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1.
Folia Microbiol (Praha) ; 55(4): 401-8, 2010 Jul.
Article in English | MEDLINE | ID: mdl-20680581

ABSTRACT

Oral administration of Lactobacillus spp. as probiotics is gaining importance in the treatment of intestinal inflammations. However, their mechanism of action is unknown. We investigated whether nonspecific binding Lactobacillus casei Shirota (LcS) and mannose-specific Lactobacillus plantarum 299v (Lp) and their spent culture supernatant (SCS) affect Salmonella enteritidis 857 (Se) growth, IL-8 and Hsp70 syntheses. In one set of experiments human enterocyte-like Caco-2 cells were infected with LcS, Lp or Se at 1-500 bacteria per cell for 1 h. In another set, cells were exposed to Se (0-200 per cell, 1 h) after exposure to lactobacilli (LB) (500 per cell, 30 min) or by co-incubation of Se and LB (1 h). The third set of experiments involved exposure of cells for 1 h to SCS or Se (100 per cell) pretreated (1 h) in SCS. The effect of LB SCS on Se growth was evaluated by agar plate diffusion test. IL-8 and Hsp70 were assessed over 2-24 h using ELISA and Western blotting, respectively. Neither LcS nor Lp affected the Se growth and IL-8 production. In addition, they did not induce Hsp70 expression by Caco-2 cells. Instead, their SCS inhibited the Se growth and IL-8 production and induced the expression of Hsp70 by both crypt- and villus-like cells. The beneficial effect of Lactobacillus spp. to the intestinal inflammations might be associated with a decrease in IL-8 levels. This effect could be mediated, at least in part, via a secreted antimicrobial product(s) either directly against the pathogens or indirectly through the synthesis of Hsp70.


Subject(s)
Immunologic Factors/biosynthesis , Interleukin-8/antagonists & inhibitors , Lacticaseibacillus casei/metabolism , Lactobacillus plantarum/metabolism , Probiotics , Salmonella enteritidis/pathogenicity , Blotting, Western , Caco-2 Cells , Coculture Techniques , Culture Media/chemistry , Enzyme-Linked Immunosorbent Assay , HSP70 Heat-Shock Proteins/biosynthesis , Humans , Interleukin-8/immunology , Salmonella enteritidis/growth & development
2.
Animal ; 4(1): 40-51, 2010 Jan.
Article in English | MEDLINE | ID: mdl-22443616

ABSTRACT

To unravel the underlying mechanisms that explain the positive effects of prefermented cereals on in vivo gastrointestinal (GI) architecture and function, an in vitro experiment using a human small intestinal epithelial cell model (Caco-2) was performed. A range of dilutions (0% to 10%) of the supernatants of three liquid experimental diets, as well as Na-lactate were used in an in vitro experiment to assess their effect on cellular growth, metabolism, differentiation and mucosal integrity using Caco-2. The experimental diets contained, in addition to a protein rich basal diet (60%), (1) a liquid control diet (C) containing 40% of a mixture of barley and wheat (ratio 3 : 1) or (2) a liquid diet (F) containing 40% prefermented barley and wheat or (3) C with the addition of the fermentation end-products (organic acids and ethanol) in concentrations similar to those in the fermented diet (FP). For F, the mixture of barley and wheat was fermented at 35°C for 48 h. Parallel to the in vitro experiment, 18 groups of eight weanling pigs were assigned to one of the experimental diets during a 14-day in vivo experiment. Each group was fed restrictively. The results of the in vitro experiment showed that the lowest dose of both F- and FP-supernatants had no clear effects on the cell proliferation, but incubation with 5% and 10% of the F- and FP-supernatants decreased the cell numbers at day 19. DNA, RNA, protein and glycoprotein synthesis in differentiated Caco-2 cells were stimulated by incubation with the lower concentrations (0.5% to 2.5%) of F- and FP-supernatants whereas the higher concentrations (5% and 10%) had no effect. Both the F- and FP-supernatants decreased the specific sucrase-isomaltase activity in a dose-dependent manner, but the effects on the specific aminopeptidase activities were less clear. Mucosal integrity initially decreased after incubation with the highest F- and FP-supernatants and started to recover between 24 and 48 h. The results of the in vivo experiment showed no dietary effects (P > 0.1) on GI morphology and brush-border enzyme activities at day 5 or at day 14. Time related changes in GI characteristics followed a normal pattern. In conclusion, the supernatants of diets containing either prefermented cereals or their fermentation end-products clearly modulate cellular growth, metabolism, differentiation and mucosal integrity in an in vitro model, although these effects were not observed in the in vivo characteristics measured in weanling pigs.

3.
Benef Microbes ; 1(2): 121-30, 2010 Jun.
Article in English | MEDLINE | ID: mdl-21840800

ABSTRACT

Invasion of the gut by pathogenic Salmonella leads to production of IL-8 that initiates inflammatory reactions to combat the bacterium. However, its persistent production causes tissue damage and interventions that suppress IL-8 production prevent tissue damage. We hypothesised that probiotics could mediate their benefits via inhibition of IL-8 synthesis. Caco-2 cells were infected with probiotic Bifidobacterium infantis W52, Lactobacillus casei W56, Lactococcus lactis W58, Lactobacillus acidophilus W70, Bifidobacterium bifidum W23, or Lactobacillus salivarius W24 or pathogenic Salmonella enterica serovar Enteritidis 857 at 0, 0.2, 1, 2, 10, 20, 100 or 200 bacterial cells/Caco-2 cell for 1 hour. Cells were also exposed to a combination of one probiotic bacterium (200 bacterial cells/Caco-2 cell) and the graded numbers of Salmonella as either co-incubation (1 hour) or pre-incubation of the probiotic bacterium (1 hour) followed by Salmonella (1 hour). The cells recovered for 2 or 24 hours. IL-8 and Hsp70 were determined by ELISA and Western blot respectively. Both probiotics and Salmonella induced a dose- and time-dependent synthesis of IL-8 but probiotics induced far lower IL-8 levels than Salmonella. The Salmonella-induced IL-8 was significantly suppressed by B. infantis W52, L. casei W56 and L. lactis W58 at low numbers of Salmonella (0.2 to 20 bacterial cells/Caco-2 cell) and within 2 hours of recovery. The observed probiotic-mediated reduction in IL-8 secretion was transient, and lost after a few hours. In addition, these three probiotics induced a significant increase in Hsp70 expression while L. acidophilus W70, B. bifidum W23 and L. salivarius W24 induced a weak Hsp70 expression and could not suppress the Salmonella-induced IL-8 synthesis. We conclude that suppression of Salmonella-induced IL-8 synthesis by Caco-2 cells is exhibited by probiotics that induce expression of Hsp70, suggesting that the protective role of probiotics could be mediated, at least in part, via Hsp70 expression. This suppression is limited to a low number of infecting pathogenic Salmonella.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Bifidobacterium/immunology , Interleukin-8/metabolism , Lactobacillus/immunology , Probiotics/pharmacology , Salmonella enteritidis/immunology , Bifidobacterium/physiology , Caco-2 Cells , Enzyme-Linked Immunosorbent Assay , HSP72 Heat-Shock Proteins/metabolism , Humans , Lactobacillus/physiology , Salmonella enteritidis/pathogenicity
4.
Folia Microbiol (Praha) ; 53(3): 189-94, 2008.
Article in English | MEDLINE | ID: mdl-18661289

ABSTRACT

The intestinal environment accommodates a wide range of contents ranging from harmless beneficial dietary and microbial flora to harmful pathogenic bacteria. This has resulted in the development of highly adapted epithelial cells lining the intestine. This adaptation involves the potential of crypt cells to proliferate and to constantly replace villous cells that are lost due to maturity or death. As a result, the normal intestinal epithelial integrity and functions are maintained. This phenomenon is eminent in intestinal defense whereby the intestinal epithelial cells serve as a physical barrier against luminal agents. The protection against agents in the gut lumen can only be effective if the epithelium is intact. Restitution of the damaged epithelium is therefore crucial in this type of defense.


Subject(s)
Antibiosis , Bacteria , Bacterial Infections/prevention & control , Gastrointestinal Diseases/prevention & control , Probiotics , Bacteria/growth & development , Bacteria/pathogenicity , Bacterial Infections/microbiology , Caco-2 Cells , Epithelial Cells/immunology , Epithelial Cells/metabolism , Epithelial Cells/microbiology , Gastrointestinal Diseases/microbiology , Heat-Shock Proteins/metabolism , Humans , Intestines/cytology , Intestines/immunology , Intestines/microbiology
5.
Folia Microbiol (Praha) ; 51(4): 268-72, 2006.
Article in English | MEDLINE | ID: mdl-17007422

ABSTRACT

Caco-2 cells (exhibiting characteristics of mature villus enterocytes) were used to determine bacteria (Salmonella enteritidis causing human gastroenteritis)-intestinal cell interactions. The interference of bacteria with the transepithelial electrical resistance (TEER) of filter-grown Caco-2 cells and the production of IL-8 after exposure of the cells to S. enteritidis 857 and/or Lactobacillus strains (L. gasseri LF221 and L. rhamnosus BGT10) was evaluated. The strain 857 decreased TEER of filter-grown Caco-2 cells; in contrast, lactobacilli had a little or no effect. The effect of S. enteritidis on the TEER decreased if Caco-2 cells were pre-incubated with lactobacilli. This strain induced high levels of IL-8 (which can lead to cell damage). Compared to the IL-8 synthesis after exposure of Caco-2 cells to S. enteritidis 857, simultaneous exposure of Caco-2 cells to S. enteritidis and lactobacilli inhibited the IL-8 synthesis after short recovery periods.


Subject(s)
Interleukin-8/metabolism , Intestinal Mucosa/microbiology , Lactobacillus/physiology , Salmonella Infections/immunology , Salmonella enteritidis/pathogenicity , Caco-2 Cells/metabolism , Caco-2 Cells/microbiology , Electric Impedance , Humans , Immunity, Mucosal , Intestinal Mucosa/metabolism
6.
Int J Food Microbiol ; 103(2): 143-55, 2005 Aug 25.
Article in English | MEDLINE | ID: mdl-16083817

ABSTRACT

We tested the effect of Lactobacillus casei strain Shirota (LcS) on the murine model of ulcerative colitis induced by dextran sodium sulphate. The effect of LcS was tested either as a prophylactic 10 days before the onset of the disease, simultaneously with ulcerative colitis induction or continued 10 days after the disease was induced. LcS was not able to prevent the disease induction in any of the experiments. However, important clinical parameters including blood anemia indicators, body weight, and organ weight were improved in the animals receiving LcS as compared with the ulcerative colitis-induced controls. Increased colonic epithelial regeneration in the LcS treated animals was observed in the chronic stage. The results seemed better for the simultaneous short LcS treatment where some parameters remained similar to the PBS controls, including disease activity scores measured in the acute stage. We can conclude that although LcS alone cannot prevent the induction of ulcerative colitis by dextran sodium sulphate, it can improve the clinical condition of the mice. This could imply important biological consequences for the human situation. Further studies including LcS or other probiotic bacteria together with the available treatment are encouraged.


Subject(s)
Colitis, Ulcerative/immunology , Lacticaseibacillus casei/physiology , Probiotics , Animals , Body Weight , Colitis, Ulcerative/blood , Colitis, Ulcerative/chemically induced , Colitis, Ulcerative/pathology , Dextran Sulfate/toxicity , Disease Models, Animal , Male , Mice , Mice, Inbred BALB C , Organ Size
7.
Clin Exp Immunol ; 141(1): 62-71, 2005 Jul.
Article in English | MEDLINE | ID: mdl-15958071

ABSTRACT

Intestinal epithelial cells secrete the chemokine interleukin (IL)-8 in the course of inflammation. Because heat shock proteins (Hsps) and butyrate confer protection to enterocytes, we investigated whether they modulate Salmonella enterica serovar Enteritidis (S. serovar Enteritidis)-induced secretion of IL-8 in enterocyte-like Caco-2 cells. Caco-2 cells incubated with or without butyrate (0-20 m M, 48 h) were infected with S. serovar Enteritidis after (1 h at 42 degrees C, 6 h at 37 degrees C) or without prior heat shock (37 degrees C). Levels of Hsp70 production and IL-8 secretion were analysed using immunostaining of Western blots and enzyme-linked immunosorbent assay (ELISA), respectively. The cells secreted IL-8 in response to S. serovar Enteritidis and produced Hsp70 after heat shock or incubation with butyrate. The IL-8 secretion was inhibited by heat shock and butyrate concentrations as low as 0.2 m M for crypt-like and 1 m M for villous-like cells. In a dose-dependent manner, higher butyrate concentrations enhanced IL-8 secretion to maximal levels followed by a gradual but stable decline. This decline was associated with increasing production of Hsp70 and was more vivid in crypt-like cells. In addition, the higher concentrations abolished the heat shock inhibitory effect. Instead, they promoted the IL-8 production in heat-shocked cells even in the absence of S. serovar Enteritidis. We conclude that heat shock and low concentrations of butyrate inhibit IL-8 production by Caco-2 cells exposed to S. serovar Enteritidis. Higher butyrate concentrations stimulate the chemokine production and override the inhibitory effect of the heat shock. The IL-8 down-regulation could in part be mediated via production of Hsp70.


Subject(s)
Butyrates/pharmacology , Enterocytes/immunology , HSP70 Heat-Shock Proteins/biosynthesis , Interleukin-8/metabolism , Salmonella Infections/immunology , Salmonella enteritidis , Caco-2 Cells , Dose-Response Relationship, Immunologic , Down-Regulation/drug effects , Enterocytes/drug effects , HSP70 Heat-Shock Proteins/immunology , Heat-Shock Response/immunology , Histone Deacetylase Inhibitors , Humans , Hydroxamic Acids , Immunity, Mucosal , Intestinal Mucosa/immunology , Salmonella Infections/metabolism
8.
Food Addit Contam ; 20(5): 427-37, 2003 May.
Article in English | MEDLINE | ID: mdl-12775461

ABSTRACT

The response of intestinal epithelial cells to short-chain fatty acids, which are increasingly used as food additives, was investigated. Human small intestinal epithelial cell model Caco-2 cells were exposed to formate, propionate and butyrate to assess their effect on cellular growth, metabolism, differentiation and protection against bacteria. The Caco-2 cells were entirely grown in the different short-chain fatty acids and respective growth patterns were determined. Differentiated cells were exposed to 0-20 mM short-chain fatty acids for 48 h and changes in DNA, RNA, (glyco)protein syntheses, sucrase isomaltase activity, transepithelial electrical resistance and protection against Salmonella enteritidis were measured. The short-chain fatty acids, altered linearly and differentially the growth pattern ranging from stimulation by formate to inhibition by butyrate. Formate inhibited cellular metabolism. Low concentrations of up to 5 mM propionate and 2 mM butyrate stimulated metabolism, while higher doses were inhibitory. Formate had no effect on sucrase isomaltase enzyme activity and transepithelial electrical resistance, whereas propionate and butyrate increased these markers of differentiation. Infection with S. enteritidis did not benefit from the short-chain fatty acid-induced transepithelial electrical resistance. It is concluded that formate, propionate and butyrate selectively and differentially modulate growth characteristics, cellular metabolism, sucrase isomaltase activity and transepithelial electrical resistance in a concentration- and carbon atom-related fashion. The short-chain fatty acid-induced transepithelial electrical resistance does not confer protection against S. enteritidis.


Subject(s)
Enterocytes/drug effects , Fatty Acids, Volatile/pharmacology , Food Additives/pharmacology , Caco-2 Cells , Cell Death/drug effects , Cell Differentiation/drug effects , Cell Division/drug effects , Dose-Response Relationship, Drug , Humans , Intestinal Mucosa/drug effects , Microvilli/enzymology , Salmonella Infections/prevention & control , Salmonella enteritidis , Sucrase-Isomaltase Complex/metabolism
9.
Cancer Lett ; 161(1): 113-20, 2000 Dec 08.
Article in English | MEDLINE | ID: mdl-11078920

ABSTRACT

The constitutional expression of heat shock proteins (HSP) 27, 70 and 90 in human breast, colon and ovarian cancer cells transplanted into severe combined immunodeficient (SCID) mice was evaluated. In addition their induced expression under chemotherapeutic stress was analyzed. The oestrogen receptor positive breast cancer cell lines (MCF-7, T47D) demonstrated an increased level of HSP 27 and 70 expression compared with oestrogen receptor negative cell lines (BT20, HBL100). After 5-fluorouracil application for 4 days, HSP 27 and 70 expression was increased in HT29 colon tumours. Hence, the human/SCID mouse model is well suited to evaluate the constitutional and induced expression of human HSPs under various experimental conditions.


Subject(s)
Heat-Shock Proteins/biosynthesis , Neoplasms, Experimental/metabolism , Adenocarcinoma/drug therapy , Adenocarcinoma/immunology , Adenocarcinoma/metabolism , Animals , Antimetabolites, Antineoplastic/pharmacology , Breast Neoplasms/immunology , Breast Neoplasms/metabolism , Colonic Neoplasms/drug therapy , Colonic Neoplasms/immunology , Colonic Neoplasms/metabolism , Female , Fluorouracil/pharmacology , HSP70 Heat-Shock Proteins/biosynthesis , HSP90 Heat-Shock Proteins/biosynthesis , Humans , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, SCID , Neoplasm Transplantation , Neoplasms, Experimental/drug therapy , Neoplasms, Experimental/immunology , Ovarian Neoplasms/immunology , Ovarian Neoplasms/metabolism , Transplantation, Heterologous , Tumor Cells, Cultured , Xenograft Model Antitumor Assays
10.
FEMS Microbiol Lett ; 185(2): 175-9, 2000 Apr 15.
Article in English | MEDLINE | ID: mdl-10754244

ABSTRACT

A nonflagellated mutant of Salmonella enterica serotype Enteritidis was constructed by disrupting the flagellin gene (fliC). Northern blot analysis indicated that the mutation did not affect expression of the downstream fliU gene. Infection experiments with differentiated Caco-2 cells revealed that the mutant was about 50-fold less invasive than the wild-type strain, while bacterial adherence was unaffected. Complementation of the mutant with an intact fliC copy restored flagella formation and efficient bacterial invasion. Our data demonstrate that the fliC gene of S. enterica serotype Enteritidis is essential for the invasion of Caco-2 cells.


Subject(s)
Flagellin/genetics , Salmonella enteritidis/genetics , Salmonella enteritidis/pathogenicity , Bacterial Adhesion , Caco-2 Cells , Conjugation, Genetic , Culture Media , Humans , Microscopy, Electron , Mutation , Polymerase Chain Reaction , Salmonella Infections/microbiology , Salmonella enteritidis/physiology , Virulence/genetics
11.
Gut ; 46(5): 679-87, 2000 May.
Article in English | MEDLINE | ID: mdl-10764712

ABSTRACT

BACKGROUND: The enterocytes of the intestinal epithelium are regularly exposed to potentially harmful substances of dietary origin, such as lectins. Expression of heat shock proteins (HSPs) by this epithelium may be part of a protective mechanism developed by intestinal epithelial cells to deal with noxious components in the intestinal lumen. AIM: To investigate if the lectins PHA, a lectin from kidney beans (Phaseolus vulgaris) and WGA, a lectin from wheat germ (Triticum aestivum) could modify the heat shock response in gut epithelial cells and to establish the extent of this effect. METHODS: Jejunal tissue sections from PHA and WGA fed rats were screened for expression of HSP70, HSP72, and HSP90 using monoclonal antibodies. Differentiated Caco-2 cells, the in vitro counterpart of villus enterocytes, were exposed to 100 microg/ml of PHA-E(4) or WGA for 48 hours and investigated for changes in DNA and protein synthesis by double labelling with [2-(14)C]thymidine and L-[methyl-(3)H]methionine. The relative concentrations of HSP60, HSP70, HSP72, and HSP90 and binding protein (BiP) in these cells exposed to lectins were analysed by polyacrylamide gel electrophoresis and immunoblotting. To establish if lectin exposed differentiated Caco-2 cells were still capable of producing a heat shock response, these cells received a heat shock (40 degrees C, 41 degrees C, and 42 degrees C) for one hour and were allowed to recover for six hours at 37 degrees C. During heat shock and recovery periods, lectin exposure was continued. RESULTS: Constitutive levels of HSPs were measured in the intestinal cells of lactalbumin fed (control) rats, as may be expected from the function of this tissue. However, in PHA and WGA fed rats a marked decline in the binding of antibodies against several HSPs to the intestinal epithelium was found. These results were confirmed by in vitro experiments using differentiated Caco-2 cells exposed to PHA-E(4) and WGA. However, after exposure to lectins, these cells were still capable of heat induced heat shock protein synthesis, and total protein synthesis was not impaired indicating specific inhibition of HSP synthesis in non-stressed cells. CONCLUSIONS: We conclude that PHA and WGA decrease levels of stress proteins in rat gut and enterocyte-like Caco-2 cells, leaving these cells less well protected against the potentially harmful content of the gut lumen.


Subject(s)
Heat-Shock Proteins/drug effects , Heat-Shock Response/physiology , Intestinal Mucosa/drug effects , Phytohemagglutinins/pharmacology , Wheat Germ Agglutinins/pharmacology , Animals , Antibodies, Monoclonal/immunology , Blotting, Western , Caco-2 Cells/drug effects , Caco-2 Cells/metabolism , Electrophoresis, Polyacrylamide Gel , Heat-Shock Proteins/biosynthesis , Heat-Shock Proteins/immunology , Humans , Intestinal Mucosa/metabolism , Male , Rats , Rats, Inbred Strains , Specific Pathogen-Free Organisms
12.
Biochem J ; 329 ( Pt 3): 551-60, 1998 Feb 01.
Article in English | MEDLINE | ID: mdl-9445382

ABSTRACT

Arcelin-1 is a lectin-like protein found in the seeds of wild varieties of the kidney bean (Phaseolus vulgaris). This protein displays insecticidal properties, but the mechanism of action is as yet unknown. In the present study we investigated the biochemical and biophysical properties of arcelin-1 from Phaseolus vulgaris cv. RAZ-2. Native arcelin-1 is a dimeric glycoprotein of 60 kDa, built from the non-covalent association of two identical monomers. This dimer resists dissociation by chaotropic agents and is highly resistant to proteolytic enzymes. Each subunit contains 10% (w/w) neutral sugars which belong to the high-mannose and complex-type glycans attached to three glycosylation sites. No interaction of the protein with simple sugars could be detected, but arcelin-1 displays an intrinsic specificity in binding complex glycans. Arcelin-1 therefore differs from the closely related phytohaemagglutinin lectins and alpha-amylase inhibitor in several respects: oligomerization states, sugar-binding affinities and the type and number of glycan chains. These features may be related to the toxicity of arcelin-1.


Subject(s)
Fabaceae/chemistry , Glycoproteins/chemistry , Insecticides/chemistry , Lectins/chemistry , Plant Proteins/chemistry , Plants, Medicinal , Polysaccharides/metabolism , Amino Acid Sequence , Animals , Caco-2 Cells , Chromatography, Ion Exchange , Chymotrypsin/metabolism , Glycoproteins/isolation & purification , Glycoproteins/metabolism , Hemagglutination Tests , Humans , Hydrolysis , Insecticides/metabolism , Lectins/metabolism , Molecular Sequence Data , Pepsin A/metabolism , Plant Lectins , Plant Proteins/isolation & purification , Plant Proteins/metabolism , Protein Binding , Rabbits , Seeds/chemistry , Trypsin/metabolism
13.
Gut ; 38(1): 47-52, 1996 Jan.
Article in English | MEDLINE | ID: mdl-8566858

ABSTRACT

The effect of Phaseolus vulgaris isolectin E4 on polyamine concentrations and ornithine decarboxylase activity of proliferating and differentiating Caco-2 cells was investigated. Values of putrescine, spermidine, and spermine in control cells were highest during the early phase of proliferative cell growth and lowest in the stationary phase. Phytohaemagglutinin E4 significantly increased cellular polyamine values during the late proliferative phase of cell growth. Ornithine decarboxylase activity was high during intensive proliferation and growth, but was lower when proliferation slowed down or ceased. Exposure of Caco-2 cells in the early proliferative phase of cell growth to increasing concentrations of the potent intestinal growth factor phytohaemagglutinin E4 greatly stimulated enzyme activity. In contrast, the activity of ornithine decarboxylase was not stimulated in Caco-2 cells of the late proliferative phase nor was there any increase in the enzyme activity in differentiating and fully differentiated cells of the stationary phase. Accordingly, when proliferating Caco-2 cells possessed the highest ornithine decarboxylase activity, the polyamine values were also at their highest. During differentiation, as the ornithine decarboxylase activity fell close to zero, polyamine values also decreased. In the early proliferative phase of cell growth ornithine decarboxylase activity coincided with DNA synthesis in cells exposed to Phaseolus vulgaris isolectin E4. These findings with Caco-2 cells were similar to those found in brush border cells of the rat small intestine.


Subject(s)
Biogenic Polyamines/metabolism , Caco-2 Cells/drug effects , Ornithine Decarboxylase/drug effects , Phytohemagglutinins/pharmacology , Caco-2 Cells/cytology , Caco-2 Cells/metabolism , Cell Division , Humans , Ornithine Decarboxylase/metabolism
14.
Br J Nutr ; 73(1): 31-9, 1995 Jan.
Article in English | MEDLINE | ID: mdl-7857913

ABSTRACT

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).


Subject(s)
Aminopeptidases/metabolism , Fabaceae/chemistry , Intestinal Mucosa/enzymology , Jejunum/enzymology , Plants, Medicinal , Swine/metabolism , Tannins/administration & dosage , Animals , Intestinal Mucosa/anatomy & histology , Intestinal Mucosa/drug effects , Jejunum/anatomy & histology , Jejunum/drug effects , Oligo-1,6-Glucosidase/metabolism , Sucrase/metabolism , Tannins/pharmacology
15.
Microb Pathog ; 15(6): 207-19, 1993 Dec.
Article in English | MEDLINE | ID: mdl-7911967

ABSTRACT

Enterotoxigenic Escherichia coli strains expressing F17 fimbriae bind to the intestinal mucosa of young calves. F17 fimbriae recognize receptors present in the mucus layer and the brush-border membranes from duodenum, jejunum and ileum. The adhesion of E. coli F17 can be inhibited by several glycoproteins. Adhesion is also inhibited by pretreatment of mucus and brush-border membranes with sodium metaperiodate. The use of glycoconjugates as potential adhesion-blockers is further discussed.


Subject(s)
Antigens, Bacterial/biosynthesis , Bacterial Adhesion , Bacterial Outer Membrane Proteins/biosynthesis , Escherichia coli/physiology , Intestinal Mucosa/microbiology , Intestine, Small/microbiology , Adhesins, Escherichia coli , Animals , Antigens, Bacterial/metabolism , Bacterial Outer Membrane Proteins/metabolism , Cattle , Escherichia coli/growth & development , Escherichia coli/metabolism , Escherichia coli Infections/microbiology , Intestine, Small/ultrastructure , Microscopy, Electron , Microvilli/microbiology
16.
Gastroenterology ; 102(5): 1516-23, 1992 May.
Article in English | MEDLINE | ID: mdl-1568561

ABSTRACT

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.


Subject(s)
Colonic Neoplasms/metabolism , Lectins/metabolism , Plant Lectins , Soybean Proteins , Colonic Neoplasms/pathology , Fucose/metabolism , Glucosamine/metabolism , Humans , Microvilli/metabolism , Phytohemagglutinins/metabolism , Thymidine/metabolism , Tumor Cells, Cultured , Uridine/metabolism
17.
Gut ; 32(8): 886-92, 1991 Aug.
Article in English | MEDLINE | ID: mdl-1885071

ABSTRACT

The interaction of plant lectins with pig small intestinal epithelium in organ culture was studied. The binding of Phaseolus vulgaris (PHA) isolectins E4 and L4 to the microvilli and microvillus vesicles in the top area of the villi was shown by immunoelectron microscopy. Differences were observed in the distribution of the isolectins. In the explants cultured for five hours with the PHA isolectins, the enterocyte height and the villus length were decreased, and a lower villus: crypt ratio was calculated. Ultrastructurally, the microvilli were shorter and irregularly positioned. After incubation with both PHA E4 and PHA L4, clusters of small vesicles, tied off from the microvilli, were seen in higher numbers when compared with control explants. The activity of the brush border enzyme sucrase-isomaltase was reduced in the PHA E4 incubated explants but did not change in the PHA L4 incubated explants. This investigation shows that explants of pig jejunal mucosa in organ culture are suitable for study of the pathological effects of lectins on the small intestinal mucosa. This method may also be used in elucidating the mechanisms by which damage to mucosal structure occurs.


Subject(s)
Fabaceae , Intestinal Mucosa/drug effects , Lectins/toxicity , Plants, Medicinal , Animals , Intestinal Mucosa/enzymology , Intestinal Mucosa/ultrastructure , Jejunum/ultrastructure , Male , Microscopy, Electron , Microvilli/enzymology , Organ Culture Techniques , Plant Lectins , Sucrase-Isomaltase Complex/metabolism , Swine
18.
Gut ; 32(2): 196-201, 1991 Feb.
Article in English | MEDLINE | ID: mdl-1864541

ABSTRACT

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).


Subject(s)
Colonic Neoplasms/metabolism , Phytohemagglutinins/pharmacokinetics , Cell Membrane/ultrastructure , Colonic Neoplasms/pathology , Colonic Neoplasms/ultrastructure , Cytoplasm/metabolism , DNA, Neoplasm/metabolism , Ferritins , Glycoproteins/biosynthesis , Humans , Microvilli/metabolism , Nucleic Acid Precursors/metabolism , RNA Precursors/metabolism , RNA, Neoplasm/metabolism , Tumor Cells, Cultured
19.
Tijdschr Diergeneeskd ; 115(22): 1051-7, 1990 Nov 15.
Article in Dutch | MEDLINE | ID: mdl-2256096

ABSTRACT

Rats were infected with Nippostrongylus brasiliensis, and changes in the histochemical composition of mucins in jejunal goblet cells were investigated. Ten days after infection, mitotic activity was extensively in jejunal crypts: both crypts and villi were characterised by hyperplasia of goblet cells. Infected rats had a markedly greater number of crypt and villi goblet cells containing neutral mucin than the control rats did. Moreover, 15 days after infection, infected rats had significantly more goblet cells containing acid mucin than control rats did. In infected rats, the acid mucins in goblet cells (day 15) appeared to be predominantly sulphomucins, whereas in control rats the acid mucins were predominantly sialomucins. The experiments established that when N. brasiliensis is excreted by rats, the histochemical composition of mucins in crypt and villi goblet cells has been changed not only quantitatively, but also qualitatively.


Subject(s)
Jejunum/metabolism , Mucins/metabolism , Nematode Infections/metabolism , Nippostrongylus , Animals , Jejunum/cytology , Male , Mucins/chemistry , Rats , Rats, Inbred Strains , Sialomucins
20.
Vet Q ; 11(2): 108-15, 1989 Apr.
Article in English | MEDLINE | ID: mdl-2662569

ABSTRACT

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.


Subject(s)
Intestinal Diseases/veterinary , Intestine, Small/drug effects , Lectins/pharmacology , Animals , Epithelium/drug effects , Intestinal Diseases/etiology
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