Structural heterogeneity of hepatocyte "tight" junctions: a quantitative analysis.

Abstract

Freeze fracture analyses of hepatocyte tight junctions reveal interconnecting strands which represent a semipermeable blood-bile barrier. In epithelia, strand number can be correlated with transepithelial resistance and junctional permeability, but a functional relationship is not always seen. We have devised an analytical method to measure accurately strand number as well as junction depth over a large area of junction. Measurements were obtained in five bile fistula rats and two marine elasmobranchs (little skates). In addition, choleretic infusions of 120 μmoles of taurodehydrocholate were administered to four additional rats over 1 hr to determine if functional changes in ion and fluid movement are associated with alternations in junctional structure. Using a microcomputer with a sonic digitizer, strand number and junction depth were obtained at intervals of 0.1 to 0.2 μ from transmission electron micrographs of fractures of rat and skate liver. Strand number averaged 4.1 ± 1.1 in control rats (798 measurements) and 4.1 ± 1.3 in the taurodehydrocholate-stimulated animals (296 measurements). Junction depths averaged 0.148 ± 0.092 μm in controls and increased to 0.167 ± 0.121 μm in taurodehydrocholate-stimulated rats, a difference which reached significance at the p < 0.06 level. Similarly, values were obtained in skate livers where central zone pigment granules facilitated dissection of portal and central portions of lobule. Central zone strand number and junction depth averaged 5.1 ± 1.6 and 0.178 ± 0.100 μm, respectively (439 measurements), and were indistinguishable from values obtained from portal regions −5.0 ± 1.9 and 0.182 ± 0.099 μm, respectively (217 measurements). Although no quantitative differences in junction structure was observed in these studies, considerable variation was noted in junction depth (0.013 to 0.670 μm in the rat and 0.008 to 0.600 μm in skates) and number. We conclude that the anatomical barrier which forms the normal hepatocyte tight junction is a highly varied structure. If junctional structure as revealed by freeze fracture analysis correlates with changes in permeability, then permeability should vary considerably at multiple points along the surface of individual hepatocytes and would not be detected by present quantitative approaches which average junctional strand number and depth. Alternatively, regulation of junctional permeability may be more dependent on structural elements which are beyond the resolution provided by freeze fracture technique.