Biomedical subjects

P Simo

Publications and source records attributed to P Simo.

8 recordsLinked to original sources

PubMed1995

Extracellular matrix components in intestinal development.

Intestinal morphogenesis and differentiation are dependent on heterotypic cell interactions between embryonic epithelial cells (endoderm) and stromal cells (mesenchyme). Extracellular matrix molecules represent attractive candidates for regulators of these interactions. The structural and functional diversity of the extracellular matrix as intestinal development proceeds is demonstrated by 1) spatio-temporal specific expression of the classically described constituents, 2) the finding of laminin and collagen IV variants, 3) changes in the ratio of individual constituent chains, and 4) a stage-specific regulation of basement membrane molecule production, in particular by glucocorticoids. The orientation/assembly of these extracellular matrix molecules could direct precise cellular functions through interactions via integrin molecules. The involvement of extracellular matrix, and in particular basement membrane molecules in heterotypic cell interactions leading to epithelial cell differentiation, has been highlighted by the use of experimental models such as cocultures, hybrid intestines and antisense approaches. These models allowed us to conclude that a correct elaboration and assembly of the basement membrane, following close contacts between epithelial and fibroblastic cells, is necessary for the expression of differentiation markers such as digestive enzymes.

Animals↗

PubMed1994

[Expression of laminin is correlated to the differentiation of human colonic cancer cells].

The level and molecular composition of laminin, a major basement membrane glycoprotein formed of three chains (A, B1 and B2) have been analyzed in various human colonic cancer cells (Caco-2 and HT29). The synthesis of laminin over a 24 h period, corresponding to cellular and secreted molecules purified by affinity chromatography, was the highest in the more differentiated cells. Immunocytochemical detection of the constituent chains of laminin in permeabilized cells as well as after separation on polyacrylamide gels showed that A, B1 and B2 chains were expressed in Caco-2 cells, whereas A chain was not detected in HT29 cells. When cancer cells were cultured on a monolayer of confluent fibroblastic cells, laminin was deposited at the basement membrane level only in the case of the most differentiated cells. In an attempt to define the role of laminin-A chain, transfection of Caco-2 cells with A chain antisense cDNA was performed; among the clones obtained, 3 were deficient for the target polypeptide. The consequences of the absence of laminin-A chain on basement membrane formation, cellular differentiation and tumor invasion will be currently determined.

Basement Membrane↗

PubMed1992

Dual and asynchronous deposition of laminin chains at the epithelial-mesenchymal interface in the gut.

The production of laminin by 14-day fetal rat intestinal endoderm and mesenchyme was investigated. The amount of neosynthesized laminin was measured after purification using affinity chromatography. Chain composition of laminin was analyzed by immunoblotting and immunofluorescence staining. The data show that both embryonic intestinal tissue components synthesize laminin and that A and B1/B2 chains were detected in both endodermal and mesenchymal cells. The cellular source of laminin found at the epithelial basement membrane has been studied by immunocytochemistry in rat/chick or mouse/chick interspecies hybrid intestines taken at various stages of development. Immunodetection of the whole laminin molecule and of the individual A and B1/B2 chains by rodent-specific polyclonal and monoclonal antibodies at the basement membrane level in these hybrid intestines revealed (a) laminin molecules, which originate from both mesenchymal and endodermal cells; (b) deposition of A and B1/B2 chains by endodermal cells, regardless of the stage of growth of the hybrid intestines; and (c) asynchronous deposition of the various chains of laminin into the basement membrane by the mesenchyme. B1/B2 chains are deposited concomitant with contact with the epithelium, whereas A chains appear only later (13 days after grafting). These data reinforce the suggestion from previous studies that cooperation between epithelium and mesenchyme is necessary for the formation of a complete basement membrane in the developing intestine.

Animals↗

PubMed1992

Mesenchyme-mediated effect of dexamethasone on laminin in cocultures of embryonic gut epithelial cells and mesenchyme-derived cells.

Previous studies have shown that glucocorticoids accelerate intestinal maturation and that this process is mediated by the mesenchymal cells. The possible involvement of laminin (LN), a basement membrane component, in this mesenchymal mediation has been analyzed. For this purpose, the influence of dexamethasone (DX) on the synthesis of LN, its chain composition and its cellular distribution has been examined biochemically and immunocytochemically in two different mesenchyme-derived cell populations, fetal intestinal mesenchymal cells and fetal skin fibroblasts, as well as in cocultures of intestinal endodermal cells seeded on top of confluent fetal skin fibroblasts. Neither the amount of metabolically labeled LN purified by affinity chromatography (expressed per mg cell proteins), nor the A versus B chain ratio monitored after separation on gel electrophoresis and immunoblotting, showed significant differences after 5 days of DX treatment. However, glucocorticoids induced a shift from secreted to cell-associated LN molecules paralleling a striking difference in the immunostaining pattern of intracellular and surface LN in the mesenchyme-derived cell monocultures; the granular intracytoplasmic LN staining in the control cultures was replaced by a fibrillar organization of LN molecules concomitantly with an increased accumulation at the cell surface. In 2-day DX-treated cocultures, there was an acceleration of LN deposition at the epithelial-fibroblastic interface, which accompanied the enhanced expression of epithelial cell differentiation markers (brush border digestive enzymes). These DX-induced changes can be blocked by the addition of anti-LN antibodies in the culture medium. These findings further support the concept that glucocorticoid action on intestinal epithelial cells involves alterations in the extracellular microenvironment, assessed here for LN molecules, occurring at the level of the mesenchymal cell compartment. These changes may contribute to an accelerated organization of LN at the epithelial-mesenchymal interface and subsequently to epithelial differentiation.

Animals↗

PubMed1991

Altered deposition of basement-membrane molecules in co-cultures of colonic cancer cells and fibroblasts.

Two human colon carcinoma cell lines, HT29 and Caco-2 were co-cultured with fetal rat or human skin fibroblasts. Their morphological features, ultra-structural characteristics at the heterologous cell interface, and the deposition of basement-membrane molecules [laminin, type-IV collagen, heparan sulfate proteoglycan (HSPG)] at the epithelial-stromal junction were analyzed. The 2 cell lines behaved differently. HT29 cells did not spread on the fibroblasts and grew as clusters, while Caco-2 cells formed a monolayer over the fibroblastic feeder layer. Only the latter carcinoma cells exhibited cytoplasmic processes towards the fibroblasts and, after 5 days in co-cultures, a structured basement membrane (BM). The immunocytochemical analysis of the BM constituents revealed the absence of the molecules studied at the sites of heterologous contacts in the case of HT29 cells. In contrast, in the co-cultures comprising Caco-2 cells, laminin and type-IV collagen were progressively deposited in a polar fashion at the epithelial-fibroblastic interface which, however, remained devoid of HSPG molecules. Together with earlier data indicating a dual origin of the BM molecules located at the epithelial-fibroblastic interface in normal intestine, the present study shows that the cancer cells as well as the fibroblastic ones under the influence of carcinoma cells display an altered capacity to synthesize and/or secrete BM molecules. The extent of such abnormalities correlates with the differentiation of the cells. Finally, these modifications occur concomitantly with alterations in cell interactions which vary among cell lines.

Adenocarcinoma↗

PubMed1991

Changes in glycosaminoglycan expression in the rat developing intestine.

Synthesis of glycosaminoglycan (GAG) chains was studied in the developing rat intestine. Intestinal segments, taken at various developmental stages, were exposed to 3H-glucosamine and 35S-sulfate for 6 hours. The amounts of 3H-GAGs (total GAGs) and of 35S-GAGs (sulfated GAGs) showed a clear age-dependence, with a broad maximum in the fetal period when dramatic growth and morphogenesis occur. Characterization of individual GAG species indicated that hyaluronic acid (HA), heparan and chondroitin sulfate (HS and CS) synthesis was modified quantitatively or qualitatively during development: decrease of HA with age; production of undersulfated HS molecules during embryonic life; shift towards a lower hydrodynamic form of HA and HS molecules after birth. We postulate that these alterations are crucial in the elaboration of an age-related specific extracellular microenvironment allowing intestinal growth and differentiation.

Aging↗

PubMed1991

Changes in the expression of laminin during intestinal development.

The expression of laminin, a major glycoprotein constituent of basement membranes, was investigated in the rat developing intestine. The biosynthesis of laminin was studied after metabolic labeling of intestinal segments taken at various stages of development; the neosynthesized laminin was purified by affinity chromatography on heparin-Sepharose. Immunoblotting and immunoprecipitation experiments allowed us to analyze its constitutive chains. The data show that laminin is synthesized in very large amounts at 16-18 days of gestation concomitant with the onset of intestinal morphogenetic movements, i.e. villus emergence. Evaluation of the relative proportion of individual laminin polypeptides shows that laminin B1/B2 chains are produced in excess of A chains whatever the developmental stage considered. Interestingly at 17 days of gestation, levels of laminin A subunits are maximal. A second rise in the A/B chain ratio starts around birth and continues until adulthood. These quantitative data are corroborated by the immunocytochemical detection of laminin A and B chains, which revealed a specific spatiotemporal pattern. The finding that laminin A chains are located in the basement membrane of growing villi and of adult crypts raises the possibility that they may be involved in the process of cell growth and/or in the establishment of cell polarity by creating a specialized extracellular microenvironment.

Animals↗

PubMed1990

Synthesis of basement membrane proteins in the small intestine.

Are the basement membrane (BM) molecules involved in epithelial-mesenchymal cell interactions known to be instrumental in intestinal development and differentiation? Several findings argue in favor of this assumption. First, quantitative and/or qualitative changes in type IV collagen, laminin-nidogen and heparan sulfate proteoglycan (HSPG) are obvious at the phases of intensive morphogenesis. Second, BM molecules deposited at the epithelial-mesenchymal interface are of dual origin: HSPG being produced by the epithelial cell population, while others like type IV collagen and laminin are mainly produced by the mesenchymal compartment. Third, the formation of the BM requires an actual contact between the epithelial and mesenchymal cells and always precedes the expression of differentiation markers in the epithelial cells. These data suggest that BM molecules display an instructive role in intestinal cell interactions.

Animals↗