Expression of IGFBP-3 by human retinal endothelial cell cultures: IGFBP-3 involvement in growth inhibition and apoptosis.
OBJECTIVE
Growth hormone (GH), insulin-like growth factor (IGF), and somatostatin (SST) modulate each other's actions. SST analogues have been successfully used to treat proliferative diabetic retinopathy (PDR) that is unresponsive to laser therapy and to retard the progression of severe nonproliferative retinopathy to PDR. In this study, the endogenous expression of IGF-binding protein (IGFBP)-3 was examined in human retinal endothelial cells (HRECs), the direct effects of IGFBP-3 on HRECs were evaluated, and the possible involvement of IGFBP-3 in mediating the growth inhibitory effects of SST receptor (SSTR) agonists in HRECs was assessed.


METHODS
The cellular localization of IGFBP-3 was examined with anti-IGFBP-3 and fluorescein-conjugated goat anti-rabbit IgG. HRECs were exposed to varying concentrations of human recombinant IGFBP-3, and growth inhibition was evaluated by thiazolyl blue (MTT) conversion. Apoptosis was examined using fluorochrome-annexin V staining. Conditioned media (CM) from SSTR2 agonist (L779976)-treated or SSTR3 agonist (L796778)-treated HRECs were analyzed by ELISA for changes in expression of IGFBP-3.


RESULTS
HREC immunostaining showed cell surface and cytoplasmic IGFBP-3. Exogenous IGFBP-3 induced a dose-dependent inhibition of HREC proliferation and staining with fluorochrome-annexin V showed numerous apoptotic HRECs. HRECs exposed to the SSTR2 or SSTR3 agonists expressed IGFBP-3 in a concentration-dependent manner.


CONCLUSIONS
Cultured HRECs expressed endogenous IGFBP-3. Exogenous administration of IGFBP-3-induced growth inhibition and apoptosis, supporting a regulatory role for IGFBP-3 in endothelial cells. SSTR agonists mediate their growth-inhibitory effect, in part, by increasing expression of IGFBP-3.
Spoerri
Polyxenie E
PE
Department of Pharmacology and Therapeutics, University of Florida, PO Box 100267, Gainesville, FL 32610-0267, USA. spoerrip@ufl.edu
Caballero
Sergio
S
Wilson
Sylvia H
SH
Shaw
Lynn C
LC
Grant
Maria B
MB
D000577:Amides / Q000494:pharmacology; D017304:Annexin A5; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D018929:Cell Culture Techniques; D002455:Cell Division / Q000187:drug effects / Q000502:physiology*; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000378:metabolism* / Q000494:pharmacology; D009578:Nitrobenzenes / Q000494:pharmacology; D017481:Receptors, Somatostatin / Q000819:agonists; D011994:Recombinant Proteins / Q000494:pharmacology; D012171:Retinal Vessels / Q000166:cytology*; D013778:Tetrazolium Salts; D013844:Thiazoles
D000577:Amides; D017304:Annexin A5; D007211:Indoles; D018972:Insulin-Like Growth Factor Binding Protein 3; C121854:L 779976; C429627:L796778; D009578:Nitrobenzenes; D017481:Receptors, Somatostatin; D011994:Recombinant Proteins; D013778:Tetrazolium Salts; D013844:Thiazoles; C022616:thiazolyl blue