Summary
Human fetal colon (14–16 weeks gestation) was cultured as explants for 15 days in serum-free Leibovitz L-15 medium at 37°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 activites, 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.
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References
Arsenault P, Ménard D (1984a) Insulin influences the maturation and proliferation of suckling mouse intestinal mucosa in serumfree organ culture. Biol Neonat 46:229–236
Arsenault P, Ménard D (1984b) Comparative study of the effect of hydrocortisone and thyroxine on suckling mouse small intestine in organ culture. Comp Biochem Physiol 77A:721–725
Arsenault P, Ménard D (1985) Influence of hydrocortisone on human fetal small intestine in organ culture. J Pediatr Gastroenterol Nutr 4:893–901
Auricchio S, Stallato A, Devizio B (1981) Development of brush border peptidases in human and rat small intestine during fetal and neonatal life. Pediatr Res 15:991–995
Autrup H, Barrett LS, Jackson FE, Jesudan ML, Stoner G, Phelps P, Trump BF, Harris CC (1978) Explant culture of human colon. Gastroenterology 74:1248–1257
Beaulieu JF, Ménard D, Calvert R (1985) Influence of epidermal growth factor (EGF) on the maturation of the fetal mouse small intestine in organ culture. J Pediatr Gastroenterol Nutr 4:476–481
Bell L, Williams L (1982) A scanning and transmission electron microscopical study of the morphogenesis of human colonic villi. Anat Embryol 165:437–455
Benjawatanopon C, Bell LE, Williams L (1982) Ultrastructural localization of alkaline phosphatase in adult human large intestine. Gut 23:134–136
Berteloot A, Chabot JG, Ménard D, Hugon JS (1979) Organ culture of adult mouse intestine. III. Behavior of the proteins, DNA content and brush border membrane enzymatic activities. In vitro 15:294–299
Calvert R, Pusterla A (1974) Epon-embedded tissue stained with aldehyde-fuchsin for autoradiography. Stain Technol 49:323–327
Dahlqvist A, Lindberg T (1966) Development of the intestinal disaccharidases and alkaline phosphatase activities in the human fetus. Clin Sci 30:517–528
Eastwood GL (1977) Gastrointestinal epithelial renewal. Gastroenterology 72:962–975
Eichholz A (1976) Structural and functional organization of brush border of intestinal epithelial cells. III. Enzyme activites and chemical composition of various fractions of Tris-disrupted brush borders. Biochim Biophys Acta 135:475–482
Giles KW, Myers A (1965) An improved diphenylamine method for the estimation of deoxyribonucleic acid. Nature 206:93
Grand RJ, Watkins JB, Torti FM (1976) Development of the human gastrointestinal tract. Gastroenterology 70:790–810
Howdle PD (1983) Organ culture in the study of the gastrointestinal tract in health and disease. Clin Sci 65:105–110
Hugon JS, Calvert R, Ménard D (1982) In vitro methods and their applications. In: Chadwick US, Philips SF (eds) BIMR Gastroenterology, vol 2, Small intestine. Butterworth, London, pp 58–72
Johnson FK (1913) The development of the mucous membrane of the large intestine and vermiform process in the human embryo. Am J Anat 14:187–233
Kopriwa BM, Leblond CP (1962) Improvements in the coating technique of radioautography. J Histochem Cytochem 10:169–184
Lacroix B, Kedinger M, Simon-Assmann P, Rousset M, Zweibaum A, Haffen K (1984) Developmental pattern of brush border enzymes in the human fetal colon. Correlation with some morphogenetic events. Early Human Dev 9:95–103
Lev R, Orlic D (1974) Histochemical and radioautographic studies of normal human fetal colon. Histochemistry 39:301–311
Lev R, Griffiths WC (1982) Colonic and small intestinal alkaline phosphatase. A histochemical and biochemical study. Gastroenterology 82:1427–1435
Lindberg T (1966) Intestinal dipeptidase: characterization, development and distribution of intestinal dipeptidases of the human fetus. Clin Sci 30:505–515
Lloyd S, Whelan W (1969) An improved method for enzymic determination of glucose in the presence of maltose. Anal Biochem 30:467–470
Lonnerholm G, Wistrand P (1983) Carbonic anhydrase in the human fetal gastrointestinal tract. Biol Neonat 44:166–176
Lowry OH, Rosebrough NF, Farr AL, Randall RJ (1951) Protein measurement with Folin phenol reagent. J Biol Chem 193:265–275
Malo C, Arsenault P, Ménard D (1983) Organ culture of the small intestine of suckling mouse in a serum-free medium. Cell Tissue Res 228:75–84
Ménard D, Arsenault P (1985) Explant culture of human fetal small intestine. Gastroenterology 88:691–700
Moog F (1982) Induction of brush border enzymes in culture. J Pediat Gastroenterol Nutr 1:161–162
Potter GD, Lester R (1984) The developing colon and nutrition. J Pediatr Gastroenterol Nutr 3:485–487
Schlegel-Haueter S, Hore P, Kerry KR, Semenza G (1972) The preparation of lactase and glucoamylase of rat small intestine. Biochim Biophys Acta 58:506–519
Simon PM, Kedinger M, Raul F, Grenier JF, Haffen K (1982) Organ culture of suckling rat intestine: Comparative study of various hormones on brush border enzymes. In vitro 18:339–346
Trier JS (1980) Organ culture of the mucosa of human small intestine. In: Harris CC, Trump DF, Stoner GD (eds) Methods in cell biology, vol 21 B, Normal tissue and cell culture. New York, Academic Press, pp 365–384
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Ménard, D., Arsenault, P. Human fetal colon in organ culture. Anat Embryol 176, 441–448 (1987). https://doi.org/10.1007/BF00310085
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DOI: https://doi.org/10.1007/BF00310085