Isolation and characterization of canalicular and basolateral plasma membrane fractions from human liver.
A method is described for the isolation of subfractions from human liver plasma membranes, enriched in canalicular domains (cLPM) and basolateral domains (blLPM), respectively, and the results are compared to those obtained with rat liver. The studies were performed in 18 human livers. The cLPM (isolated at densities 1.103-1.127 for human and 1.036-1.127 for rat cLPM) from human as well as rat liver showed a lower density than the blLPM (1.141-1.161 for human and 1.151-1.172 for rat blLPM). Human and rat blLPM were characterized by increased levels of (Na+/K+)-ATPase (relative enrichment 33 and 21, respectively). Both human and rat cLPM showed high specific activities of leucine aminopeptidase; relative enrichment factors were 42 and 31, respectively. Mg(2+)-ATPase and alkaline phosphatase, specific canalicular enzymes in rat liver, were only slightly enriched in the cLPM of human liver, which indicates that these enzymes are not suitable as marker enzymes for human liver cLPM. Both cLPM and blLPM of human and rat origin were only slightly contaminated with mitochondria, lysosomes, Golgi membranes and endoplasmic reticulum. Total recoveries of cLPM and blLPM were 0.02 mg protein/g liver each for the human membrane preparations, compared to 0.07 and 0.16 mg protein/g liver for the membranes prepared from rat liver. Analysis of membrane fluidity revealed that the human liver cLPM were more rigid than blLPM (mean difference in fluorescence polarization PDPH 0.024). They contained more cholesterol (0.43 vs. 0.30 mumol/mg protein) and phospholipids (0.54 vs. 0.39 mumol/mg protein, respectively), which was compatible to rat liver plasma membrane fractions. This study shows that besides similarities, there are several differences between human and rat liver plasma membrane fractions.
Department of Pediatrics, University of Groningen, The Netherlands.
H
H
Wolters
M
M
Spiering
A
A
Gerding
M J
MJ
Slooff
F
F
Kuipers
M J
MJ
Hardonk
R J
RJ
Vonk
D000293:Adolescent; D000328:Adult; D000368:Aged; D000818:Animals; D002458:Cell Fractionation / Q000379:methods; D002462:Cell Membrane / Q000737:chemistry* / Q000201:enzymology / Q000648:ultrastructure; D002499:Centrifugation, Density Gradient; D002648:Child; D002675:Child, Preschool; D005260:Female; D006801:Humans; D008099:Liver / Q000166:cytology* / Q000201:enzymology / Q000648:ultrastructure; D008297:Male; D008563:Membrane Lipids / Q000737:chemistry; D008875:Middle Aged; D051381:Rats; D011919:Rats, Inbred Strains
D008563:Membrane Lipids