Characterization of a far-red analog of ghrelin for imaging GHS-R in P19-derived cardiomyocytes.
Peptides
; 54: 81-8, 2014 Apr.
Article
in En
| MEDLINE
| ID: mdl-24468548
Ghrelin and its receptor, the growth hormone secretagogue receptor (GHS-R), are expressed in the heart, and may function to promote cardiomyocyte survival, differentiation and contractility. Previously, we had generated a truncated analog of ghrelin conjugated to fluorescein isothiocyanate for the purposes of determining GHS-R expression in situ. We now report the generation and characterization of a far-red ghrelin analog, [Dpr(3)(octanoyl), Lys(19)(Cy5)]ghrelin (1-19), and show that it can be used to image changes in GHS-R in developing cardiomyocytes. We also generated the des-acyl analog, des-acyl [Lys(19)(Cy5)]ghrelin (1-19) and characterized its binding to mouse heart sections. Receptor binding affinity of Cy5-ghrelin as measured in HEK293 cells overexpressing GHS-R1a was within an order of magnitude of that of fluorescein-ghrelin and native human ghrelin, while the des-acyl Cy5-ghrelin did not bind GHS-R1a. Live cell imaging in HEK293/GHS-R1a cells showed cell surface labeling that was displaced by excess ghrelin. Interestingly, Cy5-ghrelin, but not the des-acyl analog, showed concentration-dependent binding in mouse heart tissue sections. We then used Cy5-ghrelin to track GHS-R expression in P19-derived cardiomyocytes. Live cell imaging at different time points after DMSO-induced differentiation showed that GHS-R expression preceded that of the differentiation marker aMHC and tracked with the contractility marker SERCA 2a. Our far-red analog of ghrelin adds to the tools we are developing to map GHS-R in developing and diseased cardiac tissues.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Peptide Fragments
/
Myocytes, Cardiac
/
Ghrelin
/
Receptors, Ghrelin
/
Molecular Imaging
Limits:
Animals
/
Female
/
Humans
/
Male
Language:
En
Journal:
Peptides
Year:
2014
Document type:
Article
Affiliation country:
Canada
Country of publication:
United States