. In in vivo cone-beam CT, radiopacity was observed with Visible but not with Embozene™ and DCBead™. Laser diffraction revealed that 7.0% (Visible), 6.5% (Embozene™), and 22.5% (DCBead™) of microspheres were smaller than 223.5 µm. Visible and Embozene™ microspheres were very often located in bronchiolus-associated arteries, but rarely in subsegmental and capillary arteries, whereas DCBead™ were very often and often detected in bronchiolus-associated arteries and capillary arteries, respectively (and rarely in subsegmental arteries). CONCLUSION: After pulmonary artery embolization, Visible but not Embozene™ or DCBead™ provide in vivo radiopacity in cone-beam CT. In contrast to non-narrow-size-calibrated DCBead™, pulmonary artery embolization with narrow-size-calibrated Visible and Embozene™ result in a predictable arterial distribution without embolization-related hemorrhagic lung infarction.

Activation of betagamma subunits of G(i2) and G(i3) proteins by basic secretagogues induces exocytosis through phospholipase Cbeta and arachidonate release through phospholipase Cgamma in mast cells.

Mast cells are activated by Ag-induced clustering of IgE bound to FcepsilonRI receptors or by basic secretagogues that stimulate pertussis toxin-sensitive heterotrimeric G proteins. The cell response includes the secretion of stored molecules, such as histamine, through exocytosis and of de novo synthesized mediators, such as arachidonate metabolites. The respective roles of G proteins alpha and betagamma subunits as well as various types of phospholipase C (PLC) in the signaling pathways elicited by basic secretagogues remain unknown. We show that a specific Ab produced against the C-terminus of Galpha(i3) and an anti-recombinant Galpha(i2) Ab inhibited, with additive effects, both exocytosis and arachidonate release from permeabilized rat peritoneal mast cells elicited by the basic secretagogues mastoparan and spermine. A specific Ab directed against Gbetagamma dimers prevented both secretions. Anti-PLCbeta Abs selectively prevented exocytosis. The selective phosphatidylinositol 3-kinase inhibitor LY 294002 prevented arachidonate release without modifying exocytosis. Gbetagamma coimmunoprecipitated with PLCbeta and phosphatidylinositol 3-kinase. The anti-PLCgamma1 and anti-phospholipase A(2) Abs selectively blocked arachidonate release. Protein tyrosine phosphorylation was inhibited by anti-Gbetagamma Abs, LY294002, and anti PLCgamma1 Abs. These data show that the early step of basic secretagogue transduction is common to both signaling pathways, involving betagamma subunits of G(i2) and G(i3) proteins. Activated Gbetagamma interacts, on one hand, with PLCbeta to elicit exocytosis and, on the other hand, with phosphatidylinositol 3-kinase to initiate the sequential activation of PLCgamma1, tyrosine kinases, and phospholipase A(2), leading to arachidonate release.

Effect of NMDA receptor ligands on mast cell histamine release, a reappraisal.

Natural polyamines have been proposed to induce histamine release from mast cells through a direct interaction with G proteins. Alternatively, the polyamine binding site of ionotropic N-methyl-D-aspartate (NMDA) receptors has been suggested as a target for spermine on mast cells. We reexamined both hypotheses. Incubation of rat peritoneal mast cells with spermine resulted in a concentration-dependent histamine release (EC50 270 microM). Incubation with NMDA receptor agonists, glutamate or NMDA, associated to the co-agonist glycine, did not induce secretion. Western blot experiments did not reveal NMDA R1, R2a, R2b or R2c subunit expression in rat peritoneal mast cell membranes. The NMDA receptor antagonist at the glycine site, L-689,560, did not modify, at relevant concentrations, the spermine-induced secretion. The NMDA receptor antagonists, ifenprodil and LY 235959, and the NMDA channel blocker, MK801, slightly inhibited, at high concentrations, the secretory effect of spermine. The polyamine arcaine, an antagonist of the NMDA receptor polyamine binding site, induced histamine secretion (EC50 350 microM). Both spermine- and arcaine-induced effects were independent upon extracellular calcium and were largely inhibited by treatment of mast cells with pertussis toxin or benzalkonium chloride. The response to spermine and arcaine was prevented by the hydrolysis of sialic acid residues of the cell surface by neuraminidase, and was restored by permeabilization of the plasma membrane with streptolysine-O, indicating that polyamines act intracellularly. These results confirm the involvement of pertussis toxin-sensitive G proteins in the secretory effect of polyamines and demonstrate the absence of NMDA receptors on rat peritoneal mast cells. Nonselective effects of some NMDA receptor ligands on mast cells cannot be excluded.