Search PubMed⌕ Search
Home/Archive/PubMed

PubMed · 10457347

Caco-2 cell differentiation is associated with a decrease in stat protein levels and binding.

S Wang·B M Evers
View original source ↗Export metadata ↓

Abstract

Novel proteins of the Stat (signal transducers and activators of transcription) family have been associated with proliferation and differentiation of certain cells; the role of these transcription factors in gut differentiation has not been examined. The purpose of this study was to determine whether the cellular levels and actual binding of the Stat proteins are altered with intestinal differentiation using the Caco-2 cell line that spontaneously differentiates to a small bowel phenotype after confluency. We found that both Stat3 and Stat5 protein levels were increased in preconfluent and confluent Caco-2 cells; levels then decreased with postconfluency. Mobility shift assays demonstrated maximal binding of Stat3 and Stat5 at confluency and, similar to protein levels, binding activity decreased with postconfluency. The intestinal differentiation marker gene sucrase-isomaltase was increased by postconfluent day 1 with maximal levels by day 6. The progressive decrease of Stat3 and Stat5 protein levels and binding activity, occurring at a time associated with increased Caco-2 cell differentiation, suggests that a decrease in the cellular levels of these proteins may potentially play a role in subsequent intestinal cell differentiation. Delineating the cellular mechanisms responsible for intestinal differentiation is crucial to a better understanding of both normal gut development and aberrant gut growth.

Explore related subjects

Blotting, NorthernBlotting, WesternCaco-2 CellsCell DifferentiationDNA ProbesDNA-Binding ProteinsElectrophoresis, Polyacrylamide GelGene Expression Regulation, EnzymologicGene Expression Regulation, NeoplasticHumansMilk ProteinsProtein BindingRNASTAT1 Transcription FactorSTAT3 Transcription FactorSTAT5 Transcription FactorSignal TransductionSucrase-Isomaltase ComplexTrans-ActivatorsJournal ArticleResearch Support, Non-U.S. Gov'tResearch Support, U.S. Gov't, P.H.S.

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

S Wang, B M Evers. Caco-2 cell differentiation is associated with a decrease in stat protein levels and binding.. https://doi.org/10.1016/s1091-255x(99)80034-1

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related citations

PubMed2025

Northern Blotting: Protocols for Radioactive and Nonradioactive Detection of RNA.

Northern blotting is a common technique in RNA biology, allowing to detect and quantify RNAs of interest following separation by gel electrophoresis, transfer to a membrane, and hybridization of specific anti-complementary labelled probes. In this chapter, we describe our protocol for efficient RNA extraction from yeast, separation on agarose gel, and capillary transfer to a membrane. We provide two different methods for strand-specific detection of several types of RNAs using oligonucleotide probes, the first using radioactive 32P-labelled probes, the second based on nonradioactive digoxigenin-labelled probes.

Blotting, Northern↗
PubMed2001

Differential gene expression in Thermotoga neapolitana in response to growth substrate.

We have previously shown that beta-galactosidase activity expressed in Thermotoga neapolitana cells grown on lactose is subject to repression by glucose when they are grown on both substrates whereas beta-galactosidase and beta-glucosidase activities observed in cells grown on cellobiose are not repressed by growth on both glucose and cellobiose. To examine the differential expression of bgalA, bgalB, bglA and bglB in T. neapolitana, total RNA was isolated from cells growing on either glucose, lactose or cellobiose as the sole source of carbon and transcripts encoding these genes were quantitated by Northern blot analyses. BglA expression was induced by cellobiose while bglB was expressed under all three conditions at a lower level. Expression of the beta-galactosidase genes, bgalA and bgalB, was detected only in lactose-grown cells. beta-Glucosidase enzyme activity was only found in cell extracts of cellobiose-grown cells while beta-galactosidase activity was found in both lactose- and cellobiose-grown cell extracts. Our results show that in cellobiose-grown cells, the high beta-glucosidase activity is likely due to expression of bglA and that neither bgalA nor bgalB is responsible for the beta-galactosidase activity.

Blotting, Northern↗
PubMed2001

Association of thrombospondin-1 and cardiac allograft vasculopathy in human cardiac allografts.

BACKGROUND: Despite the expression of angiogenic growth factors in transplanted hearts, neovessel formation appears scant. We therefore hypothesized that cardiac allografts contain endogenous inhibitors of angiogenesis. In particular, we tested the involvement in cardiac allografts of thrombospondin-1 (TSP-1), a matrix glycoprotein that inhibits angiogenesis and facilitates smooth muscle cell (SMC) proliferation. METHODS AND RESULTS: Levels of TSP-1 mRNA in endomyocardial biopsy samples of human cardiac allografts substantially exceeded those in normal hearts. The ratio of TSP to GAPDH mRNA determined with quantitative RT-PCR was 6.54+/-1.6 in cardiac allografts versus 0.26+/-0.02 (P:=0.001) in normal hearts. Analysis in sequential biopsies revealed a strong association between persistent elevation of TSP-1 in allografts and the development of cardiac allograft vasculopathy (CAV). The CAV score was 2.4+/-0.8 in patients with persistent TSP-1 elevation compared with 0.2+/-0.2 in patients without elevation (P:=0.001). Immunohistochemistry demonstrated intense expression of TSP-1 in cardiac allografts, predominantly in cardiac myocytes and neointimal SMCs. In vitro experiments demonstrated that T cells expressed TSP-1, acidic fibroblast growth factor, and vascular endothelial cell growth factor on allogeneic stimulation. Cytokines known to be elevated in cardiac allografts (interleukin-1beta, interferon-gamma, and tumor necrosis factor-alpha) induced TSP-1 in SMCs but inhibited TSP-1 in endothelial cells. CONCLUSIONS: Persistent elevation of TSP-1 in cardiac allografts correlates with the development of CAV. Allogeneic stimulation induces angiogenic growth factors and TSP-1 in T cells. Cytokines differentially regulate TSP-1 expression in SMCs versus endothelial cells. Increased levels of TSP-1 in human cardiac allografts may alter vascular responses to angiogenic growth factors by inhibiting angiogenesis and promoting SMC proliferation characteristic of CAV.

Blotting, Northern↗