ovalbumin and lucifer-yellow

Absorption of dietary fat in the small intestine involves epithelial exposure to potentially harmful molecules such as bile salts and free fatty acids. We used organ culture of porcine jejunal explants incubated with a pre-digested mixture of fat (plant oil), bile and pancreatin to mimick the physiological process of dietary fat absorption, and short exposures to the fat mixture caused fat droplet accumulation within villus enterocytes. Lucifer yellow (LY), a fluorescent membrane-impermeable polar tracer was included to monitor epithelial integrity. Both in controls and during fat absorption LY penetrated the epithelium and accumulated in the basal lamina and the lamina propria. LY was also seen in the paracellular space, whereas villus enterocytes were generally only weakly labeled except for small amounts taken up by apical endocytosis. In the crypts, however, fat absorption induced cell permeabilization with LY accumulating in the cytosol and nucleus. Morphologically, both apical and basolateral membranes appeared intact, indicating that the leakiness was caused by minor lesions in the membrane. Albeit to a lesser extent, bile alone was capable of permeabilizing crypt cells, implying that the surfactant properties of bile salts are involved in the process. In addition to LY, crypt enterocytes also became permeable for albumin, ovalbumin and insulin. In conclusion, during fat absorption the permeability of the gut epithelium is increased mainly in the crypts. A possible explanation is that cell membranes of immature crypt cells, lacking detergent-resistant lipid raft microdomains, are less resistant to the deleterious effects of bile salts and free fatty acids.

Topics: Albumins; Animals; Bile Acids and Salts; Cell Membrane Permeability; Dietary Fats; Enterocytes; Insulins; Intestinal Absorption; Intestinal Mucosa; Isoquinolines; Ovalbumin; Surface-Active Agents; Swine

The effect of interleukin-4 (IL-4), a cytokine associated with allergy and inflammation, on the permeability of the intestinal epithelium was investigated. IL-4 reduced transepithelial electrical resistance (TER) and increased permeation to horseradish peroxidase (HRP) and Lucifer Yellow (LY) of human intestinal T84 cell monolayers. The increased permeation due to IL-4 treatment was also observed at 4 degrees C. The permeability of T84 cell monolayers to beta-lactogulobulin (beta-Lg), ovalbumin (OVA), and fluorescein isothiocyanate (FITC)-dextran of various molecular sizes was also high in the IL-4-treated cell monolayers. Sodium azide (NaN(3)), which inhibits ATP synthesis of the cells, did not inhibit the increases in these substances. Even 150 kDa FITC-dextran significantly permeated the T84 cells when the monolayers were treated with IL-4. These results suggest that fairly large molecules are able to permeate intestinal epithelial monolayers via the energy-independent paracellular pathway when the monolayers are exposed to excessive IL-4.

Topics: Adenosine Triphosphate; Animals; Biological Transport; Cell Line, Tumor; Dextrans; Fluorescein-5-isothiocyanate; Horseradish Peroxidase; Humans; Interleukin-4; Intestinal Mucosa; Isoquinolines; Lactoglobulins; Macromolecular Substances; Ovalbumin; Permeability; Sodium Azide; Temperature

Salmonella species invade and replicate within epithelial cells in membrane-bound vacuoles. In this report we show that upon infection of HeLa epithelial cells, Salmonella typhimurium residues in vacuoles that contain lysosomal membrane glycoproteins (lgps). Four to six hours after invasion, intracellular bacteria induce the formation of stable filamentous structures containing lgps that are connected to the bacteria-containing vacuoles. Formation of these lgp-rich structures requires viable intracellular bacteria and is blocked by inhibitors of vacuolar acidification. These structures are not present in uninfected cells or in cells infected with another invasive bacteria, Yersinia enterocolitica. Tracers added to the extracellular medium are not delivered to the Salmonella-induced filaments, suggesting that these structures are different from previously described tubular lysosomes. Initiation of intracellular bacterial replication correlates with formation of these lgp-containing filaments. Certain avirulent Salmonella mutants that are defective for intracellular replication fail to induce formation of these structures. These observations suggest that Salmonella-induced filaments containing lgps are linked to intracellular bacterial replication.

Topics: Cell Compartmentation; Epithelium; HeLa Cells; Humans; Hydrogen-Ion Concentration; In Vitro Techniques; Intracellular Membranes; Isoquinolines; Lysosomes; Membrane Glycoproteins; Nocodazole; Ovalbumin; Salmonella typhimurium