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Characterization of a far-red analog of ghrelin for imaging GHS-R in P19-derived cardiomyocytes.
Douglas, Gregory A F; McGirr, Rebecca; Charlton, Carlie L; Kagan, Dov B; Hoffman, Lisa M; Luyt, Leonard G; Dhanvantari, Savita.
Affiliation
  • Douglas GA; Departments of Pathology, Chemistry, Medical Imaging, Medical Biophysics, and Oncology, Western University, Richmond Street, London, Ontario, Canada; Imaging Program, Lawson Health Research Institute, 268 Grosvenor Street, London, Ontario N6A 4V2, Canada.
  • McGirr R; Departments of Pathology, Chemistry, Medical Imaging, Medical Biophysics, and Oncology, Western University, Richmond Street, London, Ontario, Canada; Imaging Program, Lawson Health Research Institute, 268 Grosvenor Street, London, Ontario N6A 4V2, Canada.
  • Charlton CL; Departments of Pathology, Chemistry, Medical Imaging, Medical Biophysics, and Oncology, Western University, Richmond Street, London, Ontario, Canada; Imaging Program, Lawson Health Research Institute, 268 Grosvenor Street, London, Ontario N6A 4V2, Canada.
  • Kagan DB; Departments of Pathology, Chemistry, Medical Imaging, Medical Biophysics, and Oncology, Western University, Richmond Street, London, Ontario, Canada; Imaging Program, Lawson Health Research Institute, 268 Grosvenor Street, London, Ontario N6A 4V2, Canada.
  • Hoffman LM; Departments of Pathology, Chemistry, Medical Imaging, Medical Biophysics, and Oncology, Western University, Richmond Street, London, Ontario, Canada; Imaging Program, Lawson Health Research Institute, 268 Grosvenor Street, London, Ontario N6A 4V2, Canada.
  • Luyt LG; Departments of Pathology, Chemistry, Medical Imaging, Medical Biophysics, and Oncology, Western University, Richmond Street, London, Ontario, Canada; Imaging Program, Lawson Health Research Institute, 268 Grosvenor Street, London, Ontario N6A 4V2, Canada.
  • Dhanvantari S; Departments of Pathology, Chemistry, Medical Imaging, Medical Biophysics, and Oncology, Western University, Richmond Street, London, Ontario, Canada; Imaging Program, Lawson Health Research Institute, 268 Grosvenor Street, London, Ontario N6A 4V2, Canada. Electronic address: sdhanvan@lawsonimaging.c
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.
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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
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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