Convection of macromolecules is the dominant mode of transport across horizontal 0.4- and 3-microns filters in diffusion chambers: significance for biologic monolayer permeability assessment.
The purpose of the present study was to evaluate the permeability characteristics of fluorescein-labeled hydroxyethyl starch (FITC-HES, 20 less than molecular radius, aE less than 111 A) across large pore (0.4- and 3-microns polycarbonate) filters used in endothelial cell monolayer diffusion studies. Although the apparent permeability (P) of the FITC-HES macromolecules across these filters declined as the molecular radius increased, this decline was less than that associated with each solute's free diffusion (D0). Thus, the P/D0 anomalously increased as aE increased, a pattern not seen for free diffusion (flat P/D0 with increased aE) or restricted diffusion (decline in P/D0 with increased aE). Substantial natural convection across the porous filters produced the anomalous P/D0 curve for the following reasons: (1) this effect was elevated with positive pressure and ameliorated by zero driving pressure, and (2) the effect was much greater across filters with large (3 microns) vs small (0.4 microns) pore sizes. In addition, we estimated that less than half of macromolecular transport above 50 A probe radius at zero transmural pressure arrives by diffusion. The findings suggest that the property of restricted diffusion of biologic layers on these filters will be artifactually exaggerated when the measured resistance of the filter is subtracted from that of the filter plus biologic monolayer. Remedies for this problem may include using smaller pore filters, filters layered with extracellular material, or structural methods to determine true filter permeability.
Katz
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MA
Benjamin W. Zweifach Microcirculation Laboratories, Tucson Veterans Affairs Medical Center, Arizona 85723.
Schaeffer
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RC
D002463:Cell Membrane Permeability; D015194:Diffusion Chambers, Culture; D004730:Endothelium, Vascular / Q000378:metabolism; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D005456:Fluorescent Dyes; D006358:Hot Temperature; D006895:Hydroxyethyl Starch Derivatives / Q000378:metabolism; D046911:Macromolecular Substances; D008851:Micropore Filters; D013861:Thiocyanates
D005452:Fluoresceins; D005456:Fluorescent Dyes; D006895:Hydroxyethyl Starch Derivatives; D046911:Macromolecular Substances; D013861:Thiocyanates; D016650:Fluorescein-5-isothiocyanate