scholarly journals Epithelioid cell cultures from rat small intestine. Characterization by morphologic and immunologic criteria.

1979 ◽  
Vol 80 (2) ◽  
pp. 248-265 ◽  
Author(s):  
A Quaroni ◽  
J Wands ◽  
R L Trelstad ◽  
K J Isselbacher
Keyword(s):  
Basement Membrane ◽  
Amorphous Material ◽  
Population Doubling ◽  
Antigenic Determinants ◽  
Intestinal Epithelial ◽  
Epithelioid Cells ◽  
Intestinal Crypt ◽  
Ultrastructural Studies ◽  
Small Intestinal

Rat small intestinal epithelial cell lines have been established in vitro and subcultured serially for periods up to 6 mo. These cells have an epithelioid morphology, grow as monolayers of closely opposed polygonal cells, and during the logarithmic phase of growth have a population doubling time of 19--22 h. Ultrastructural studies revealed the presence of microvilli, tight junctions, an extensive Golgi complex, and the presence of extracellular amorphous material similar in appearance to isolated basement membrane. These cells exhibit a number of features characteristic of normal cells in culture; namely, a normal rat diploid karyotype, strong density inhibition of growth, lack of growth in soft agar, and a low plating efficiency when seeded at low density. They did not produce tumors when injected in syngeneic animals. Immunochemical studies were performed to determine their origin using antisera prepared against rat small intestinal crypt cell plasma membrane, brush border membrane of villus cells and isolated sucrase-isomaltase complex. Antigenic determinants specific for small intestinal epithelial (crypt and villus) cells were demonstrated on the surface of the epithelioid cells, but they lacked immunological determinants specific for differentiated villus cells. An antiserum specifically staining extracellular material surrounding the cells cultured in vitro demonstrated cross-reactivity to basement membrane in rat intestinal frozen sections. It is concluded that the cultured epithelioid cells have features of undifferentiated small intestinal crypt cells.

Differentiation of rat small intestinal epithelial cells by extracellular matrix

1988 ◽  
Vol 254 (3) ◽  
pp. G355-G360 ◽  
Author(s):  
K. M. Carroll ◽  
T. T. Wong ◽  
D. L. Drabik ◽  
E. B. Chang
Keyword(s):  
Extracellular Matrix ◽  
Basement Membrane ◽  
Glucose Absorption ◽  
Cell Maturation ◽  
Intestinal Cell ◽  
Synthetic Activity ◽  
Intestinal Epithelial ◽  
Intestinal Cell Line ◽  
Small Intestinal

The role of extracellular matrix as a determinant of intestinal cell maturation was explored by growing a normal, but immature, rat small intestinal cell line (IEC-6) on basement membrane extract from Engelbreth-Holm-Swarm (EHS) sarcoma cells (ECM). Grown on plastic or glass, these cells are relatively immature and proliferate rapidly. In contrast, cells on ECM attached more rapidly, stopped proliferating, and rapidly organized into multicellular complex structures. Ultrastructurally, cells grown on ECM displayed significantly more mitochondria, rough endoplasmic reticulum, apical microvilli, and complex golgi apparatus, consistent with greater maturity and synthetic activity. By indirect immunofluorescence, sucrase, alkaline phosphatase, and cellular apolipoprotein B were present in cells grown on ECM only. In contrast to cells grown on glass, these cells also demonstrated Na-dependent glucose absorption, a function unique to mature villus cells (7). We conclude that the basement membrane may be a key determinant of intestinal epithelial cell maturation. The development of a mature villuslike intestinal cell in vitro may have wide application for future studies of induction and regulation of intestinal maturation and function.

Schistosoma mansoni: a comparative study of artificially transformed schistosomula and schistomula recoverd after cercarial penetration of isolated skin

Parasitology ◽  
1977 ◽  
Vol 74 (1) ◽  
pp. 73-86 ◽  
Author(s):  
Linda H. Brink ◽  
Diane J. McLaren ◽  
S. R. Smithers
Keyword(s):  
Comparative Study ◽  
Schistosoma Mansoni ◽  
Amorphous Material ◽  
Surface Membrane ◽  
Antigenic Determinants ◽  
Agglutination Reaction ◽  
Rat Serum ◽  
Mechanical Separation ◽  
B Blood Group

A comparison was made of the ultrastructure, development and antigenic nature of the surfaces and of the viability of three types of schistosomula of Schistosoma mansoni: schistosomula formed afrer cercariae had penetrated isolated skin (SS), schistosomula produced after mechanical separation of cercarial tails from bodies (MS), and schistosomula transformed from cercariae after incubation in fresh rat serum (RS).Within 2 h of transformation, the surface membrane of all three types of schistosomula had changed from trilaminate to heptalaminate structures and SS and MS had lost their cercarial glycocalyx. Initially a dense amorphous material was demonstrated on the surfaces of RS, which was thought to be the result of an interaction between a factor in rat serum and the glycocalyx: this material was greatly reduced within 2 h of transformation. The pre-acetabular glands of SS were emptied while those of MS and RS retained their contents. Immunofluorescent studies showed that all schistosomula bound serum from mice immune to S. mansoni, but the binding was stronger with MS and RS. The mixed agglutination reaction demonstrated the presence of human A and B blood group-like antigenic determinants on approximately 30% of 3 h old SS; these determinants were not detected on MS or RS. In vitro, the development of MS and RS was similar to SS; the first schistosomula reached the ‘gut-closed’ stage by day 10; 50–70% of SS reached this stage by day 12, in contrast to only 25–50% of MS and RS. Between 28 and 45% of all schistosomula developed to maturity when injected intravenously into mice.It was concluded that the two types of artificially prepared schistosomula fultil the main criteria of transformation from cercaria to schistosomulum. Further, it is suggested that MS are the most appropriate source of material for immunochemical and physiological studies.

scholarly journals Intestinal crypt stem cells possess high levels of cytoskeletal-associated phosphotyrosine-containing proteins and tyrosine kinase activity relative to differentiated enterocytes.

1989 ◽  
Vol 109 (5) ◽  
pp. 2139-2144 ◽  
Author(s):  
D R Burgess ◽  
W P Jiang ◽  
S Mamajiwalla ◽  
W Kinsey
Keyword(s):  
Stem Cells ◽  
Growth Factors ◽  
Tyrosine Kinase ◽  
Protein Tyrosine Kinase ◽  
Kinase Activity ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Tyrosine Kinase Activity ◽  
Intestinal Crypt

Growth and differentiation of stem cells is thought to be regulated by growth factors and responding protein tyrosine kinase activities. Comparing mitotic stem cells from the adult intestinal epithelium, isolated from the crypts of Lieberkuhn, with isolated differentiated absorbtive cells we find major differences in the levels of phosphotyrosine-containing proteins. Crypt stem cells possess two major phosphotyrosine-containing polypeptides of 36 and 17 kD which have greater than 15 times more phosphotyrosine than that present in the polypeptides of differentiated enterocytes. Tyrosine kinase activity and similar phosphotyrosine-containing proteins are associated with the Triton cytoskeleton. Moreover, crypt tyrosine kinase(s) is active in vitro in phosphorylating similar cytoskeleton-associated substrates. These results suggest that cytoskeleton-associated phosphotyrosine kinase(s) and their substrates may play a role in growth and differentiation of adult intestinal epithelial cells.

Proinsulin stimulates growth of small intestinal crypt-like cells acting via specific receptors

1999 ◽  
Vol 276 (2) ◽  
pp. E262-E268 ◽  
Author(s):  
Peter M. Jehle ◽  
Rolf D. Fussgaenger ◽  
Niklas K. O. Angelus ◽  
Robert J. Jungwirth ◽  
Bernhard Saile ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Specific Binding ◽  
Physiological Role ◽  
Intestinal Epithelial ◽  
Binding Studies ◽  
C Peptide ◽  
Intestinal Crypt ◽  
Igf I ◽  
Confluent Cell ◽  
Small Intestinal

The mechanisms that regulate cell turnover in the intestinal epithelium are incompletely understood. Here we tested the hypothesis that proinsulin, present in serum and pancreatic juice in picomolar concentrations, stimulates growth of the rat small intestinal crypt-like cell line IEC-6 under serum-free conditions. Proinsulin binding was assessed by competitive ligand binding studies. Proinsulin and insulin-like growth factor I (IGF-I) stimulated cell proliferation up to threefold above controls, with half-maximal action already in the picomolar range and with additive effects. In early confluent cell monolayers, proinsulin bound with higher affinity (IC50 1.3 ± 0.05 nM) and capacity (87,200 ± 2,500 receptors/cell) than IGF-I (4.0 ± 0.6; 23,700 ± 2,200, P < 0.05). C-peptide competed with 10-fold lower affinity for binding of125I-proinsulin but not for125I-IGF-I or125I-insulin, suggesting a specific binding epitope of the proinsulin molecule within or close to the C-peptide region. In contrast, insulin showed ∼100-fold lower binding affinity and growth-promoting potency than proinsulin or IGF-I. We conclude that proinsulin stimulates growth of small intestinal crypt cells through specific binding and may play a physiological role in the regulation of intestinal epithelial cell proliferation.

scholarly journals Bovine dairy complex lipids improve in vitro measures of small intestinal epithelial barrier integrity

PLoS ONE ◽  
2018 ◽  
Vol 13 (1) ◽  
pp. e0190839 ◽  
Author(s):  
Rachel C. Anderson ◽  
Alastair K. H. MacGibbon ◽  
Neill Haggarty ◽  
Kelly M. Armstrong ◽  
Nicole C. Roy
Keyword(s):  
Epithelial Barrier ◽  
Intestinal Epithelial ◽  
Intestinal Epithelial Barrier ◽  
Barrier Integrity ◽  
Complex Lipids ◽  
Small Intestinal

Transcellular transport of calcium and inorganic phosphate in the small intestinal epithelium

1981 ◽  
Vol 240 (6) ◽  
pp. G409-G416 ◽  
Author(s):  
H. Murer ◽  
B. Hildmann
Keyword(s):  
Brush Border ◽  
Inorganic Phosphate ◽  
Membrane Vesicles ◽  
Calcium Flux ◽  
Intestinal Epithelial ◽  
Transcellular Transport ◽  
Lateral Membrane ◽  
Sodium Flux ◽  
Small Intestinal

Transport mechanisms involved in the small intestinal handling of inorganic phosphate and calcium have been studied by different in vitro methods during the last few years. In concordance with studies on intact epithelial preparations, studies with brush-border and basal-lateral membrane vesicles isolated from the small intestinal epithelial cell revealed that transcellular calcium and inorganic phosphate fluxes are coupled to transcellular sodium flux, i.e., secondary active via coupling to the primary active sodium flux. A sodium-coupled mechanism in the brush-border membrane leads to cellular accumulation of inorganic phosphate. A sodium-coupled mechanism leads to extrusion of calcium from the cell into the serosal interstitium. A primary active transport mediated by the Ca-ATPase and located in the basal-lateral membrane also exists for calcium. Regulation of transcellular phosphate and calcium flux proceeds via altered influx rates at the luminal cell pole.

scholarly journals Development of an endogenous epithelial Na+/H+exchanger (NHE3) in three clones of Caco-2 cells

1999 ◽  
Vol 277 (2) ◽  
pp. G292-G305 ◽  
Author(s):  
Andrzej J. Janecki ◽  
Marshall H. Montrose ◽  
C. Ming Tse ◽  
Fermin Sanchez de Medina ◽  
Alain Zweibaum ◽  
...  
Keyword(s):  
Membrane Domains ◽  
Intestinal Epithelial ◽  
Major Mechanism ◽  
Kinase C ◽  
Cell Clones ◽  
Nhe1 Activity ◽  
Epidermal Growth ◽  
Small Intestinal ◽  
Vitro Model

Expression of endogenous Na+/H+exchangers (NHEs) NHE3 and NHE1 at the apical (AP) and basolateral (BL) membrane domains was investigated in three clones (ATCC, PF-11, and TC-7) derived from the human adenocarcinoma cell line Caco-2. In all three clones, NHE1 was the only isoform detected at the BL domain during 3 to 22 postconfluent days (PCD). In clone PF-11, the BL NHE1 activity increased up to 7 PCD and remained stable thereafter. Both NHE1 and NHE3 were found at the AP domain at 3 PCD and contributed 67 and 33% to the total AP Na+/H+exchange, respectively. The AP NHE3 activity increased significantly from 3 to 22 PCD, from 93 to 450 μM H+/s, whereas AP NHE1 activity decreased from 192 to 18 μM H+/s during that time. Similar results were obtained with the ATCC clone, whereas very little AP NHE3 activity was observed in clone TC-7. Surface biotinylation and indirect immunofluorescence confirmed these results and also suggested an increase in the number of cells expressing NHE3 being the major mechanism of the observed overall increase in NHE3 activity in PF-11 and ATCC clones. Phorbol 12-myristate 13-acetate (PMA, 1 μM) acutely inhibited NHE3 activity by 28% of control, whereas epidermal growth factor (EGF, 200 ng/ml) stimulated the activity by 18%. The effect of PMA was abolished by the protein kinase C (PKC) inhibitor 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine, suggesting involvement of PKC in the PMA-induced inhibition of NHE3. Similar magnitude of inhibition by PMA and stimulation by EGF was observed at 7 and 17 PCD, suggesting the development of regulatory mechanisms in the early postconfluent period. Taken together, these data suggest a close similarity of membrane targeting and regulation of endogenous NHE3 between Caco-2 cells and native small intestinal epithelial cells and support the usefulness of some Caco-2 cell clones as an in vitro model for studies on physiology of NHE3 in the intestinal epithelium.

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