Поняття “леґітимність” та “леґітимація” як теоретичні інновації М.Вебера: зміст і роль у сучасній соціології

Forssman antigen, a neutral glycosphingolipid carrying five monosaccharides, was localized in epithelial MDCK cells by the immunogold technique. Labeling with a well defined mAb and protein A-gold after freeze-substitution and low temperature embedding in Lowicryl HM20 of aldehyde-fixed and cryoprotected cells, resulted in high levels of specific labeling and excellent retention of cellular ultrastructure compared to ultra-thin cryosections. No Forssman glycolipid was lost from the cells during freeze-substitution as measured by radio-immunostaining of lipid extracts. Redistribution of the glycolipid between membranes did not occur. Forssman glycolipid, abundantly expressed on the surface of MDCK II cells, did not move to neighboring cell surfaces in cocultures with Forssman negative MDCK I cells, even though they were connected by tight junctions. The labeling density on the apical plasma membrane was 1.4-1.6 times higher than basolateral. Roughly two-thirds of the gold particles were found intracellularly. The Golgi complex was labeled for Forssman as were endosomes, identified by endocytosed albumin-gold, and lysosomes, defined by double labeling for cathepsin D. In most cases, the nuclear envelope was Forssman positive, but the labeling density was 10-fold less than on the plasma membrane. Mitochondria and peroxisomes, the latter identified by catalase, remained free of label, consistent with the notion that they do not receive transport vesicles carrying glycosphingolipids. The present method of lipid immunolabeling holds great potential for the localization of other antigenic lipids

Palmitic acid and linoleic acid metabolism in Caco-2 cells: Different triglyceride synthesis and lipoprotein secretion

Polarized monolayers of intestinal Caco-2 cells were used to study the effects of saturated palmitic acid (16:0) and polyunsaturated linoleic acid (18:2) on triglyceride synthesis and lipoprotein secretion. Monolayers were incubated for 24 h, at the apical or lumenal side, with palmitic acid (16:0) or linoleic acid (18:2) in physiological concentrations. Incubation with 1.0 mM 16:0 or 18:2 resulted in differences in the composition and amount of secreted lipoproteins. Radiolabeled lipids in the lipoproteins secreted during incubation with 18:2 were found in the chylomicron/VLDL (very low density lipoprotein) density whereas with 16:0 the secreted lipoproteins were in the intermediate density/low density lipoprotein (IDL/LDL) density range. More triglyceride was secreted into the (basolateral) medium during incubation with 1.0 mM 18:2 (41 ± 12% of total triglyceride synthesized) than with 1.0 mM 16:0 (18 ± 3% of total). The biochemical findings correlate with conspicuous morphological changes in the cells in the presence of 16:0, but not 18:2. Increasing concentrations of 16:0 (0.1-1.8 mM) caused gradual accumulation of intracellular membrane. Microvilli became strongly reduced in number. With 18 mM palmitic acid we found an increased incorporation of [1- 14C]palmitic acid into phosphatidic acid (14.8% of total incorporation into phospholipid with 16:0 vs. <0.5% with 18:2) and diacylglycerol (12.5% with 16:0 vs 0.5% with 18:2) and the amount of intracellular phospholipid doubled. The morphological changes were completely reversed after 24 h with 1.0 mM 18:2. We conclude from our results that, compared to 18:2, 16:0 is not efficiently incorporated into triglycerides. 16:0 is incorporated into cellular phospholipids in a greater proportion than 18:2, causing accumulation of intracellular phospholipid and the precursors phosphatidic acid and diacylglycerol. Different processing of 18:2 and 16:0 by Caco-2 cells resulted in profound differences in triglyceride synthesis and lipoprotein composition and secretion

MHV-A59 enters polarized murine epithelial cells through the apical surface but is released basolaterally

AbstractCoronaviruses have a marked tropism for epithelial cells. Entry and release of the porcine transmissible gastroenteritis virus (TGEV) is restricted to apical surfaces of polarized epithelial cells, as we have recently shown (J. W. A. Rossen, C. P. J. Bekker, W. F. Voorhout, G. J. A. M. Strous, A. van der Ende, and P. J. M. Rottier, 1994, J. Virol. 68, 7966-7973). In this paper we analyze the interactions of mouse hepatitis coronavirus A59 (MHV-A59) with polarized murine kidney cells (mTAL) grown on permeable supports. After inoculation from the apical or basolateral side, virus entry was found to take place only through the apical membrane. The virus utilized a protein of the carcinoembryonic antigen family as its receptor. In contrast to TGEV, MHV-A59 was released preferentially from the basolateral plasma membrane domain, as evidenced by the accumulation of viral proteins and infectivity in the basolateral culture fluid as well as by electron microscopical observations in the mouse, MHV initially replicates in the nasal epithelium before being disseminated throughout the body; the basolateral release of MHV from epithelial cells into the animal's circulation may be the first step in the establishment of a systemic infection

Coronaviruses in polarized epithelial cells

Coronaviruses have a marked tropism for epithelial cells. In this paper the interactions of the porcine transmissible gastroenteritis virus (TGEV) and mouse hepatitis virus (MHV-A59) with epithelial cells are compared. Porcine (LLC-PK1) and murine (mTAL) epithelial cells were grown on permeable supports. By inoculation from the apical or basolateral side both TGEV and MHV-A59 were found to enter the polarized cells only through the apical membrane. The release of newly synthesized TGEV from LLC-PK1 cells occurred preferentially from the apical plasma membrane domain, as evidenced by the accumulation of viral proteins and infectivity in the apical culture fluid. In contrast, MHV was released preferentially from the basolateral membrane of mTAL cells. The apical release of TGEV and the basolateral release of MHV may explain the in vivo establishment of a local and systemic infection, respectivel