ISSN:
0021-9541
Keywords:
Life and Medical Sciences
;
Cell & Developmental Biology
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Biology
,
Medicine
Notes:
We have applied a multiple isotope dilution technique to examine junctional permeability of human umbilical vein endothelial cells (HUVEC) in vitro. Primary cultures were grown to confluence on porous Cytodex-3 microcarrier beads, packed into 0.3 ml columns (3 × 106 cells) and perfused at varying flow rates (0.3-1.2 ml/min) with HEPES-buffered Tyrodes solution containing unlabeled cyanocobalamin, insulin, and albumin. Columns were challenged periodically with mixtures of radioactive tracers of different molecular size. Permeability to 22Na+, [57Co]cyanocobalamin (1.3 kD), [125l]insulin (6 kD) or [125l]albumin (66 kD) was assessed relative to [131l]lgG (160 kD, impermeant reference tracer) by comparing column elution profiles. Although the single passage extraction of [125l]albumin by beads alone approximated 40%, the presence of confluent HUVEC rendered these beads effectively impermeable to albumin. High junctional extractions were measured for cyanocobalamin (0.79 ± 0.02, n = 28) and insulin (0.51 ± 0.05, n = 14) in cultures perfused at 0.3-0.4 ml/min, and tracer extraction decreased as perfusion rates increased. Permeability coefficients for cyanocobalamin (9.66 × 10-5 cm/s) and insulin (4.18 × 10-5 cm/s) increased significantly during perfusion with thrombin (10 U/ml) or cytochalasin D (1 μg/ml), whereas permeability to albumin (0.39 × 10-5 cm/s) remained unchanges. Morphological studies, using the glycocalyx stain ruthenium red, revealed that thrombin or cytochalasin D increased the penetration of the stain into junctions between endothelial cells.
Additional Material:
7 Ill.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1002/jcp.1041490112
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