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Biomedical subjects

D Ménard

Publications and source records attributed to D Ménard.

At least 19 recordsLinked to original sources
PubMed1992

Insulin decreases chylomicron production in human fetal small intestine.

The objective of this study was to examine the effect of insulin on lipoprotein synthesis and secretion in human fetal intestine. Jejunal explants were cultured with [14C]oleic acid in Leibovitz medium for 42 h. Although the addition of insulin (30 U) did not alter the incorporation of [14C]oleic acid into triglycerides, phospholipids and cholesteryl esters in the tissue, it significantly decreased (P < 0.05) the level of triglycerides (20%) in the medium. Among the three lipoprotein classes (chylomicron, VLDL, and HDL) isolated by ultracentrifugation, the chylomicron level was found significantly (P < 0.05) diminished in the medium (29%). Neither the lipid chemical composition of CM or that of VLDL, LDL and HDL was affected by the presence of insulin. These results suggest that chylomicron synthesis is modulated by insulin, whereas lipoprotein distribution and lipid composition are not regulated by this hormone.

Chylomicrons↗
PubMed1992

The expression of lactase enzymatic activity and mRNA in human fetal jejunum. Effect of organ culture and of treatment with hydrocortisone.

Very sensitive procedures were developed for the parallel determination of intestinal lactase (LPH) activity and the cognate mRNA. Between 14 and 20 weeks of gestation, lactase activity is low and varies only slightly; at 37 weeks, a relatively high level of activity is observed. The amounts of LPH mRNA correlates with the enzymatic activity (r = 0.64). Culture of fetal jejunal explants for 5 days induces by itself a 2-fold increase in LPH mRNA, without any significant change in lactase enzymatic activity. This increase may reflect the loss of a negative transcriptional regulation operative in vivo, and suggests an additional post-transcriptional regulatory component. The addition of hydrocortisone (50 ng/ml) during culture induces a doubling of lactase activity without variation in LHP mRNA, indicating a post-transcriptional modulation by hydrocortisone. The intestinal lysosomal acid beta-galactosidase activity was shown to be unaffected by hydrocortisone treatment. This observation clearly illustrates that the two intestinal beta-galactosidases are regulated differently. Our results suggests a complex developmental regulation of human intestinal lactase and that the perinatal increase in lactase activity could be modulated at a post-transcriptional level by hydrocortisone.

Gene Expression Regulation, Enzymologic↗
PubMed1992

Apamin, a highly potent fetal L-type Ca2+ current blocker in single heart cells.

Apamin, a bee venom polypeptide, was reported to block the naturally occurring Ca2+ slow action potentials (APs) in cultured cell reaggregates from old chick hearts [Bkaily, G. et al. Am. J. Physiol. 248 (Heart Circ. Physiol. 17): H961-H965, 1985] as well as the tetrodotoxin (TTX)- and Mn(2+)-insensitive slow Na+ current in young embryonic chick heart cells (Bkaily, G. In Vitro Toxicology. Academic, In press; Bkaily et al. J. Mol. Cell. Cardiol. 23: 25-39, 1991). With the use of the whole cell voltage-clamp technique in single ventricular cells from 10-day-old chick embryos and 17- to 20-wk-old human fetuses, two types of Ca2+ currents (ICa), T and L, were found. These two types of slow inward current in both heart preparations were nearly similar in their voltage, kinetics, and pharmacology. Apamin, a slow Ca2+ action potential blocker in old embryonic chick heart, was found to block the L-type ICa (IL) in a dose-dependent manner without affecting the T-type ICa in both heart cell preparations. The blockade of the IL by apamin was completely reversible upon washout with apamin-free solution. Therefore, when compared with nifedipine or to PN 200-110, apamin seems to be a highly potent L-type Ca2+ channel blocker in heart cells.

Animals↗
PubMed1992

Ontogeny of intestinal lipid and lipoprotein synthesis.

Despite increasing evidence that the fetal intestine produces apoproteins, there is limited knowledge about its ability to absorb fat. The present study focused on the intracellular processing of lipid formation and transport by lipoproteins in the developing fetus. Explants of fetal jejunum (14-20 weeks) were maintained in serum-free organ culture for 42 h with [14C]-oleate. Esterified lipids extracted from the tissue showed an abundance of phospholipids (PL; 54-77%), while those harvested from the medium displayed a predominance for triglycerides (TG; 68-73%). Only a minor percentage of radioactivity was recovered in cholesterol ester (CE) from the tissue (1.8-2.5%) or medium (1.3-2.5%). Separation of PL by thin-layer chromatography revealed a prevalence of phosphatidylcholine followed by phosphatidylethanolamine. Over the fetal period studied a trend towards increase was noted in TG, CE and most of the PL classes but not phosphatidylcholine. In parallel, we observed a progressive increase in these lipoprotein fractions produced by the fetal intestine during development, i.e., chylomicrons, very low-density lipoproteins and high-density lipoproteins. Our data stress not only the ability of the fetal intestine to absorb fat, but also the ontogenetic changes of lipid and lipoprotein synthesis.

Fetus↗
PubMed1992

Intestinal lipids and lipoproteins in the human fetus: modulation by epidermal growth factor.

The aim of the present investigation was first, to examine the ability of human fetal intestine (17-20 wk) to incorporate fatty acid into esterified lipids; and second, to study in vitro lipoprotein synthesis and secretion by fetal explants, as well as the effect of epidermal growth factor (EGF) on these processes. Cultured fetal jejunal explants were incubated in Leibovitz medium for 42 h with [14C]oleate. Both triglycerides (TG) and phospholipids (PL) were the major labeled products. Whereas TG were predominant (80%) in the culture medium, PL accounted for more than 50% of total tissue lipids. More than 60% of the radioactivity in PL was associated with phosphatidylcholine. Some labeling (< 5%) was also recovered in the cholesteryl ester fraction. Active exocytosis was demonstrated by the accumulation of newly synthesized esterified lipids in the medium and the presence of lipoproteins in the basolateral membrane region and intercellular spaces. Most of the newly synthesized lipids were found in lipoproteins of d < 0.97 g/ml (51.2%) and d < 1.21 g/ml (39.3%), whereas the rest were recovered in d < 1.006 g/ml (9.8%) and 1.063 g/ml (5.6%). A similar trend characterized the lipoprotein secretion. The synthesis of the d < 0.97 g/ml fraction (30,653 +/- 4,122 dpm/mg protein) was significantly greater than the 1.006 g/ml fraction (5,897 +/- 1,734), P < 0.005. The secretion of d < 0.97 g/ml particles into the medium was also five fold higher than that of the d < 1.006 g/ml fraction (P < 0.01). The addition of EGF to the culture medium (25, 50, and 100 ng/ml) significantly enhanced the d < 0.97 g/ml lipoprotein secretion (25-40%) and decreased the d 1.006 g/ml and 1.063 g/ml fraction output. The lipid composition of these lipoprotein fractions was never altered by the presence of EGF, suggesting that the number of lipoprotein particles, rather than size, was modified by the growth factor. The present findings provide the first evidence that the human fetal intestine has the capacity to elaborate lipoprotein fractions for the transport of newly synthesized lipids. Furthermore, our data suggest that EGF, present in significant quantity in saliva, amniotic fluid, and bile, can modulate the release of TG-rich lipoproteins by fetal intestinal explants.

Epidermal Growth Factor↗
PubMed1991

Radioautographic localization of epidermal growth factor receptors in human fetal gut.

The present investigation was undertaken to study the localization and accessibility of epidermal growth factor binding sites in the human fetal gut (15-19 weeks of gestation) using light-microscopic and quantitative autoradiography. Exposure of colonic explants to 5 nmol/L 125I-labeled epidermal growth factor for 60 minutes at 22 degrees C revealed extensive accumulation at binding sites in undifferentiated cells of the crypts and at the base of the villus, as well as in the inner circular layer of the muscularis externa bordering the submucosa. Some labeling was also present in the mesenchymal and vascular elements of the lamina propria. Labeling was virtually absent on the brush border at all levels of the epithelium. Quantitative analysis revealed a distinct gradient in grain density along the various compartments of the crypt-villus axis. Epithelial cells in the deep portions of the crypts showed the highest grain density (9.2 grains/microns 2) with values gradually decreasing to 6.5 in upper crypt and 3.9 in lower villus cells. The upper third of the villus showed very little labeling (0.4 grains/microns 2). Cellular distribution of silver grains in lower villous cells revealed a polarization of labeling in the basolateral infranuclear region. Experiments performed at 4 degrees C and at various incubation times showed similar results. Using isolated loops of intact colon and jejunum, segments in which labeled epidermal growth factor was only accessible on the serosal side showed extensive labeling and distribution similar to that found in explanted tissue. On the other hand, labeled epidermal growth factor could not access these same receptor sites when infused into the lumen, either at 22 degrees C or 4 degrees C. These results show that in the human fetal gut (a) the greatest concentration of epidermal growth factor binding sites is found in regions of high proliferative activity and (b) binding sites are absent from the brush border, suggesting that, under normal circumstances, systemic but not luminal epidermal growth factor has free access to its specific receptor.

Autoradiography↗
PubMed1990

Presence and binding characteristics of calcitriol receptors in human fetal gut.

In the present study, we show for the first time the presence of calcitriol-specific binding sites in hypertonic extracts of cells isolated from human fetal small intestine and colon from 13-21 weeks of gestation. Woolf plot analysis of the binding characteristics revealed the presence of a single class of high affinity receptors. The presence of specific receptors for calcitriol in fetal intestine and colon opens interesting possibilities as to the role of this hormone in human gut development.

Calcitriol↗
PubMed1990

Cell proliferation in developing human stomach.

Cell proliferation during morphogenesis of human stomach was investigated using radioautography and biochemical determinations of [3H]-thymidine incorporation into DNA. Labeling indices in the epithelium, mesenchyme and muscle layer were established on radioautographs and the heights (mm) of the gastric glands were measured between 10 and 17 weeks of gestation. At 11-12 weeks, the appearance of the first pit/gland was noted, and the labeling index ranged from 9.2 to 10.2%. Labeled cells were present at all levels of the stratified epithelium. Between 14 and 16 weeks, the total epithelial labeling index declined sharply (8.1 to 5.4%) with a concomitant increase of the height of the pit/gland structures (0.055 to 0.080 mm). High proliferative activity was also recorded in the mesenchyme and the muscle layer, the labeling indices decreasing between 10 and 17 weeks. The biochemical quantitation of the [3H]-thymidine uptake into the total gastric DNA clearly supported the continuous decrease of the cell proliferation determined by radioautography. Detailed analysis of the epithelium showed that proliferative cells were more numerous at the base of the gland at the earliest stage (11 weeks) but concentrated in the pit/neck regions by 13-14 weeks. As the pit/gland development proceeded (14 to 17 weeks) labeled cells remained more abundant in the pit/neck regions of the gland (10.9%) and were rarely seen on the surface epithelium (2%). The present investigation provides basic quantitative data regarding cell proliferation in developing human stomach, and indicates that the morphogenesis of the gastric glands is correlated with the high proliferative capacity of the pit/neck cells.

Cell Division↗
PubMed1990

Differential effects of epidermal growth factor and hydrocortisone in human fetal colon.

The influence of epidermal growth factor (EGF) and hydrocortisone on the functional development of human fetal colon was studied in organ cultures. Fetal colon (14 to 17 weeks gestation) was cultured for 5 days at 37 degrees C in serum-free Leibovitz L-15 medium alone or supplemented with 1, 10, and 100 ng of EGF/ml or with 50 ng of hydrocortisone/ml of culture medium. The overall morphology of the colonic explants was not altered by the hormonal addition. In the continuous presence of EGF (1, 10, and 100 ng/ml) for 5 days, a significant decrease of [3H]thymidine incorporation into DNA was observed. At the brush border level, the addition of EGF induced a significant drop in sucrase, maltase, and alkaline phosphatase activities. These enzymic modifications occurred between the third and fifth day of culture, whereas variation in DNA synthesis was already evident within 24 h. The addition of hydrocortisone at a dose affecting the small intestine (50 ng/ml) did not significantly influence colonic DNA synthesis nor the digestive enzymic activities. These observations show for the first time that EGF, but not hydrocortisone, influences the proliferation and differentiation of human fetal colonic mucosa.

Cell Division↗
PubMed1989

Cell proliferation during morphogenesis of the human colon.

Cell proliferation during morphogenesis of human colon from 8 to 18 weeks of gestation was investigated using radioautography. The sites of [3H]-thymidine uptake were localized in the epithelium, the mesenchyme, and the muscularis externa. As long as the epithelium was stratified, the proliferative pool was very large, and the highest labeling indices (up to 28%) were found during this period. With the formation of villi, the proliferative pool decreased and focused exclusively in the developing crypts by 14-15 weeks. Between 8 and 11 weeks, the labeling index was similar both in the epithelium and the mesenchyme. By 13 weeks of gestation, the small intestinal cell proliferation pattern (epithelium greater than mesenchyme greater than muscular layer) was established in the fetal colon. The biochemical quantitation of the [3H]-thymidine uptake into the total colonic DNA supported the decreasing cell proliferation pattern determined by radioautography. The present investigation establishes basic quantitative data regarding cell proliferation during a particular developmental phase of the human colon.

Autoradiography↗
PubMed1988

Presence and characteristics of epidermal growth factor receptors in human fetal small intestine and colon.

In the present study, we demonstrate for the first time the presence of important concentrations of EGF binding sites in isolated epithelial cells of both human fetal small intestine and colon as early as 12 weeks gestation. The pattern of EGF binding in the small intestine between 12 and 17 weeks show that binding was significantly higher (2.5-fold) in younger fetuses than in older fetuses. Moreover, the fetal colon exhibited a much higher binding capacity (1.5-2.5 times) than corresponding intestinal cells for all age groups studied. Analysis of Scatchard representations reveal that the concentration of high- and low-affinity binding sites in colonic epithelial cells are twice the values observed in corresponding intestinal cells. The present data raise interesting possibilities as to the role of this growth factor in human fetal gut development.

Binding Sites↗
PubMed1988

Biologic effects of epidermal growth factor in human fetal jejunum.

The influence of epidermal growth factor (EGF) on the differentiation and proliferation of human fetal jejunum was studied in organ cultures. Fetal intestine (11-14-wk gestation) was cultured for 5 days at 37 degrees C in serum-free Leibovitz L-15 medium alone or supplemented with 25, 50, and 100 ng EGF/ml culture medium. The addition of hormone did not modify the morphology of the intestinal explants. Biochemical studies revealed that lactase activity was significantly increased with the addition of 50 and 100 ng EGF/ml culture medium. On the other hand, the increase in sucrase, trehalase, and glucoamylase activities that normally occurs during the culture was repressed in the presence of increasing concentrations of EGF. Deoxyribonucleic acid synthesis was significantly decreased after 5 days of culture even in the presence of the lowest EGF concentration used. Concomitantly, the labeling index of the epithelial cells dropped drastically in the presence of EGF. The EGF-induced variation in DNA synthesis was already evident within 24 h of culture, whereas enzymic modifications occurred only between the third and fifth day of culture. The simultaneous addition of EGF and hydrocortisone (50 ng/ml) did not reveal any synergistic action between the two hormones on the hydrolytic activities of the brush border. However, EGF did inhibit hydrocortisone-stimulated DNA synthesis. The present work provides for the first time some basic data on the influence of EGF on brush border hydrolytic activities and on epithelial cell proliferation of human fetal jejunum. These observations strongly suggest that EGF plays an important role in the fetal development of the human gastrointestinal tract.

Cell Division↗
PubMed1988

Raman spectroscopy of cytoplasmic muscle fiber proteins. Orientational order.

The polarized Raman spectra of glycerinated and intact single muscle fibers of the giant barnacle were obtained. These spectra show that the conformation-sensitive amide I, amide III, and C-C stretching vibrations give Raman bands that are stronger when the electric field of both the incident and scattered radiation is parallel to the fiber axis (Izz). The detailed analysis of the amide I band by curve fitting shows that approximately 50% of the alpha-helical segments of the contractile proteins are oriented along the fiber axis, which is in good agreement with the conformation and composition of muscle fiber proteins. Difference Raman spectroscopy was also used to highlight the Raman bands attributed to the oriented segments of the alpha-helical proteins. The difference spectrum, which is very similar to the spectrum of tropomyosin, displays amide I and amide III bands at 1,645 and 1,310 cm-1, respectively, the bandwidth of the amide I line being characteristic of a highly alpha-helical biopolymer with a small dispersion of dihedral angles. A small dichroic effect was also observed for the band due to the CH2 bending mode at 1,450 cm-1 and on the 1,340 cm-1 band. In the C-C stretching mode region, two bands were detected at 902 and 938 cm-1 and are both assigned to the alpha-helical conformation.

Amides↗
PubMed1987

The biosynthesis of intestinal sucrase-isomaltase in human embryo is most likely controlled at the level of transcription.

Although sucrase-isomaltase appears in the small intestine at quite different stages of development in man as compared with most mammals, we find that in human embryo also the appearance of sucrase-isomaltase mRNA closely parallels that of sucrase and isomaltase activities, as we have previously found to be the case in baby rabbits. Also, in the proximal-distal gradient of human embryonic intestine (proximal small intestine greater than distal small intestine greater than colon) the levels of these enzyme activities and those of the corresponding mRNA correlate closely. Finally, glucocorticosteroid treatment of a human colon carcinoma cell line (Caco-2) in vitro or of baby rabbits in vivo leads to a parallel increase of both sucrase and isomaltase activities and of sucrase-isomaltase mRNA. We conclude that in man also, in spite of the different timing in development, the biosynthesis of sucrase-isomaltase is most likely to be controlled at the level of transcription or perhaps of the mRNA stability.

Animals↗
PubMed1987

Maturation of human fetal esophagus maintained in organ culture.

The purpose of this work was to study the human fetal esophagus maintained in organ culture. Esophageal explants from 8 fetuses aged from 12 to 16 weeks of gestation were cultured up to 21 days at 37 degrees C in Leibovitz L-15 serum-free medium. Between 12 and 16 weeks of gestation, the esophagus has a stratified columnar ciliated epithelium, and glycogen aggregates are present in all cell layers. This morphology remains the same up to 5 days in culture. After 7 to 9 days, a vacuolization in the upper half layer occurs, leading to a lifting off of the ciliated layer and a flattening of the subjacent cells. After 15 days of culture, the esophageal epithelium is stratified squamous and the cells are exfoliated at the surface of the explants. Glycogen aggregates are still present in all layers. Islets of ciliated cells resting on the basal cell layers develop within the squamous epithelium. With the extension of the culture period up to 21 days, the general morphology of the epithelium does not change. The ultrastructural features of the newly formed squamous epithelium, with its basal lamina, are similar to that reported for human adult esophageal epithelium. During the course of the culture, the DNA synthesis continues as determined by autoradiography. It is concluded that it is possible to maintain viable human fetal esophagus in organ culture and that an accelerated maturation takes place leading to the formation of the adult esophageal epithelium.

Cell Differentiation↗
PubMed1987

Human fetal colon in organ culture.

Human fetal colon (14-16 weeks gestation) was cultured as explants for 15 days in serum-free Leibovitz L-15 medium at 37 degrees C. The overall morphology of the colonic explants was well maintained throughout the culture period and all epithelial cell types retained their ultrastructural characteristics. The incorporation of [3H]-leucine continued and even increased, reflecting sustained synthesis of proteins. Even though the incorporation of [3H]-thymidine into the total DNA decreased during culture, the synthesis of DNA continued. The sites of [3H]-thymidine incorporation into the different layers of the colonic wall were studied by radioautography. The incorporation of the radioactive precursor occurs mainly in the epithelium and to lesser degrees in the mesenchyme and the muscular layer. Labeled epithelial nuclei were located in the intervillous areas but not on the villi. The labeling index of the epithelial cells remained constant throughout the culture period indicating the preservation of the proliferative capacity of the epithelium. Brush-border hydrolytic activities, namely those of sucrase, maltase, lactase, trehalase, glucoamylase and alkaline phosphatase, were assayed in the colonic tissue. These enzymic activities generally decreased in the tissue and increased in the medium during the course of culture. These observations clearly demonstrate that fetal colon can be maintained viable for at least 15 days in a serum-free medium. Organ culture now provides the opportunity to study the normal function and metabolism of human colon during its development.

Autoradiography↗
PubMed1987

Stimulatory effects of epidermal growth factor on deoxyribonucleic acid synthesis in the gastrointestinal tract of the suckling mouse.

The effects of epidermal growth factor (EGF), cortisone and thyroxine on deoxyribonucleic acid (DNA) synthesis in the esophagus, stomach, small intestine and colon have been studied in suckling mouse. Daily administration of EGF [4 micrograms/g body weight (bw)/day] during 3 days to 8-day-old mice induced a significant increase of the incorporation of [3H]thymidine into DNA in the stomach, the small intestine, and the two halves of the colon. The DNA synthesis in the esophagus remained unaffected by the EGF treatment. The maximal increase of [3H]thymidine incorporation into DNA was observed in the colon, and represented 112%. Daily administration of cortisone acetate (25 micrograms/g bw/day) or thyroxine (1 microgram/g bw/day) during 3 days to 8-day-old mice had no significant influence of the DNA synthesis of any part of the gastrointestinal tract. These results show that EGF is able to affect the DNA synthesis in the stomach, small intestine and colon of suckling mice.

Animals↗