Guy Dandrifosse

Spermine and spermidine induce intestinal maturation in the rat

In the present study, we aimed to induce precocious intestinal maturation in neonatal rats by the oral administration of polyamines. Groups of 5 rats received either saline, spermidine (10 mumol daily), or spermine (6 mumol daily) orally on the 12th, 13th, and 14th postnatal days. The rats were killed on the 15th postnatal day. After the small bowel was removed, a 1-cm distal ileal segment was removed for histologic examination and the remaining small bowel tissue was homogenized for further biochemical analysis. Polyamine administration was shown to induce structural and biochemical mucosal changes characteristic of postnatal maturation. Lactase, sucrase, and maltase specific activities (micromoles of substrate hydrolyzed per minute per gram of protein) were 80 +/- 10, 10 +/- 3, and 116 +/- 19 for the saline-treated rats; 51 +/- 7, 34 +/- 2, and 315 +/- 37 for the spermidine-treated rats; and 25 +/- 2, 46 +/- 5, and 419 +/- 63 for the spermine-treated rats, respectively. Similar results were obtained with rats, first treated with spermine (6 mumol) on the 7th postnatal day, receiving spermine (6 mumol) daily as described above and killed on the 10th postnatal day. Dose-response experiments performed as reported above in rats whose treatment began on the 12th postnatal day showed that the maturational effects of orally administered spermine are dose-dependent.

Polyamine Concentration in Rat Milk and Food, Human Milk, and Infant Formulas 1

ABSTRACT: The polyamine concentration in rat milk and food, human milk, and infant formulas was estimated by HPLC. In rat milk, the concentration of putrescine and spermine was low (generally under 2.5 nmol.mL−1 for putrescine and under 1 nmol.mL−1 for spermine). The spermidine concentration was higher and seemed to increase during lactation. The rat food was richer in polyamines than the rat milk (about 150 times for putrescine and spermine, about 30 times for spermidine). We already proved that ingestion of spermine or spermidine can induce precocious maturation of the rat intestine. The present observations suggest that polyamines contained in rat food could play an important role in postnatal maturation of the rat intestine. The polyamine concentration of human milk was measured from 60 different mothers during a period extending from the 1st wk to the 6th mo of lactation. Great variation was observed. During the 1st mo of lactation, the general pattern was as follows: putrescine concentration generally varied little (from 1 to 3 nmol.mLr−1), spermine and spermidine concentrations showed a similar pattern (the highest values appeared at the end of the 1st wk of suckling). After the 4th mo of lactation, putrescine concentration increased slightly, whereas spermine and spermidine concentration stayed almost stable. The concentrations of polyamines in 18 powdered milks for babies were estimated. Spermine and spermidine contents were lower than those in human milk. A protective effect of spermine or spermidine against alimentary allergies is suggested.

Development of a serum-free co-culture of human intestinal epithelium cell-lines (Caco-2/HT29-5M21)

BackgroundThe absorptive and goblet cells are the main cellular types encountered in the intestine epithelium. The cell lineage Caco-2 is a model commonly used to reproduce the features of the bowel epithelium. However, there is a strong debate regarding the value of Caco-2 cell culture to mimick in vivo situation. Indeed, some authors report in Caco-2 a low paracellular permeability and an ease of access of highly diffusible small molecules to the microvilli, due to an almost complete lack of mucus. The HT29-5M21 intestinal cell lineage is a mucin-secreting cellular population. A co-culture system carried out in a serum-free medium and comprising both Caco-2 and HT29-5M21 cells was developed. The systematic use of a co-culture system requires the characterization of the monolayer under a given experimental procedure.ResultsIn this study, we investigated the activity and localization of the alkaline phosphatase and the expression of IAP and MUC5AC genes to determine a correlation between these markers and the cellular composition of a differentiated monolayer obtained from a mixture of Caco-2 and HT29-5M21 cells. We observed that the culture conditions used (serum-free medium) did not change the phenotype of each cell type, and produced a reproducible model. The alkaline phosphatase expression characterizing Caco-2 cells was influenced by the presence of HT29-5M21 cells.ConclusionThe culture formed by 75% Caco-2 and 25% HT29-5M21 produce a monolayer containing the two main cell types of human intestinal epithelium and characterized by a reduced permeability to macromolecules.

Are milk polyamines preventive agents against food allergy

Insufficient polyamine intake could play a role in the induction of sensitization to dietary allergens. This proposal is based essentially on investigations made in sucking rats and in children. In sucking rats it has been established that oral administration of spermine can induce all the modifications occurring in the digestive tract at weaning. In the intestine events occur in two phases. The early event consists of desquamation of the epithelium resulting from an activation of apoptosis. The late event appears to involve an hormonal cascade in which adrenocorticotropic hormone, cytokines, bombesin and corticosterone are included. Observations in human subjects show that: (1) the spermine and spermidine concentrations are generally lower in infant formulas than in human breast milk. Mothers seem consistently to have relatively high or relatively low concentrations of spermine and spermidine in their milk. These individual variations may be due to diet, lifestyle or genetic background; (2) the probability of developing allergy can reach 80 % if the mean spermine concentration in the milk is lower than 2 nmol/ml milk. It is approximately 0 % if the mean spermine concentration is higher than 13 nmol/ml milk; (3) preliminary results show that the intestinal permeability to macromolecules differs in premature babies when they are fed on breast milk compared with infant formulas (J Senterre, J Rigo, G Forget, G Dandrifosse and N Romain, unpublished results). This difference does not seem to be present when powdered milk is supplemented with polyamines at the concentration found in breast milk; (4) spermine increases proliferation and differentiation of lymphocytes isolated from the tonsils of children.

Metabolism of phospholipids of anterior or posterior gills of the crab Eriocheir sinensis M. EDW, during the adaptation of this animal to media of various salinities

Abstract 1. 1. The lipid composition and the fatty-acid pattern from anterior and posterior gills taken from Eriocheir sinensis adapted to fresh water or to sea water are analysed. 2. 2. Total phospholipids of posterior gills increase when crabs are acclimated in dilute media; lipid content of anterior gills remains constant in fresh water and sea water animals. 3. 3. Fatty-acid pattern of anterior and posterior gills does not change following a variation in the salinity of the exterior medium. 4. 4. Velocity of total phospholipid renewal from inorganic phosphate (Pi) and acido-soluble organic phosphate (PoAS) is increased when the crab is living in sea water. 5. 5. The velocity of synthesis of PE, PS and DPG rises at the level of the posterior gills extracted from freshwater animals. 6. 6. is postulated that PE and especially PS play an important part in the control of the active transport of ions located in the posterior gills of Eriocheir sinensis.

Immunochemical, biomolecular and biochemical characterization of bovine epithelial intestinal primocultures

BackgroundCultures of enterocytes and colonocytes represent valuable tools to study growth and differentiation of epithelial cells. In vitro models may be used to evaluate passage or toxicity of drugs, interactions of enteropathogenes bacteria strains with intestinal epithelium and other physiologic or pathologic phenomenon involving the digestive tract.ResultsCultures of bovine colonocytes and jejunocytes were obtained from organoid-enriched preparations, using a combination of enzymatic and mechanical disruption of the intestine epithelium, followed by an isopicnic centrifugation discarding most single cells.Confluent cell monolayers arising from plated organoids exhibited epithelium typical features, such as the pavement-like structure, the presence of apical microvilli and tight junctions. Accordingly, cells expressed several markers of enterocyte brush border (i.e. maltase, alkaline phosphatase and fatty acid binding protein) as well as an epithelial cytoskeleton component (cytokeratin 18). However, enterocyte primocultures were also positive for the vimentin immunostaining (mesenchyme marker). Vimentin expression studies showed that this gene is constitutively expressed in bovine enterocytes. Comparison of the vimentin expression profile with the pattern of brush border enzymes activities, suggested that the decrease of cell differentiation level observed during the enterocyte isolation procedure and early passages of the primoculture could result from a post-transcriptional de-repression of vimentin synthesis. The low differentiation level of bovine enterocytes in vitro could partly be counteracted adding butyrate (1–2 mM) or using a glucose-deprived culture medium.ConclusionThe present study describes several complementary approaches to characterize bovine primary cultures of intestinal cells. Cultured cells kept their morphologic and functional characteristics during several generations.

Dietary polyamines and non-neoplastic growth and disease.

This review presents the data that are now available concerning the effects of dietary polyamines at either postnatal or adult stages in non-neoplastic growth and disease. Polyamines provided by food have a potential role in growth and development of the digestive system in neonatal mammals (and fishes). In humans, this property could be of importance in preventing the appearance of food allergies. Dietary polyamines also seem necessary for the maintenance of normal growth and general properties of adult digestive tract. Their possible therapeutic effects have been investigated in gastric, intestinal, and, more recently, whole-body healing.

Enhancement of Transfection Efficiency Through Rapid and Noncovalent Post-PEGylation of Poly(Dimethylaminoethyl Methacrylate)/DNA Complexes

AbstractPurpose. The aim of this work was to develop a new strategy to intro- duce poly(ethylene glycol) (PEG) into methacrylate-based polymer/DNA complexes in order to produce hemocompatible particles able to transfect cells in the presence of serum. Methods. Atom transfer radical polymerization was used to synthesize a well-defined poly(2-(dimethylamino)ethyl methacrylate) homopolymer (PDMAEMA) and a poly(2-(dimethylamino)ethyl methacrylate-b-poly(ethylene glycol) α-methyl ether, ω-methacrylate) palm-tree-like copolymer (P(DMAEMA-b-MAPEG)). The complexes obtained by self assembly of the pCMVβ plasmid and the polymers were used to transfect Cos-7 cells. Their physical properties—particle size and zeta potential—were characterized respectively by dynamic light scattering and electrophoretic mobility measurements. Ex vivo hemocompatibility was also determined. Results. The PDMAEMA/pCMVβ complexes transfected Cos-7 cells exclusively in the absence of serum. Although the P(DMAEMA-b-MAPEG) copolymer had no transfection activity per se, the addition of the latter to pre-formed PDMAEMA/DNA complexes significantly enhanced the activity and allowed transfection even in the presence of serum. The presence of palm-tree-like copolymers also improved the hemocompatibility properties of the complexes. No effect on platelet counts was observed for P(DMAEMA-b-MAPEG)/pCMVβ complexes, whereas a decrease of platelets was clearly observed when blood cells were incubated with PDMAEMA/pCMVβ complexes. Conclusions. Such a synergistic effect of noncovalent PEGylation of poly(amino methacrylate)/DNA complexes allows a new and versatile approach to tune up transfection efficiency.

Analysis of dansyl derivatives of di- and polyamines in mouse brain, human serum and duodenal biopsy specimens by high-performance liquid chromatography on a standard reversed-phase column

The concentrations of putrescine, spermine and spermidine were measured in human serum, childrens duodenal biopsy specimens and mouse brain homogenates by high-performance liquid chromatography. The chromatographic analysis was performed on dansyl derivatives of the polyamines using a reverse-phase system with an ion-pairing retention mechanism (heptane sulphonate). Capacity factors were determined at different concentrations of acetonitrile. Simple linear gradients were set up for fast (15 min) or routine (25 min) analysis. Three fluorescence detectors were compared for these determinations and their detection limits determined. The minimum detectable amount of polyamines was 25 fmol compared to 500 fmol with standard detectors. While samples prepared from tissues did not require a high sensitivity, a detector of better performance was needed to assay the polyamines in human serum.

Intestinal development in suckling rats: direct or indirect spermine action?

The present investigation addresses the question of whether spermine orally given to unweaned rats directly or indirectly exerts its effects on the intestinal brush border disaccharidases and if the adrenal gland secretions play a role in this phenomenon. The results showed that spermine, surgically placed in the lower part of the distal small intestine, induced sucrase, stimulated maltase-specific activity and decreased lactase-specific activity in both proximal and distal segments of the small intestine. Introduction of spermine into the lumen of the large intestine stimulated the specific activities of disaccharidases in the whole small intestine. Intraperitoneal injection had no effect except a slight reduction of lactase-specific activity in the distal intestine. Adrenalectomy prevented the oral effect of spermine on sucrase- and maltase-specific activity but not on lactase-specific activity. Addition of spermine to intestinal explants in organ cultures fails to reproduce any of these effects. It even reduced maltase-specific activity. These findings suggest that dietary polyamines have either direct and indirect effects on properties of rat immature intestine.