Apolipoprotein E localization in rat hepatocytes by immunogold labeling of cryothin sections.
The distribution of apolipoprotein (apo) E in rat hepatocytes was investigated with an affinity-purified polyclonal antibody raised against apoE isolated from hepatogeneous very low density lipoproteins (VLDL). The distribution of this antibody was visualized with colloidal gold complexed to anti-rabbit IgG. By epipolarization microscopy, apoE was found uniformly along the basolateral surfaces of all hepatic parenchymal cells, showing a striking intensity along the sinusoidal front. Punctate deposits of colloidal gold appeared to be randomly distributed within all hepatocytes. Widely scattered Kupffer cells also stained for apoE. Electron microscopic examination of immunogold-labeled cryothin sections showed that hepatocytic microvilli projecting into the space of Disse consistently contained clusters of immunogold. The gold particles were variably associated with evident lipoprotein particles, raising the possibility that apoE alone may bind to receptors or other macromolecules at the surface of hepatocytes. Endosomes near the sinusoidal front and multivesicular bodies in the Golgi/biliary area labeled intensely for apoE, consistent with a high content of apoE associated with triglyceride-rich lipoprotein remnants contained within these organelles. Some but not all nascent VLDL particles within putative forming Golgi secretory vesicles were labeled, but many other Golgi vesicles and cisternae that lacked evident VLDL particles were also labeled. These results suggest that at least some apoE associates with nascent VLDL in forming Golgi secretory vesicles. Unexpectedly, the matrix of all hepatocytic peroxisomes was heavily labeled. Immunoblots with the affinity-purified anti-rat apoE IgG against proteins from highly purified peroxisomes isolated from rat hepatocytes revealed a protein with an apparent molecular mass of 34.5 kDa, similar to that of rat apoE in rat blood plasma. In addition, gold was sometimes found in the area either adjacent to peroxisomes or between multivesicular bodies and the bile canaliculus not evidently associated with a membranous compartment. These observations suggest that apoE may participate in interorganellar cholesterol transport within hepatocytes.
Hamilton
R L
RL
Department of Anatomy, University of California, San Francisco 94143-0130.
Wong
J S
JS
Guo
L S
LS
Krisans
S
S
Havel
R J
RJ
D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity / Q000276:immunology; D001057:Apolipoproteins E / Q000032:analysis*; D005629:Frozen Sections; D015151:Immunoblotting; D007150:Immunohistochemistry; D008099:Liver / Q000737:chemistry* / Q000166:cytology; D008854:Microscopy, Electron; D051381:Rats; D013347:Subcellular Fractions / Q000737:chemistry
D000911:Antibodies, Monoclonal; D001057:Apolipoproteins E