Development and molecular characterization of HCT-116 cell lines resistant to the tumor promoter and multiple stress-inducer, deoxycholate

Cara L. Crowley-Weber, Claire M. Payne, Mary Gleason-Guzman, George S Watts, Bernard W Futscher, Caroline N. Waltmire, Cheray Crowley, Katerina Dvorakova, Carol Bernstein, Mary Craven, Harinder Garewal, Harris Bernstein

Research output: Contribution to journalArticle

69 Citations (Scopus)

Abstract

Evidence from live cell bioassays shows that the flat mucosa from patients with colon cancer exhibits resistance to bile salt-induced apoptosis. Three independent cell lines derived from the colonic epithelial cell line HCT-116 were selected for resistance to bile salt-induced apoptosis. These cell lines were developed as tissue culture models of apoptosis resistance. Selection was carried out for resistance to apoptosis induced by sodium deoxycholate (NaDOC), the bile salt found in highest concentrations in human fecal water. Cultures of HCT-116 cells were serially passaged in the presence of increasing concentrations of NaDOC. The resulting apoptosis resistant cells were able to grow at concentrations of NaDOC (0.5 mM) that cause apoptosis in a few hours in unselected HCT-116 cells. These cells were then analyzed for changes in gene expression. Observations from cDNA microarray, 2-D gel electrophoresis/MALDI-mass spectroscopy, and confocal microscopy of immunofluorescently stained preparations indicated underexpression or overexpression of numerous genes at either the protein or mRNA level. Genes that may play a role in apoptosis and early stage carcinogenesis have been identified as upregulated in these cell lines, including Grp78, Bcl-2, NF-κB50), NF-κB(p65), thioredoxin peroxidase (peroxiredoxin) 2, peroxiredoxin 4, maspin, guanylate cyclase activating protein-1, PKCζ, EGFR, Ras family members, PKA, PI(4,5)K, TRAF2 and BIRC1 (IAP protein). Under-expressed mRNAs included BNIP3, caspase-6, caspase-3 and serine protease 11. NF-κB was constitutively activated in all three resistant cell lines, and was responsible, in part, for the observed apoptosis resistance, determined using antisense oligonucleotide strategies. Molecular and cellular analyses of these resistant cell lines has suggested potential mechanisms by which apoptosis resistance may develop in the colonic epithelium in response to high concentrations of hydrophobic bile acids that are associated with a Western-style diet. These analyses provide the rationale for the development of hypothesis-driven intermediate biomarkers to assess colon cancer risk on an individual basis.

Original languageEnglish (US)
Pages (from-to)2063-2080
Number of pages18
JournalCarcinogenesis
Volume23
Issue number12
DOIs
StatePublished - Dec 1 2002

Fingerprint

HCT116 Cells
Deoxycholic Acid
Carcinogens
Apoptosis
Cell Line
Peroxiredoxins
Bile Acids and Salts
Colonic Neoplasms
Guanylate Cyclase-Activating Proteins
TNF Receptor-Associated Factor 2
Caspase 6
Messenger RNA
Antisense Oligonucleotides
Matrix-Assisted Laser Desorption-Ionization Mass Spectrometry
Electrophoresis, Gel, Two-Dimensional
Serine Proteases
Oligonucleotide Array Sequence Analysis
Confocal Microscopy
Caspase 3
Biological Assay

ASJC Scopus subject areas

  • Cancer Research

Cite this

Development and molecular characterization of HCT-116 cell lines resistant to the tumor promoter and multiple stress-inducer, deoxycholate. / Crowley-Weber, Cara L.; Payne, Claire M.; Gleason-Guzman, Mary; Watts, George S; Futscher, Bernard W; Waltmire, Caroline N.; Crowley, Cheray; Dvorakova, Katerina; Bernstein, Carol; Craven, Mary; Garewal, Harinder; Bernstein, Harris.

In: Carcinogenesis, Vol. 23, No. 12, 01.12.2002, p. 2063-2080.

Research output: Contribution to journalArticle

Crowley-Weber, CL, Payne, CM, Gleason-Guzman, M, Watts, GS, Futscher, BW, Waltmire, CN, Crowley, C, Dvorakova, K, Bernstein, C, Craven, M, Garewal, H & Bernstein, H 2002, 'Development and molecular characterization of HCT-116 cell lines resistant to the tumor promoter and multiple stress-inducer, deoxycholate', Carcinogenesis, vol. 23, no. 12, pp. 2063-2080. https://doi.org/10.1093/carcin/23.12.2063
Crowley-Weber, Cara L. ; Payne, Claire M. ; Gleason-Guzman, Mary ; Watts, George S ; Futscher, Bernard W ; Waltmire, Caroline N. ; Crowley, Cheray ; Dvorakova, Katerina ; Bernstein, Carol ; Craven, Mary ; Garewal, Harinder ; Bernstein, Harris. / Development and molecular characterization of HCT-116 cell lines resistant to the tumor promoter and multiple stress-inducer, deoxycholate. In: Carcinogenesis. 2002 ; Vol. 23, No. 12. pp. 2063-2080.
@article{6752b3e0c6e34bf982a2d94e7905f6cb,
title = "Development and molecular characterization of HCT-116 cell lines resistant to the tumor promoter and multiple stress-inducer, deoxycholate",
abstract = "Evidence from live cell bioassays shows that the flat mucosa from patients with colon cancer exhibits resistance to bile salt-induced apoptosis. Three independent cell lines derived from the colonic epithelial cell line HCT-116 were selected for resistance to bile salt-induced apoptosis. These cell lines were developed as tissue culture models of apoptosis resistance. Selection was carried out for resistance to apoptosis induced by sodium deoxycholate (NaDOC), the bile salt found in highest concentrations in human fecal water. Cultures of HCT-116 cells were serially passaged in the presence of increasing concentrations of NaDOC. The resulting apoptosis resistant cells were able to grow at concentrations of NaDOC (0.5 mM) that cause apoptosis in a few hours in unselected HCT-116 cells. These cells were then analyzed for changes in gene expression. Observations from cDNA microarray, 2-D gel electrophoresis/MALDI-mass spectroscopy, and confocal microscopy of immunofluorescently stained preparations indicated underexpression or overexpression of numerous genes at either the protein or mRNA level. Genes that may play a role in apoptosis and early stage carcinogenesis have been identified as upregulated in these cell lines, including Grp78, Bcl-2, NF-κB50), NF-κB(p65), thioredoxin peroxidase (peroxiredoxin) 2, peroxiredoxin 4, maspin, guanylate cyclase activating protein-1, PKCζ, EGFR, Ras family members, PKA, PI(4,5)K, TRAF2 and BIRC1 (IAP protein). Under-expressed mRNAs included BNIP3, caspase-6, caspase-3 and serine protease 11. NF-κB was constitutively activated in all three resistant cell lines, and was responsible, in part, for the observed apoptosis resistance, determined using antisense oligonucleotide strategies. Molecular and cellular analyses of these resistant cell lines has suggested potential mechanisms by which apoptosis resistance may develop in the colonic epithelium in response to high concentrations of hydrophobic bile acids that are associated with a Western-style diet. These analyses provide the rationale for the development of hypothesis-driven intermediate biomarkers to assess colon cancer risk on an individual basis.",
author = "Crowley-Weber, {Cara L.} and Payne, {Claire M.} and Mary Gleason-Guzman and Watts, {George S} and Futscher, {Bernard W} and Waltmire, {Caroline N.} and Cheray Crowley and Katerina Dvorakova and Carol Bernstein and Mary Craven and Harinder Garewal and Harris Bernstein",
year = "2002",
month = "12",
day = "1",
doi = "10.1093/carcin/23.12.2063",
language = "English (US)",
volume = "23",
pages = "2063--2080",
journal = "Carcinogenesis",
issn = "0143-3334",
publisher = "Oxford University Press",
number = "12",

}

TY - JOUR

T1 - Development and molecular characterization of HCT-116 cell lines resistant to the tumor promoter and multiple stress-inducer, deoxycholate

AU - Crowley-Weber, Cara L.

AU - Payne, Claire M.

AU - Gleason-Guzman, Mary

AU - Watts, George S

AU - Futscher, Bernard W

AU - Waltmire, Caroline N.

AU - Crowley, Cheray

AU - Dvorakova, Katerina

AU - Bernstein, Carol

AU - Craven, Mary

AU - Garewal, Harinder

AU - Bernstein, Harris

PY - 2002/12/1

Y1 - 2002/12/1

N2 - Evidence from live cell bioassays shows that the flat mucosa from patients with colon cancer exhibits resistance to bile salt-induced apoptosis. Three independent cell lines derived from the colonic epithelial cell line HCT-116 were selected for resistance to bile salt-induced apoptosis. These cell lines were developed as tissue culture models of apoptosis resistance. Selection was carried out for resistance to apoptosis induced by sodium deoxycholate (NaDOC), the bile salt found in highest concentrations in human fecal water. Cultures of HCT-116 cells were serially passaged in the presence of increasing concentrations of NaDOC. The resulting apoptosis resistant cells were able to grow at concentrations of NaDOC (0.5 mM) that cause apoptosis in a few hours in unselected HCT-116 cells. These cells were then analyzed for changes in gene expression. Observations from cDNA microarray, 2-D gel electrophoresis/MALDI-mass spectroscopy, and confocal microscopy of immunofluorescently stained preparations indicated underexpression or overexpression of numerous genes at either the protein or mRNA level. Genes that may play a role in apoptosis and early stage carcinogenesis have been identified as upregulated in these cell lines, including Grp78, Bcl-2, NF-κB50), NF-κB(p65), thioredoxin peroxidase (peroxiredoxin) 2, peroxiredoxin 4, maspin, guanylate cyclase activating protein-1, PKCζ, EGFR, Ras family members, PKA, PI(4,5)K, TRAF2 and BIRC1 (IAP protein). Under-expressed mRNAs included BNIP3, caspase-6, caspase-3 and serine protease 11. NF-κB was constitutively activated in all three resistant cell lines, and was responsible, in part, for the observed apoptosis resistance, determined using antisense oligonucleotide strategies. Molecular and cellular analyses of these resistant cell lines has suggested potential mechanisms by which apoptosis resistance may develop in the colonic epithelium in response to high concentrations of hydrophobic bile acids that are associated with a Western-style diet. These analyses provide the rationale for the development of hypothesis-driven intermediate biomarkers to assess colon cancer risk on an individual basis.

AB - Evidence from live cell bioassays shows that the flat mucosa from patients with colon cancer exhibits resistance to bile salt-induced apoptosis. Three independent cell lines derived from the colonic epithelial cell line HCT-116 were selected for resistance to bile salt-induced apoptosis. These cell lines were developed as tissue culture models of apoptosis resistance. Selection was carried out for resistance to apoptosis induced by sodium deoxycholate (NaDOC), the bile salt found in highest concentrations in human fecal water. Cultures of HCT-116 cells were serially passaged in the presence of increasing concentrations of NaDOC. The resulting apoptosis resistant cells were able to grow at concentrations of NaDOC (0.5 mM) that cause apoptosis in a few hours in unselected HCT-116 cells. These cells were then analyzed for changes in gene expression. Observations from cDNA microarray, 2-D gel electrophoresis/MALDI-mass spectroscopy, and confocal microscopy of immunofluorescently stained preparations indicated underexpression or overexpression of numerous genes at either the protein or mRNA level. Genes that may play a role in apoptosis and early stage carcinogenesis have been identified as upregulated in these cell lines, including Grp78, Bcl-2, NF-κB50), NF-κB(p65), thioredoxin peroxidase (peroxiredoxin) 2, peroxiredoxin 4, maspin, guanylate cyclase activating protein-1, PKCζ, EGFR, Ras family members, PKA, PI(4,5)K, TRAF2 and BIRC1 (IAP protein). Under-expressed mRNAs included BNIP3, caspase-6, caspase-3 and serine protease 11. NF-κB was constitutively activated in all three resistant cell lines, and was responsible, in part, for the observed apoptosis resistance, determined using antisense oligonucleotide strategies. Molecular and cellular analyses of these resistant cell lines has suggested potential mechanisms by which apoptosis resistance may develop in the colonic epithelium in response to high concentrations of hydrophobic bile acids that are associated with a Western-style diet. These analyses provide the rationale for the development of hypothesis-driven intermediate biomarkers to assess colon cancer risk on an individual basis.

UR - http://www.scopus.com/inward/record.url?scp=0036963898&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0036963898&partnerID=8YFLogxK

U2 - 10.1093/carcin/23.12.2063

DO - 10.1093/carcin/23.12.2063

M3 - Article

C2 - 12507930

AN - SCOPUS:0036963898

VL - 23

SP - 2063

EP - 2080

JO - Carcinogenesis

JF - Carcinogenesis

SN - 0143-3334

IS - 12

ER -