Gap junctions contain different amounts of cholesterol whichundergo unique sequestering processes during fiber cell differentiationin the embryonic chicken lens

[摘要] Purpose: To determine the possible changes in the distribution ofcholesterol in gap junction plaques during fiber cell differentiationand maturation in the embryonic chicken lens. The possible mechanism bywhich cholesterol is removed from gap junction plaques is alsoinvestigated.Methods: Filipin cytochemistry in conjunction with freeze-fractureTEM was used to visualize cholesterol, as represented byfilipin-cholesterol complexes (FCCs) in gap junction plaques.Quantitative analysis on the heterogeneous distribution of cholesterolin gap junction plaques was conducted from outer and inner corticalregions. A novel technique combining filipin cytochemistry withfreeze-fracture replica immunogold labeling (FRIL) was used to labelCx45.6 and Cx56 antibodies in cholesterol-containing gap junctions.Filipin cytochemistry and freeze-fracture TEM and thin-section TEM wereused to examine the appearance and nature of the cholesterol-containingvesicular structures associated with gap junction plaques.Results: Chicken lens fibers contain cholesterol-rich,cholesterol-intermediate and cholesterol-free gap junction populationsin both outer and inner cortical regions. Filipin cytochemistry and FRILstudies confirmed that cholesterol-containing junctions were gapjunctions. Quantitative analysis showed that approximately 86% of gapjunctions in the outer cortical zone were cholesterol-rich gapjunctions, whereas approximately 81% of gap junctions in the innercortical zone were cholesterol-free gap junctions. A number ofpleiomorphic cholesterol-rich vesicles of varying sizes were oftenobserved in the gap junction plaques. They appear to be involved in theremoval of cholesterol from gap junction plaques through endocytosis.Conclusions: Gap junctions in the young fibers are enriched withcholesterol because they are assembled in the unique cholesterol-richcell membranes in the lens. A majority of cholesterol-rich gap junctionsin the outer young fibers are transformed into cholesterol-free ones inthe inner mature fibers during fiber cell maturation. A distinctendocytotic process appears to be involved in removing cholesterol fromthe cholesterol-containing gap junctions, and it may play a major rolein the transformation of cholesterol-rich gap junctions intocholesterol-free ones during fiber cell maturation.

[学科分类] 生物化学/生物物理