An effective method for the separation of insulin-like growth factors 1 and 2 during the purification process

1988 ◽  
Vol 8 (3) ◽  
pp. 293-297 ◽  
Author(s):  
M. Tally ◽  
K. Florell ◽  
G. Enberg
Keyword(s):  
Growth Factors ◽  
Human Insulin ◽  
Cation Exchanger ◽  
Radioreceptor Assay ◽  
Cross Reaction ◽  
Purification Process ◽  
Purification Step ◽  
Igf Receptor ◽  
Insulin Like Growth Factors

The separation of human insulin-like growth factors hIGF-1 and hIGF-2 was greatly improved by an additional purification step using the cation exchanger Mono-S (FPLC) compared to previous studies. Cross-reactions between hIFG-1 and hIGF-2 were strongly reduced. The more highly purified hIGF-1 had a cross-reaction of less than 1% in the RIA for hIGF-2, and was equivalent to recombinant hIGF-1. The pure hIGF-2 had a cross-reaction of less than 1% in the RIA for hIGF-1. In the human placental hIGF-2 radioreceptor assay, the hIGF-1 polypeptide competed less than 1% with hIGF-2 when the type 1 IGF receptor was blocked with insulin.

scholarly journals Binding properties and biological potencies of insulin-like growth factors in L6 myoblasts

1986 ◽  
Vol 233 (1) ◽  
pp. 223-230 ◽  
Author(s):  
F J Ballard ◽  
L C Read ◽  
G L Francis ◽  
C J Bagley ◽  
J C Wallace
Keyword(s):  
Protein Synthesis ◽  
Growth Factors ◽  
Radioligand Binding ◽  
Cross Linking ◽  
Protein Accumulation ◽  
Igf Receptor ◽  
H Protein ◽  
Insulin Like Growth Factors

Protein synthesis in rat L6 myoblasts is stimulated and protein breakdown inhibited in a co-ordinate manner by insulin-like growth factors (IGF) or insulin. For both processes, bovine IGF-1 was somewhat more potent than human IGF-1, which was effective at a tenth the concentration of insulin, rat IGF-2 or human IGF-2. A similar order of potency is noted when DNA synthesis or protein accumulation is monitored over a 24 h period, but between 20- and 50-fold higher concentrations of each growth factor are required than those needed to produce effects in the 4 h protein-synthesis or -breakdown measurements. Binding experiments with labelled human or bovine IGF-1 as ligand demonstrated competition at concentrations of IGF-2, especially human IGF-2, lower than that of either IGF-1 preparation. This pattern was much more pronounced when the radioligand was either human IGF-2 or rat IGF-2. Insulin competed 10-15% for the binding of labelled IGF-1, but not at all with labelled IGF-2. Ligand-receptor cross-linking experiments showed that labelled bovine IGF-1 bound approximately equally to the type 1 IGF receptor (Mr 130000 after reduction) and to the type 2 IGF receptor (Mr 270000 after reduction), and that unlabelled IGF-1 competed equally with radioligand binding to both receptors. On the other hand, rat IGF-2 competed more effectively for binding to the type-2 receptor, and insulin competed only for binding to the type-1 receptor. Further cross-linking experiments with rat IGF-2 as radioligand demonstrated binding only to the type-2 receptor and to proteins with Mr values after reduction of 230000 and 200000. This binding was prevented by high rat IGF-2 concentrations, less effectively by bovine IGF-1 and not at all by insulin. The apparently conflicting biological potencies and receptor binding of the different growth factors can be explained if all the biological actions are mediated via the type-1 IGF receptor, rather than through the abundant type-2 receptor.

Expression of mRNA encoding insulin-like growth factors I and II and the type 1 IGF receptor in the bovine corpus luteum at defined stages of the oestrous cycle

Reproduction ◽  
2000 ◽  
pp. 293-302 ◽  
Author(s):  
KJ Woad ◽  
G Baxter ◽  
CO Hogg ◽  
TA Bramley ◽  
R Webb ◽  
...  
Keyword(s):  
Growth Factors ◽  
Corpus Luteum ◽  
Luteal Phase ◽  
Oestrous Cycle ◽  
Luteal Cells ◽  
Bovine Corpus Luteum ◽  
Igf I ◽  
Igf Receptor ◽  
Insulin Like Growth Factors

Previous studies have implicated insulin-like growth factors I and II (IGF-I and -II), in the regulation of ovarian function. The present study investigated the localization of mRNA encoding IGF-I and -II and the type 1 IGF receptor using in situ hybridization to determine further the roles of the IGFs within the bovine corpus luteum at precise stages of the oestrous cycle. Luteal expression of mRNA encoding IGF-I and -II and the type 1 IGF receptor was detected throughout the oestrous cycle. The expression of IGF-I mRNAvaried significantly during the oestrous cycle. IGF-I mRNA concentrations were significantly higher on day 15 than on day 10, and IGF-I mRNA in the regressing corpus luteum at 48 h after administration of exogenous prostaglandin was significantly greater than in the early or mid-luteal phase (days 5 and 10). In contrast, there was no significant effect of day of the oestrous cycle on expression of mRNA for IGF-II and the type 1 IGF receptor in the corpus luteum. Expression of IGF-II mRNA was localized to a subset of steroidogenic luteal cells and was also associated with cells of the luteal vasculature. mRNA encoding the type 1 IGF receptor was widely expressed in a pattern indicative of expression in large and small luteal cells. These data demonstrate that the bovine corpus luteum is a site of IGF production and reception throughout the luteal phase. Furthermore, this study highlights the potential of IGF-II in addition to IGF-I in the autocrine and paracrine regulation of luteal function.

scholarly journals The C Region of Human Insulin-like Growth Factor (IGF) I Is Required for High Affinity Binding to the Type 1 IGF Receptor

1989 ◽  
Vol 264 (19) ◽  
pp. 11004-11008 ◽  
Author(s):  
M L Bayne ◽  
J Applebaum ◽  
D Underwood ◽  
G G Chicchi ◽  
B G Green ◽  
...  
Keyword(s):  
Growth Factor ◽  
Human Insulin ◽  
Affinity Binding ◽  
Insulin Like Growth Factor ◽  
High Affinity Binding ◽  
High Affinity ◽  
Igf I ◽  
Igf Receptor

scholarly journals Comparative binding of bovine, human and rat insulin-like growth factors to membrane receptors and to antibodies against human insulin-like growth factor-1

1986 ◽  
Vol 233 (1) ◽  
pp. 215-221 ◽  
Author(s):  
L C Read ◽  
F J Ballard ◽  
G L Francis ◽  
R C Baxter ◽  
C J Bagley ◽  
...  
Keyword(s):  
Growth Factors ◽  
Polyclonal Antiserum ◽  
Competitive Binding ◽  
Membrane Receptors ◽  
Binding Studies ◽  
Comparative Binding ◽  
Placental Membranes ◽  
Insulin Like Growth Factors

The immunological properties of human, bovine and rat insulin-like growth factors (IGF) and insulin were compared in competitive binding studies with Tr10 and NPA polyclonal antisera raised in rabbits against human IGF-1. Bovine IGF-1 was 11-19% as effective as human IGF-1 in competing for binding with 125I-labelled human IGF-1, whereas IGF-2 reacted poorly and insulin did not compete. Similar competitive binding curves were obtained with the mouse monoclonal anti-(human IGF-1) antibody 3D1, except that bovine IGF-1 showed a severalfold greater affinity for the monoclonal antibody than for either polyclonal antiserum. Membranes isolated from human placenta, sheep placenta and foetal-human liver were used as sources of cellular receptors. In human placental membranes, most of the binding of IGF-1 tracers could be attributed to a type-1 receptor, because insulin inhibited up to 65% of tracer binding. The other two tissues apparently contain only type-2 receptors, as evidenced by the very low potency of bovine or human IGF-1 in competing for binding with IGF-2 tracers and the absence of any competition by insulin. In competition for binding with labelled bovine or human IGF-1 to human placental membranes, bovine IGF-1 had a similar potency to human IGF-1, whereas bovine IGF-1 was more potent in binding studies with tissues rich in type-2 receptors. Rat IGF-2 was considerably less effective than human IGF-2 in competition for receptors on any of the membrane preparations.

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