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At least 109 records · Page 6Linked to original sources
PubMed2005

Involvement of SDS-stable higher M(r) forms of bovine normal milk alpha-lactalbumin in inducing intestinal IEC-6 cell death.

Monomeric 14-kDa bovine alpha-lactalbumin was purified with a preparation of lower molecular weight whey protein concentrate from Holstein cow normal milk followed by size exclusion chromatography. The protein showed a stimulatory rather than an inhibitory effect on the proliferation of a cultured IEC-6 cell line from the rat small intestine. But incubation in 30% trifluoroethanol/acetate buffer (pH 5.5) at 37 degrees C for 5 d in a slowly rotating test tube rendered it highly cytotoxic with concomitant appearance of SDS-stable 20- and 30-kDa forms of alpha-lactalbumin on electrophoresis. Furthermore, alpha-lactalbumin obtained by a one-step purification procedure by affinity chromatography on an anti-alpha-lactalbumin antibody column from the lower molecular weight whey protein concentrate, which had been found to contain several SDS-stable higher M(r) forms of alpha-lactalbumin, exhibited potent inhibitory activity on IEC-6 cell growth. These results indicate the involvement of SDS-stable higher M(r) forms of bovine normal milk alpha-lactalbumin in inducing cell death on the intestinal IEC-6 cell line.

Animals↗
PubMed1993

Role of tyrosine kinase and phosphotyrosine phosphatase in growth of the intestinal crypt cell (IEC-6) line.

The roles of gastrin and sodium vanadate in proliferation were examined in cultured IEC-6 cells that are mitotically active and derived originally from jejunal crypts of the rat intestine. Incubation of the cells in the presence of gastrin at a concentration of 250 ng/ml or of sodium vanadate at a concentration of 0.2 mM leads to a 60% increase in cell growth in 24 hr. The stimulated growth in both cases was inhibited by genistein, a tyrosine kinase inhibitor. Incubation in the presence of gastrin and sodium vanadate together produced a small, albeit significant, potentiation of growth of the cells. Gastrin as well as sodium vanadate also promoted the phosphorylation on tyrosine of a similar group of proteins with molecular masses of 42, 45, 52, 60, 78, and 120 kDa. The phosphorylations were rapidly occurring as early as 5 min and lasted for only 15 min. Several proteins were detected in normal IEC-6 cells, including GTPase activating protein, raf1 kinase, phospholipase C gamma-1, and phosphoinositide 3-kinase. The results suggest that gastrin and sodium vanadate induce growth of IEC-6 cells by stimulation of tyrosine kinase and/or inhibition of tyrosine phosphatase. The gastrin and sodium vanadate effects also involve the phosphorylation of a number of proteins, the identities of which are not known at present but may include some of the kinases that are frequently associated with cell growth, such as mitogen-activated protein kinase, raf1 kinase, phosphoinositide 3-kinase, and others.

Animals↗
PubMed2005

[Quantitatively determining trace elements Co and Bi in gelatin by inductively coupled plasma atomic emission spectrometry method--applying IEC model to correct the spectral interference of Fe to Co].

By ICP-AES, the authors established the method to quantitatively determine the trace elements Co and Bi in gelatin. The authors processed the unknown sample with wet digesting method. Trace elements in gelatin could be quantitatively determined at the same time. To correct the spectral interference of Fe to Co at specific wavelength, the Interelement Correction(IEC) model was established. The results of the experiment showed that the IEC model could effectively erase the spectral interference of Fe to Co. The result of the spectral correction was greatly superior to the result where the spectral interference weren't corrected by the IEC model. The method was accurate, quick with high performance and wide linear range. The linear correlation coefficient of the tested elements was over 0.999 90. The relative standard deviation (RSD%) was less than 2.00%. The recovery rates were in the range of 98%-107%.

Bismuth↗
PubMed1990

Expression of sodium-linked nucleoside transport activity in monolayer cultures of IEC-6 intestinal epithelial cells.

Mature, confluent monolayer cultures of IEC-6 rat intestinal epithelial cells in conventional growth media express both Na(+)-linked, concentrative nucleoside transport (NT) activity and equilibrative, inhibitor-sensitive NT activity, but do not show morphologic differentiation. Na(+)-dependent fluxes of Ado and formycin B were minor in early subconfluent IEC-6 monolayers, but increased severalfold to become the major component of influx of these agents in confluent monolayers grown in medium containing Nu-Serum, a commercial medium supplement with a low serum content. In monolayers cultured in medium with fetal bovine serum, cell proliferation rates were similar to those in medium supplemented with Nu-Serum, but expression of Na(+)-linked NT activity was 6-8-fold lower than in monolayers grown in the latter medium. Inclusion of hydrocortisone in growth medium with Nu-Serum caused a 2-fold increase in the expression of Na(+)-linked NT activity. Experiments in which components of medium supplementation were withheld showed that insulin and epidermal growth factor were important in expression of the Na(+)-linked NT activity. Because the Na(+)-linked NT system has a brush border location in fresh intestinal epithelium, it is concluded that the regulated expression of this activity in the IEC-6 monolayers is a differentiative change.

Adenosine↗
PubMed1985

Response of cultured IEC-17 normal rat intestinal epithelial cells to X radiation.

The growth parameters and radiosensitivity of normal rat intestinal epithelial cells, IEC-17, were studied. The cells were cultured by standard methods and exposed to an array of doses (1-12 Gy) of 250 kVp X rays. The survival curves generated exhibited no initial shoulder and were bimodal. The Do of the first component was about 0.2 Gy and the second component. 5.0 Gy. The ability of this cell line to repair sublethal lesions was examined by fractionation studies; repair was completed within 60 min after the first dose. When Chinese hamster ovary (CHO) cells were grown under the same conditions used for the IEC-17 cells and then irradiated with single doses, a typical survival curve with a Do of 1.4 Gy was obtained. The survival curves obtained for the IEC-17 cell line are consistent with the response of a morphologically distinct single population containing two functionally separate types of cells.

Animals↗
PubMed1997

Cytoskeletal association of epidermal growth factor receptor and associated signaling proteins is regulated by cell density in IEC-6 intestinal cells.

Epidermal growth factor (EGF) mediates a variety of physiologic responses in rat intestine. EGF receptor (EGFR) responsiveness to EGF is mediated by the surface expression of high affinity EGFR, which is associated with the cytoskeleton (CSK). EGFR signal transduction appears to be mediated by the CSK association of EGFR and related signaling proteins. In the nontransformed intestinal cell line IEC-6, expression of EGFR, Src homology and collagen protein (SHC), phospholipase C gamma 1 (PLC gamma), and their tyrosine phosphorylation in response to EGF was assayed by immunoblot. The distribution of EGFR and tyrosine-phosphorylated EGFR was regulated by cell density. At confluence, EGFR and tyrosine-phosphorylated EGFR were predominantly associated with the Triton X-100-insoluble CSK at confluence, while predominantly Triton X-100-soluble at subconfluence. PLC gamma was predominantly soluble at both states of confluence. Confluent but not subconfluent IEC-6 cells demonstrated a cascade of EGF-mediated events consisting of a transient CSK association of PLC gamma with EGFR, a brief expression of tyrosine-phosphorylated PLC gamma, a brief increase in PLC gamma CSK association, and a prolonged soluble association of PLC gamma with the EGFR. EGF led to an increase in the CSK association of SHC at both states of confluence and was greater at confluence. EGFR association with SHC was primarily soluble at subconfluence, while at confluence EGFR association was markedly increased and predominantly in the CSK. Thus, cell density regulates the CSK association of the EGFR and its ability to associate and activate signaling pathways in intestinal cells.

Adaptor Proteins, Signal Transducing↗
PubMed2005

Chloride channels in the small intestinal cell line IEC-18.

Small intestinal crypt cells play a critical role in modulating Cl- secretion during digestion. The types of Cl- channels mediating Cl- secretion in the small intestine was investigated using the intestinal epithelial cell line, IEC-18, which was derived from rat small intestine crypt cells. In initial radioisotope efflux studies, exposure to forskolin, ionomycin or a decrease in extracellular osmolarity significantly increased 36Cl efflux as compared to control cells. Whole cell patch clamp techniques were subsequently used to examine in more detail the swelling-, Ca2+-, and cAMP-activated Cl- conductance. Decreasing the extracellular osmolarity from 290 to 200 mOsm activated a large outwardly rectifying Cl- current that was voltage-independent and had an anion selectivity of I- > Cl-. Increasing cytosolic Ca2+ by ionomycin activated whole cell Cl- currents, which were also outwardly rectifying but were voltage-dependent. The increase in intracellular Ca2+ levels with ionomycin was confirmed with fura-2 loaded IEC-18 cells. A third type of whole cell Cl- current was observed after increases in intracellular cAMP induced by forskolin. These cAMP-activated Cl- currents have properties consistent with cystic fibrosis transmembrane regulator (CFTR) Cl- channels, as the currents were blocked by glibenclamide or NPPB but insensitive to DIDS. In addition, the current-voltage relationship was linear and had an anion selectivity of Cl- > I-. Confocal immunofluorescence studies and Western blots with two different anti-CFTR antibodies confirmed the expression of CFTR. These results suggest that small intestinal crypt cells express multiple types of Cl- channels, which may all contribute to net Cl- secretion.

Animals↗
PubMed1997

Induction of CYP1A1 by beta-naphthoflavone in IEC-18 rat intestinal epithelial cells and potentiation of induction by dibutyryl cAMP.

We have examined the inducibility of CYP1A1 by beta-naphthoflavone (BNF) in a rat intestinal epithelial cell line, IEC-18, and the associated interaction between cAMP and BNF. CYP1A1 was not constitutively expressed in IEC-18 cells. Upon treatment with BNF, CYP1A1 RNA, protein, and microsomal 7-ethoxyresorufin O-deethylase activity were detected. Treatment with dibutyryl cAMP resulted in a 2-fold increase in the extent of induction at both RNA and protein levels, with corresponding increases in CYP1A1 enzymatic activity. These results support the involvement of protein kinase A in Ah receptor-mediated induction of CYP1A1 and provide an in vitro model for further studies on the mechanisms underlying regional and cellular differences in the regulation of CYP1A1 gene expression in the small intestine.

Animals↗
PubMed1998

Effect of laminin on the nuclear localization of nucleolin in rat intestinal epithelial IEC-6 cells.

Laminin is a major component of extracellular matrix. The mechanism of action of laminin on cell proliferation, differentiation, and migration is not fully understood. In this study, we investigated the role of extracellular matrix, especially laminin, on the cellular localization of the nuclear protein, nucleolin, and on cell proliferation. Immunofluorescent and western blot analysis indicated that nucleolin was translocated most efficiently to the nucleus in the small intestinal rat epithelial cell line (IEC-6) when cultured on laminin-coated plates. Specifically, nucleolin was observed predominantly in cytoplasm in the cells cultured without laminin. In contrast, nuclear localization was observed in the cells cultured on laminin. This effect of laminin on nucleolin translocation was time-dependent. Laminin was also observed to stimulate proliferation of IEC-6 cells in serum free medium. Our results suggest that laminin alters the distribution of nucleolin which may be an early signal for cell proliferation.

Animals↗
PubMed1995

IL-1 alpha is constitutively expressed in the rat intestinal epithelial cell line IEC-6.

The rat small intestinal epithelial cell line, IEC-6, was examined for the presence of IL-1 mRNAs using the reverse transcription/polymerase chain reaction method. IL-1 alpha but not IL-1 beta transcripts were detected in plastic adherent cells. The levels of IL-1 alpha transcripts were similar in cells cultured at different densities. IL-1 activity was not detected, by bioassay, in the culture supernatant of the cells, nor was it membrane associated. IL-1 activity was detected in cell lysates, although its measurement was made difficult by the presence of an inhibitor of the bioassay. Intracellular IL-1 alpha was detectable using Western blots. Interleukin-1 receptor antagonist mRNA was also detectable in plastic adherent cells. Treatment with TGF-beta 1, IL-1 beta, IL-6, TNF-alpha, or combinations of any two of these cytokines failed to induce the secretion of the IL-1 alpha or to significantly change the levels of mRNA. The IEC-6 cell line is similar to other epithelial cells in having intracellular pools of IL-1 alpha.

Animals↗
PubMed1993

Paraquat uptake in the cultured gastrointestinal epithelial cell line, IEC-6.

Although the major target organ of paraquat toxicity is the lung, the primary route of absorption of the herbicide is across the gastrointestinal epithelium. Thus, uptake of paraquat was investigated in the rat duodenal crypt cell line, IEC-6. The herbicide was toxic; as measured by cell number, the LD50 was 75 microM. Using 2 microM [14C]paraquat as tracer, uptake of the herbicide was found to be slow but linear over 24 hr at 37 degrees C. No accumulation was observed at 4 degrees C. Inhibition studies showed paraquat inhibited both putrescine and spermidine uptake after only a 15-min incubation. The Km for putrescine was increased when incubated in the presence of paraquat whereas Vmax was not changed, suggesting a competitive mode of inhibition. In the reverse situation, putrescine also inhibited paraquat uptake (IC50 10 microM). Paraquat uptake was reduced in a dose-dependent manner by the putative calmodulin antagonist W-7, but was not affected by KN-62, a Ca2+/calmodulin kinase II specific inhibitor. These results provide evidence that paraquat uptake occurs through the polyamine transport system in IEC-6 cells. Furthermore, this process is modulated by intracellular events involving a major signaling protein, Ca2+/calmodulin.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1996

Regulation of complement C3 synthesis by interleukin-1 and transforming growth factor-beta in rat non-transformed intestinal epithelial cell line, IEC-6.

Intestinal epithelial cells are an important source of many biologically active molecules that modulate immune responses in the mucosa. The purpose of this study was to demonstrate the synthesis of complement C3 component in the rat non-transformed crypt-like intestinal epithelial cell line, IEC-6. Unstimulated IEC-6 cells secreted a low level of C3 protein and showed weak expression of C3 mRNA. The addition of interleukin (IL)-1 beta induced a dose- and time-dependent increase in C3 production. These effects of IL-1 beta were observed at a concentration as low as 0.01 ng/ml and reached a plateau at a concentration of 5 ng/ml. The effects were observed at the mRNA level as early as 6 h after the beginning of incubation. Transforming growth factor (TGF)-beta alone had no effect. However, TGF-beta at low concentrations (0.001-1 ng/ml) enhanced the effect of IL-1 beta in increasing C3 production; this enhancement was not observed at high concentrations (5-10 ng/ml). These effects of TGF-beta were also observed at the mRNA level. The present findings indicate that intestinal epithelial cells are indeed capable of synthesizing complement C3 in response to IL-1 beta and TGF-beta.

Animals↗
PubMed2005

Augmented internalisation of ferroportin to late endosomes impairs iron uptake by enterocyte-like IEC-6 cells.

Absorption of iron occurs by duodenal enterocytes, involving uptake by the divalent metal transporter-1 (DMT1) and release by ferroportin. Ferroportin responds to the hepatocyte-produced 25-amino-acid-peptide hepcidin-25 by undergoing internalisation to late endosomes that impair iron release. Ferroportin is also expressed on the apical membrane of polarised Caco-2 cells, rat intestinal cells and in IEC-6 cells (an intestinal epithelial cell line). A blocking antibody to ferroportin also impairs the uptake, but not the release, of iron. In this study IEC-6 cells were used to study the mechanism of impairment or recovery from impairment produced by the blocking antibody and the fate of DMT1 and ferroportin. Uptake of 1 muM Fe(II) was studied by adding the antibody from time 0 and after adding or removing the antibody once a steady state had been reached. Surface binding, maximum iron transport rate V(max) and transporter affinity (K(m)) were measured after impairment of iron uptake. Ferroportin and DMT1 distribution were assessed by immunofluorescence microscopy. Antibody-mediated impairment, or recovery from impairment, of Fe(II) uptake occurred within minutes. Impairment was lost when the antibody was combined with the immunizing peptide. DMT1 and ferroportin undergo internalisation to late endosomes and, in the presence of the antibody, augmented internalisation of DMT1 and ferroportin caused swelling of late endosomes. Surface binding of Fe(II) and iron transport V(max) were reduced by 50%, indicating that the antibody removed membrane-bound DMT1. The ferroportin antibody induced rapid turnover of membrane ferroportin and DMT1 and its internalisation to late endosomes, resulting in impaired Fe(II) uptake.

Animals↗
PubMed1989

Differentiation of intestinal epithelial cell line (IEC-18) by an acid extract of rat small intestine.

A factor which may induce differentiation of intestinal epithelial cell lines in vitro was found in an acid extract of adult rat small intestine. The addition of a partially purified acetic acid extract of rat small intestine to IEC-18 cell culture dishes increased sucrase activity within 48 h. Thymidine incorporation markedly decreased within 24 h. Significant development of microvilli-like structures was observed on the acid extract-treated IEC-18 cells, compared with controls. This activity of rat acid extract was heat-stable and the apparent molecular weight of the factor was 400-800. These findings suggested that the factor may be related to the epithelial differentiation of rat small intestinal crypt cells.

Animals↗
PubMed2005

IEC 61511 and the capital project process--a protective management system approach.

This year, the process industry has reached an important milestone in process safety-the acceptance of an internationally recognized standard for safety instrumented systems (SIS). This standard, IEC 61511, documents good engineering practice for the assessment, design, operation, maintenance, and management of SISs. The foundation of the standard is established by several requirements in Part 1, Clauses 5-7, which cover the development of a management system aimed at ensuring that functional safety is achieved. The management system includes a quality assurance process for the entire SIS lifecycle, requiring the development of procedures, identification of resources and acquisition of tools. For maximum benefit, the deliverables and quality control checks required by the standard should be integrated into the capital project process, addressing safety, environmental, plant productivity, and asset protection. Industry has become inundated with a multitude of programs focusing on safety, quality, and cost performance. This paper introduces a protective management system, which builds upon the work process identified in IEC 61511. Typical capital project phases are integrated with the management system to yield one comprehensive program to efficiently manage process risk. Finally, the paper highlights areas where internal practices or guidelines should be developed to improve program performance and cost effectiveness.

Accident Prevention↗
PubMed2005

Identification and characterization of 1,25D3-membrane-associated rapid response, steroid (1,25D3-MARRS)-binding protein in rat IEC-6 cells.

We report the presence of a mammalian equivalent of the avian Membrane-Associated Rapid Response, Steroid (1,25D3-MARRS)-binding protein specific for 1,25(OH)2D3 in a rat small intestinal cell line, IEC-6, that demonstrates rapid responses to the steroid hormone. Identification of transcript and protein was achieved using RT-PCR with several specific primer sets, Western blot analysis with two separate antibodies recognizing distinct regions of the protein, ribozyme knockdown and immunohistochemistry. Promoter analysis of the 1000-bp upstream region of the 1,25D3-MARRS gene in several species revealed the presence of a conserved smad-3 element in the 5' proximal promoter region, but no classical vitamin D response element (VDRE). Treatment of IEC-6 cells with transforming growth factor beta1 (TGFbeta1) increased steady-state levels of 1,25D3-MARRS (mRNA and protein) approximately two-fold over a 24-h period. In contrast, treatment with 1,25(OH)2D3 failed to significantly change 1,25D3-MARRS protein or mRNA levels. Localization studies showed rapid nuclear translocation of a pool of 1,25D3-MARRS protein after 1,25(OH)2D3 treatment, suggesting that the protein is subject to membrane-initiated signal pathway activation. Together these data point to complex interactions between the two important 1,25(OH)2D3 sensitive response systems in intestinal cells, 1,25D3-MARRS protein and the well-studied nVDR, that together work to fine tune intestinal Ca2+ absorption in a variety of avian and mammalian species.

Animals↗
PubMed2004

Function block applications in control systems based on IEC 61804.

A variety of fieldbus technologies and digital fieldbus devices have been introduced within the process industries over the last ten years. There has been a gradual acceptance of the fact that a variety of communication technologies are needed to fully address the application requirements of a manufacturing facility. However, engineers responsible for the specification, engineering, and implementation of control systems require that a common interface and functionality be provided in the control system. This capability should be independent of the underlying fieldbus technology or manufacturer of the fieldbus device. The draft IEC 61804 standard defines how a control system can be structured to provide this flexibility in the utilization of fieldbus technology. In this paper, we discuss how a consistent function block capability may be provided for all fieldbus technology utilized in a control system. Examples will be given of how this standard has been applied in modern control systems to give a consistent interface to Foundation Fieldbus and PROFIBUS. Some detail will be presented on the standard means that is defined for manufacturers to describe function block capability of a field device. An analysis is given of the impact and benefit that the IEC 61804 standard will have on the process industry and on manufacturers of control systems.

Journal Article↗
PubMed1994

Transforming growth factor-alpha increases tyrosine phosphorylation of microtubule-associated protein kinase in a small intestinal crypt cell line (IEC-6).

The small intestinal crypt cell line (IEC-6) is an undifferentiated, untransformed, mitotically active cell used in this study to determine the effect of transforming growth factor-alpha (TGF-alpha) on tyrosine phosphorylation levels of cellular proteins. Thymidine incorporation increased maximally after addition of 2 ng/ml TGF-alpha for 24 h. At the same dose, TGF-alpha induced the tyrosine phosphorylation of proteins with approximate molecular masses of 42, 44, 52, 80, 150 and 175 kDa as shown by Western blots treated with anti-phosphotyrosine antibody. The most intense phosphorylation was seen in the 42 kDa (p42) and 44 kDa (p44) proteins, which were identified as two isoforms of microtubule-associated protein kinase (MAPK). This phosphorylation was seen as early as 5 min post stimulation and was dose dependent. Both p42 and p44 were found in the nucleus after stimulation, although a basal level of unphosphorylated protein was present before stimulation. The observed tyrosine phosphorylation of p42 and p44 was inhibited by genistein, a tyrosine kinase inhibitor, and tyrphostin 23, an epidermal growth factor receptor tyrosine kinase inhibitor. We conclude that MAPK is tyrosine phosphorylated in response to TGF-alpha stimulation of IEC-6 cells.

Animals↗