Development of a Sensitive Bioassay for the Analysis of IGF-Related Activation of AKT/mTOR Signaling in Biological Matrices.
AKT phosphorylation
BIRA assay
IGF
Western immunoblotting
bioactivity
bioassay
capillary immuno-electrophoresis
cerebrospinal fluid
colostrum
mTOR
milk
serum
Journal
Cells
ISSN: 2073-4409
Titre abrégé: Cells
Pays: Switzerland
ID NLM: 101600052
Informations de publication
Date de publication:
24 02 2021
24 02 2021
Historique:
received:
24
01
2021
revised:
15
02
2021
accepted:
21
02
2021
entrez:
6
3
2021
pubmed:
7
3
2021
medline:
16
11
2021
Statut:
epublish
Résumé
The bioactivity of the IGF system is not a function of isolated hormone concentrations in a given biological matrix. Instead, the biological activities of IGFs are regulated by IGFBPs, IGFBP proteases, and inhibitors of IGFBP proteases. Therefore, assays based on IGF-related bioactivity may describe functions of the complete IGF system in a given biological matrix. Of particular interest are the IGF system effects on the AKT/mTOR pathway, as a dominant system for controlling growth, metabolism, and aging. In order to improve the sensitivity of IGF-dependent bioactivity, we made use of the known short-term and enhancing effects of IGFBP2 on the intracellular PI3K pathway. As a specific readout of this pathway, and further as a marker of the mTOR pathway, we assessed the phosphorylation of AKT-Ser473. Preincubation using IGFBP2 enhanced IGF1-dependent AKT-Ser473 phosphorylation in our experimental system. The assay's specificity was demonstrated by inhibition of IGF1 receptors outside or inside the cell, using antiserum or small molecule inhibitors, which reduced AKT phosphorylation in response to exogenous IGF1 (
Identifiants
pubmed: 33668197
pii: cells10030482
doi: 10.3390/cells10030482
pmc: PMC7995968
pii:
doi:
Substances chimiques
IGF1 protein, human
0
Insulin-Like Growth Factor I
67763-96-6
MTOR protein, human
EC 2.7.1.1
Proto-Oncogene Proteins c-akt
EC 2.7.11.1
TOR Serine-Threonine Kinases
EC 2.7.11.1
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Subventions
Organisme : H. Wilhelm Schaumann Stiftung
ID : no grant number
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