J. M. V. M. Mouwen

Nippostrongylus brasiliensis Histochemical changes in the composition of mucins in goblet cells during infection in rats

Changes in the quality of mucins in jejunal goblet cells were investigated during an infection with Nippostrongylus brasiliensis in rats. At 10 days after infection, when proliferative activity in the crypts is excessive and both crypts and villi are characterized by hyperplasia of goblet cells, the histochemical composition of the population of goblet cells in comparison with controls shows a marked increase in crypt and villous goblet cells containing neutral mucins. At 15 days after infection both crypts and villi display a significant increase in goblet cells containing acid mucin and decrease in goblet cells containing neutral mucin. The acid mucins in crypt and villous goblet cells on day 15 appear to be sulphomucins predominantly, whereas in controls sialomucin-containing goblet cells dominate both in the crypts and on the villi. These experiments establish that the explusion of N. brasiliensis from the intestine of the rat coincides not only with quantitative, but also with remarkable qualitative changes in the histochemical composition of mucins in goblet cells.

Biological and pathobiological aspects of the glycocalyx of the small intestinal epithelium. A review

The literature on the glycocalyx of small intestinal epithelium is reviewed. The structure, general and barrier functions, synthesis, and degradation of the glycocalyx, and pathobiological aspects of the glycocalyx in relation to its barrier function are mentioned. Topics for future research are indicated.

Interaction of legume lectins with the cellular metabolism of differentiated Caco-2 cells

The binding of the legume lectins Phaseolus vulgaris E4 and L4, Glycine max agglutinin, Vicia faba agglutinin, and Pisum sativum agglutinin to intact differentiated Caco-2 cells and to brush border membranes of differentiated Caco-2 cells was investigated, and their impact on the cellular metabolism and the microvilli of these cells was assessed. P. vulgaris isolectin E4 showed the most intense staining after binding of fluorescein isothiocyanate-labeled lectin to intact Caco-2 cells. P. sativum agglutinin showed the weakest staining intensity. The dissociation constant for P. vulgaris isolectin E4 and P. sativum agglutinin binding was 0.11 x 10(-5) and 1.69 x 10(-5) mol/L, respectively. The values of the dissociation constants for P. vulgaris isolectin L4, G. max agglutinin, and V. faba agglutinin were situated in between these extremes. Stimulation of thymidine, glucosamine, and fucose incorporation was observed after exposure to P. vulgaris isolectins and soybean agglutinin. V. faba agglutinin had an inhibitory effect, whereas P. sativum agglutinin showed little or no effect. Compared with control cells and P. vulgaris isolectin L4- and P. sativum agglutinin-incubated cells, the microvilli of P. vulgaris isolectin E4-, soybean agglutinin-, and V. faba agglutinin-incubated cells were shortened significantly. The data provide evidence that a correlation exists, not only between the dissociation constants of the lectins and the fluorescent staining intensity, but also between the dissociation constants of the lectins and the extent of the legume lectin-induced changes in the cellular metabolism.

Quantitative determination of the lectin binding capacity of small intestinal brush-border membrane. An enzyme linked lectin sorbent assay (ELLSA)

A test to determine quantitatively the lectin binding sites in brush-border membranes has been developed. Highly purified bovine small intestinal brush-border membranes were prepared, and subsequently coated directly to the bottom of a microtiter plate. Soybean agglutinin conjugated with peroxidase was coupled to its binding sites in the brush-border membranes and the peroxidase activity was determined in a spectrophotometer. The number of soybean agglutinin binding sites in the brush-border membranes has been established by means of iterized computer fit analysis of the data, indicating values for maximal binding of 7.10(-7) M soybean agglutinin per mg of brush-border membrane protein and a dissociation constant of 1.5.10(-5) M.

Dietary effects of faba-bean (Vicia faba L.) tannins on the morphology and function of the small-intestinal mucosa of weaned pigs

The objective of the present study was to evaluate effects of condensed tannins in faba beans (Vicia faba L.) on morphological and functional variables of the small-intestinal mucosa of piglets. In an experiment with young piglets (8-17 kg body weight), fed on either a control diet or a diet containing 200 g/kg of low- or high-tannin faba bean hulls (with < 0.10 and 3.3% catechin equivalents of condensed tannins respectively), morphological and functional characteristics of the jejunal mucosa were determined. Results of the study showed that the morphological variables of the mucosa of the three groups of piglets were similar. Also, no changes due to dietary tannins were observed in sucrase (EC3.2.1.48)-isomaltase (EC 3.2.1.10) activity in homogenates of mucosa plus submucosa. However, aminopeptidase (EC 3.4.11.2) activity in these homogenates in the proximal part of the small intestine of the animals of the group fed on the high-tannin diet was significantly lower than that in the animals fed on the control diet or the diet with low-tannin hulls (P < 0.05).

Actin cytoskeletal lesions in differentiated human colon carcinoma Caco-2 cells after exposure to soybean agglutinin

We have investigated the effects of soybean agglutinin on the cytoskeletal element actin in differentiated Caco‐2 cells. The actin cytoskeleton of the cells was visualized by fluorescence microscopy using 7‐nitrobenz‐2‐oxa‐1, 3‐diazole phallacidin as a specific marker for F‐actin. Compared with control Caco‐2 cells no changes in the fluorescence pattern were observed after incubation with soybean agglutinin. However, using the deoxyribonuclease‐I inhibition assay a dose‐related response was noted in the increase of intracellular G‐actin after a 2‐hour incubation period with soybean agglutinin. Already after exposure for 15 min to soybean agglutinin a decrease in intracellular F‐actin was demonstrable. This apparent depolymerization could be prevented by incubating the Caco‐2 cells with soybean agglutinin and the appropriate monosaccharide simultaneously. The increase in the amount of G‐actin appeared to be correlated with a shortening of microvilli on the Caco‐2 cells.

Binding of kidney bean (Phaseolus vulgaris) isolectins to differentiated human colon carcinoma Caco-2 cells and their effect on cellular metabolism.

The binding of Phaseolus vulgaris (PHA) isolectins L4 and E4 to the brush border membrane of differentiated Caco-2 cells was studied and the impact on cellular metabolism and microvilli was assessed. Computer analysis of the data based on binding experiments with peroxidase conjugated isolectins gave mean (SD) values for maximal binding of 2540 (151).10(-9) M for PHA-L4 and 2104 (140).10(-9) M for PHA-E4 per mg of brush border membrane protein. The dissociation constants for L4 and E4 binding were 4.3 (1.4).10(-6) M and 1.1 (0.8).10(-6) M, respectively. Incubation of differentiated Caco-2 cells for 30 minutes with ferritin conjugated PHA isolectins showed that, as indicated by the number of ferritin particles, PHA-E4 bound to the microvilli to a greater extent than PHA-L4. Ferritin particles were also localised intracellularly over endocytotic invaginations and vesicles. After incubation for 48 hours with PHA-L4 or PHA-E4, the relative incorporation of precursors for DNA, RNA, and (glyco)protein synthesis into the trichloroacetic acid insoluble fraction of the Caco-2 cells was determined. Both isolectins stimulated the incorporation of thymidine and glucosamine, but neither PHA-L4 nor PHA-E4 were able to influence the incorporation of uridine. With respect to fucose, methionine, and N-acetyl mannosamine, the stimulatory effect remained confined to PHA-E4. Since PHA-L4 and PHA-E4 were tested at the same concentrations, PHA-E4 is more effective than PHA-L4. The changes in the uptake of radioactive precursors were lost after heat inactivation of PHA-E4. Compared with control and PHA-L4 incubated Caco-2 cells, the microvilli of PHA-E4 incubated cells were shortened significantly (p less than 0.01).

Biological and pathological aspects of the mammalian small intestinal permeability to macromolecules.

The literature on the biology and pathology of mammalian small intestinal permeability to macromolecules is reviewed. In mammals, macromolecules may penetrate the epithelial layer of the small intestinal mucosa, especially in the neonatal period. The neonatal uptake and transport of immunoglobulins is important in the acquisition of passive immunity in the newborn. In the mature small intestine the uptake of macromolecules almost ceases, except in M-cells. Excessive uptake and transport of macromolecules has been demonstrated in several experimental and spontaneous gastrointestinal diseases, in which increased concentration of macromolecules in the small intestinal lumen and/or damage of one or more components of the small intestinal mucosal barrier is present. Finally, methods to study macromolecular permeation of the small intestine are discussed.

Biology and pathology of the intestinal M-cell. A review.

The literature of the intestinal M-cell, which is part of the epithelium covering the gut-associated lymphoid tissue (GALT), is reviewed. Attention is paid to the localization, structure, origin and function of this cell, under both normal and pathological conditions.

The interaction between plant lectins and the small intestinal epithelium: a primary cause of intestinal disturbance.

The literature concerning the effects of plant lectins on the small intestinal epithelium is reviewed. It appears that after oral intake, intact plant lectins can reach the small intestinal lumen. Their binding to the mucosal surface evokes an increased synthesis of glycoproteins and a degeneration of the intestinal epithelium. The epithelial alterations may result in hyperregenerative villus atrophy and endogenous nitrogen loss. These changes ultimately can lead to less efficient feed conversion, diminished growth, scouring, wasting and death. The possible significance of plant lectins in digestive disturbances in farm animals is suggested.