Elevated IGFBP3 Levels in Diabetic Tears: A Negative Regulator of IGF-1 Signaling in the Corneal Epithelium

• Published in: The ocular surface Vol. 10; no. 2; pp. 100 - 107
• Main Authors: Wu, Yu-Chieh, PhD, Buckner, Benjamin R., BS, Zhu, Meifang, MS, Cavanagh, H. Dwight, MD, PhD, Robertson, Danielle M., OD, PhD
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.04.2012
• Subjects: Adult; Blotting, Western; Cells, Cultured; cornea; Corneal Diseases - metabolism; Diabetes; Diabetes Mellitus, Type 1 - metabolism; Diabetes Mellitus, Type 2 - metabolism; Enzyme-Linked Immunosorbent Assay; epithelium; Epithelium, Corneal - metabolism; Female; Humans; IGF-1; IGF-1R; IGFBP3; Insulin-Like Growth Factor Binding Protein 3 - genetics; Insulin-Like Growth Factor Binding Protein 3 - metabolism; Insulin-Like Growth Factor I - metabolism; Male; Middle Aged; Ophthalmology; Real-Time Polymerase Chain Reaction; Receptor, IGF Type 1 - genetics; tears; Tears - metabolism; tears; cornea; IGFBP3; epithelium; IGF-1; Diabetes; IGF-1R
• ISSN: 1542-0124; 1937-5913; 1542-0124
• DOI: 10.1016/j.jtos.2012.01.004

Abstract To determine the ratio of IGFBP3:IGF-1 in normal and diabetic human tears, and in telomerase-immortalized human corneal epithelial cells (hTCEpi) cultured under elevated glucose conditions and to correlate these changes with total and phosphorylated levels of IGF-1R. Tear samples were collected noninvasively from diabetic subjects and non-diabetic controls; corneal sensitivity was assessed using a Cochet-Bonnet Aesthesiometer. Conditioned media were collected following culture of hTCEpi cells in normal (5 mM) and elevated (25 mM) glucose conditions; mannitol was used as an osmotic control. IGFBP3, IGF-1, and phosphorylated IGF-1R levels were assessed by ELISA. IGFBP3 and IGF-1R mRNA were assessed by real-time polymerase chain reaction (PCR). Total and phosphorylated IGF-1R expression in whole cell lysates was assessed by western blot. There was a 2.8-fold increase in IGFBP3 in diabetic tears compared to non-diabetic controls ( P =0.006); IGF-1 levels were not significantly altered. No difference in corneal sensitivity was detected between groups. The concentration of IGFBP3 in tears was independent of IGF-1. Consistent with human tear measurements in vivo, IGFBP3 secretion was increased 2.2 fold ( P <0.001) following culture of hTCEpi cells under elevated glucose conditions in vitro. Treatment with glucose and the mannitol control reduced IGFBP3 mRNA ( P <0.001). Total IGF-1R levels were unchanged. The increase in the IGFBP3:IGF-1 ratio detected in diabetic tears compared to normal controls blocked phosphorylation of the IGF-1R by IGF-1 ( P <0.001) when tested in vitro. Taken together, these in vivo and confirmatory in vitro findings suggest that the observed increase in IGFBP3 found in human tears may attenuate IGF-1R signaling in the diabetic cornea. A long-term increase in IGFBP3 may contribute to epithelial compromise and the pathogenesis of ocular surface complications reported in diabetes.