Daniel Ménard

Insulin modulation of newly synthesized apolipoproteins B-100 and B-48 in human fetal intestine: Gene expression and mRNA editing are not involved

We investigated insulins effect on intestinal lipid transport and, particularly, the biogenesis of apolipoproteins crucial to lipoprotein secretion. Adding insulin (3 mU) to the serum‐free medium of cultured jejunal explants from human fetuses (17–20 weeks) reduced triglyceride and chylomicron production and inhibited apo B‐48 and apo B‐100 secretion. When apo B mRNA was assayed by RT‐PCR and its editing by primer extension, no change was detectable following the addition of insulin. HDL lipid content, apo A‐1 synthesis and RNA level were unaffected by insulin. Collectively, these results suggest that the insulin‐stimulated decline in intestinal chylomicron output may involve apo B co‐ or post‐translational modifications.

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.

Epidermal growth factor (EGF) accelerates the maturation of fetal mouse intestinal mucosa in utero.

Pregnant Swiss ICR mice were injected i.p. with 0.5 μg of epidermal growth factor (EGF) per g b. wt at 15, 16 and 17 days of gestation and fetuses were removed at 18 days of gestation. EGF treatment had no effect on the weight of the fetuses and on the length of the small intestine. No modification of the protein and DNA contents was noted. However brush border alkaline phosphatase and trehalase activities were significantly increased as well as endoplasmic reticulum membrane-bound glucose-6-phosphatase.

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.

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.

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 all 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 β‐galactosidase activity was shown to be unaffected by hydrocortisone treatment. This observation clearly illustrates that the two intestinal β‐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.

Epidermal growth factor influences cell proliferation, glycoproteins, and lipase activity in human fetal stomach

BACKGROUND & AIMS The role of epidermal growth factor (EGF) in the functional development of human stomach is unknown. The aim of this study was to establish the distribution and cellular localization of EGF receptors in developing gastric mucosa and to determine the effects of EGF on epithelial cell proliferation and differentiation. METHODS Quantitative radioautography with 125I-EGF and indirect immunofluorescence using an antibody for human EGF receptor were performed using fetal gastric tissues (12-20 weeks of gestation). The effects of EGF (1, 10, and 100 ng/mL) on DNA synthesis, glycoprotein synthesis, and lipase and pepsin activities in fetal gastric explants maintained in serum-free organ culture were determined. RESULTS EGF receptors were present as early as 12 weeks of gestation and localized on basolateral membranes of all gastric epithelial cells. DNA and glycoprotein synthesis were significantly increased after 24 hours of culture in the presence of EGF. Unlike pepsin activity, lipase activity was modulated by EGF, and a significant diminution of the tissue lipolytic activity was noted after 5 days of culture. CONCLUSIONS This study clearly indicates the influence of EGF on the proliferation and differentiation of gastric epithelium, suggesting an important role for EGF in fetal development of the human gastric mucosa.

Calcitriol regulates the expression of the genes encoding the three key vitamin D3 hydroxylases and the drug‐metabolizing enzyme CYP3A4 in the human fetal intestine

background and aims The human fetal jejunum has been shown to harbour the vitamin D3 (D3) nuclear receptor (VDRn) and to be responsive to calcitriol/1,25‐dihydroxyvitamin D3[1,25(OH)2D3] through modulation of proliferation and differentiation processes. The aim of the study was to evaluate the presence as well as the effect of 1,25(OH)2D3 exposure on the expression levels of the three key D3‐hydroxylase gene transcripts (25‐hydroxylase, CYP27A; 24‐hydroxylase, CYP24; 1α‐hydroxylase, CYP27B1) as well as that of the 1,25(OH)2D3‐responsive endobiotic/xenobiotic metabolizing enzyme CYP3A4 (which is also considered a major detoxifiying enzyme) in the human proximal and distal intestine.methods Specimens from normal fetuses ranging from 15 to 20 weeks of gestation were obtained following elective termination of normal pregnancies. Intestinal explants were cultured for a period of 24 h or 48 h with 10−7 m 1,25(OH)2D3. All data were compared to paired‐control cultures without 1,25(OH)2D3. Total RNA was extracted and cDNA synthesized by RT‐PCR. The cDNA obtained was amplified by radioactive PCR, the signal intensity evaluated by densitometric analyses and expressed in relation to the levels of GAPDH.

Expression of CYP27A, a gene encoding a vitamin D-25 hydroxylase in human liver and kidney

Vitamin D3 (D3) is not active but must be hydroxylated at C‐25 in liver before acquiring its hormonal potential in the kidney. The sterol‐27 hydroxylase (gene symbol: CYP27A) catalyses the oxidation of sterol side chain in bile acid synthesis but the enzyme is also known as a D3−25 hydroxylase.

Peutz-Jeghers syndrome. Association of duodenal and bilateral breast cancers in the same patient.

SummaryA female patient with Peutz-Jeghers syndrome successively developed bilateral breast carcinoma and malignant transformation of a duodenal hamartomatous polyp, from which she died.