Identification of specific sites in the TNF-α molecule promoting insulin resistance in H-411E cells

S. K. Mishra, S. S. Solomon, C. S. Cwik, M. R. Palazzolo, Arnold Postlethwaite, J. M. Seyer

Research output: Contribution to journalArticle

Abstract

Data from a number of laboratories supports a potential role for Tumor Necrosis Factor-α (TNF-α) in the pathogenesis of insulin resistance (IR) in animal models of/and human disease. We designed experiments to establish a dose-response relationship for TNF-α and IR in H-411E cells in culture. IR was measured by the ability of graded amounts of insulin to stimulate expression of calmodulin (CaM) mRNA in these cells. This was assessed by autoradiographs of Northern blot(s) of CaM mRNA probed with labelled oligonucleotide cDNA for rat CaM I. We found that TNF-α at 0.1, 1.0, and 10.0ng/ml opposed 10,000μ U/ml insulin, i.e. % IR: 20%; 67%; and 88%, respectively, At 1.0 ng/ml TNF-α, insulin 1,000 μU/ml stimulated CaM mRNA 41% and at 10,000 μU/ml, 63%. Furthermore, oligopeptide homologues (at 1000 × M concentration of TNF-α) IV, 69-100 and VII, 133-157 conferred 66% and 101% IR, respectively, while all other peptide fragments were essentially without effect. Studies done with both monoclonal and polyclonal antibody to the TNF-α receptor demonstrated blocking activity of polyclonal, but not monoclonal antibody. This supports the concept that activity of the peptide fragments occurs through the TNF-α receptor and not by non-specific translocation across the plasma membrane. These data demonstrate the probable molecular site for IR, induced by TNF-α and its peptide fragments.

Original languageEnglish (US)
JournalJournal of Investigative Medicine
Volume44
Issue number1
StatePublished - Jan 1 1996

Fingerprint

Insulin Resistance
Tumor Necrosis Factor-alpha
Calmodulin
Insulin
Peptide Fragments
Molecules
Tumor Necrosis Factor Receptors
Messenger RNA
Monoclonal Antibodies
Animal Disease Models
Oligopeptides
Oligonucleotides
Northern Blotting
Complementary DNA
Cell Culture Techniques
Cell Membrane
Cell membranes
Rats
Animals
Antibodies

All Science Journal Classification (ASJC) codes

  • Medicine(all)
  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Identification of specific sites in the TNF-α molecule promoting insulin resistance in H-411E cells. / Mishra, S. K.; Solomon, S. S.; Cwik, C. S.; Palazzolo, M. R.; Postlethwaite, Arnold; Seyer, J. M.

In: Journal of Investigative Medicine, Vol. 44, No. 1, 01.01.1996.

Research output: Contribution to journalArticle

Mishra, S. K. ; Solomon, S. S. ; Cwik, C. S. ; Palazzolo, M. R. ; Postlethwaite, Arnold ; Seyer, J. M. / Identification of specific sites in the TNF-α molecule promoting insulin resistance in H-411E cells. In: Journal of Investigative Medicine. 1996 ; Vol. 44, No. 1.
@article{8058d2cc24d349d2b29d63e5ca531cbb,
title = "Identification of specific sites in the TNF-α molecule promoting insulin resistance in H-411E cells",
abstract = "Data from a number of laboratories supports a potential role for Tumor Necrosis Factor-α (TNF-α) in the pathogenesis of insulin resistance (IR) in animal models of/and human disease. We designed experiments to establish a dose-response relationship for TNF-α and IR in H-411E cells in culture. IR was measured by the ability of graded amounts of insulin to stimulate expression of calmodulin (CaM) mRNA in these cells. This was assessed by autoradiographs of Northern blot(s) of CaM mRNA probed with labelled oligonucleotide cDNA for rat CaM I. We found that TNF-α at 0.1, 1.0, and 10.0ng/ml opposed 10,000μ U/ml insulin, i.e. {\%} IR: 20{\%}; 67{\%}; and 88{\%}, respectively, At 1.0 ng/ml TNF-α, insulin 1,000 μU/ml stimulated CaM mRNA 41{\%} and at 10,000 μU/ml, 63{\%}. Furthermore, oligopeptide homologues (at 1000 × M concentration of TNF-α) IV, 69-100 and VII, 133-157 conferred 66{\%} and 101{\%} IR, respectively, while all other peptide fragments were essentially without effect. Studies done with both monoclonal and polyclonal antibody to the TNF-α receptor demonstrated blocking activity of polyclonal, but not monoclonal antibody. This supports the concept that activity of the peptide fragments occurs through the TNF-α receptor and not by non-specific translocation across the plasma membrane. These data demonstrate the probable molecular site for IR, induced by TNF-α and its peptide fragments.",
author = "Mishra, {S. K.} and Solomon, {S. S.} and Cwik, {C. S.} and Palazzolo, {M. R.} and Arnold Postlethwaite and Seyer, {J. M.}",
year = "1996",
month = "1",
day = "1",
language = "English (US)",
volume = "44",
journal = "Journal of Investigative Medicine",
issn = "1081-5589",
publisher = "Lippincott Williams and Wilkins",
number = "1",

}

TY - JOUR

T1 - Identification of specific sites in the TNF-α molecule promoting insulin resistance in H-411E cells

AU - Mishra, S. K.

AU - Solomon, S. S.

AU - Cwik, C. S.

AU - Palazzolo, M. R.

AU - Postlethwaite, Arnold

AU - Seyer, J. M.

PY - 1996/1/1

Y1 - 1996/1/1

N2 - Data from a number of laboratories supports a potential role for Tumor Necrosis Factor-α (TNF-α) in the pathogenesis of insulin resistance (IR) in animal models of/and human disease. We designed experiments to establish a dose-response relationship for TNF-α and IR in H-411E cells in culture. IR was measured by the ability of graded amounts of insulin to stimulate expression of calmodulin (CaM) mRNA in these cells. This was assessed by autoradiographs of Northern blot(s) of CaM mRNA probed with labelled oligonucleotide cDNA for rat CaM I. We found that TNF-α at 0.1, 1.0, and 10.0ng/ml opposed 10,000μ U/ml insulin, i.e. % IR: 20%; 67%; and 88%, respectively, At 1.0 ng/ml TNF-α, insulin 1,000 μU/ml stimulated CaM mRNA 41% and at 10,000 μU/ml, 63%. Furthermore, oligopeptide homologues (at 1000 × M concentration of TNF-α) IV, 69-100 and VII, 133-157 conferred 66% and 101% IR, respectively, while all other peptide fragments were essentially without effect. Studies done with both monoclonal and polyclonal antibody to the TNF-α receptor demonstrated blocking activity of polyclonal, but not monoclonal antibody. This supports the concept that activity of the peptide fragments occurs through the TNF-α receptor and not by non-specific translocation across the plasma membrane. These data demonstrate the probable molecular site for IR, induced by TNF-α and its peptide fragments.

AB - Data from a number of laboratories supports a potential role for Tumor Necrosis Factor-α (TNF-α) in the pathogenesis of insulin resistance (IR) in animal models of/and human disease. We designed experiments to establish a dose-response relationship for TNF-α and IR in H-411E cells in culture. IR was measured by the ability of graded amounts of insulin to stimulate expression of calmodulin (CaM) mRNA in these cells. This was assessed by autoradiographs of Northern blot(s) of CaM mRNA probed with labelled oligonucleotide cDNA for rat CaM I. We found that TNF-α at 0.1, 1.0, and 10.0ng/ml opposed 10,000μ U/ml insulin, i.e. % IR: 20%; 67%; and 88%, respectively, At 1.0 ng/ml TNF-α, insulin 1,000 μU/ml stimulated CaM mRNA 41% and at 10,000 μU/ml, 63%. Furthermore, oligopeptide homologues (at 1000 × M concentration of TNF-α) IV, 69-100 and VII, 133-157 conferred 66% and 101% IR, respectively, while all other peptide fragments were essentially without effect. Studies done with both monoclonal and polyclonal antibody to the TNF-α receptor demonstrated blocking activity of polyclonal, but not monoclonal antibody. This supports the concept that activity of the peptide fragments occurs through the TNF-α receptor and not by non-specific translocation across the plasma membrane. These data demonstrate the probable molecular site for IR, induced by TNF-α and its peptide fragments.

UR - http://www.scopus.com/inward/record.url?scp=33749570608&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33749570608&partnerID=8YFLogxK

M3 - Article

VL - 44

JO - Journal of Investigative Medicine

JF - Journal of Investigative Medicine

SN - 1081-5589

IS - 1

ER -