IGF-I mRNA and Signaling in the Diabetic Retina

Abstract

IGF-I promotes the survival of multiple cell types by activating the IGF-I receptor (IGF-IR), which signals downstream to a serine/threonine kinase termed Akt. Because in diabetes vascular and neural cells of the retina undergo accelerated apoptosis, we examined IGF-I synthesis and signaling in the human and rat diabetic retina. In retinas obtained postmortem from six donors aged 64 ± 8 years with a diabetes duration of 7 ± 5 years, IGF-I mRNA levels were threefold lower than in the retinas of six age-matched nondiabetic donors (P = 0.005). In the retinas of rats with 2 months' duration of streptozotocin-induced diabetes, IGF-I mRNA levels were similar to those of control rats, but after 5 months of diabetes they failed to increase to the levels recorded in agematched controls (P < 0.02). Retinal IGF-I expression was not altered by hypophysectomy, proving to be growth-hormone independent. IGF-IR levels were modestly increased in the human diabetic retinas (P = 0.02 vs. nondiabetic retinas) and were unchanged in the diabetic rats. Phosphorylation of the IGF-IR could be measured only in the rat retina, and was not decreased in the diabetic rats (94 ± 18% of control values). In the same diabetic rats, phosphorylation of Akt was 123 ± 21% of control values. There was not yet evidence of increased apoptosis of retinal microvascular cells after 5 months of streptozotocin-induced diabetes. Hence, in the retina of diabetic rats, as in the retina of diabetic human donors, IGF-I mRNA levels are substantially lower than in agematched nondiabetic controls, whereas IGF-IR activation and signaling are not affected, at least for some time. This finding suggests that in the diabetic retina, the activation of the IGF-IR is modulated by influences that compensate for, or are compensated by, decreased IGF-I synthesis. Diabetes 50:175-183, 2001

A Role for the Polyol Pathway in the Early Neuroretinal Apoptosis and Glial Changes Induced by Diabetes in the Rat

“…Blood was obtained by cardiac puncture for the measurement of GHb (Glyc-Affin GHb assay; PerkinElmer, Norton, OH). From each animal, one eye was fixed in 10% buffered formalin at room temperature for the preparation of retinal wholemounts or sections, and the contralateral retina was isolated under the dissecting microscope and either homogenized for protein studies (14) or weighed and extracted in 6% ice-cold perchloric acid to measure sorbitol and fructose content. The measurements of retinal sorbitol and fructose were performed by the laboratory of Dr. Peter Oates at Pfizer using fluorometric enzyme assays (15,16).…”

“…The retinas were homogenized in ice-cold lysis buffer (14), and the proteins were resolved by electrophoresis on 12% SDS-PAGE and electroblotted. The blots were probed with the GFAP antibodies (1:40,000), and immunoreactive bands were detected and quantitated as previously described (14). The results are expressed as densitometric units per microgram of protein.…”

The PI3-K/Akt pathway: roles related to alterations in vasomotor responses in diabetic models

“…However, baseline Akt phosphorylation of serine-473, although not of threonine-308, and Akt activity (as determined by in vitro assay) showed reductions in STZ-diabetic rats (Laviola et al, 2001). Gerhardinger and coworkers (Gerhardinger et al, 2001) found that STZ-diabetic rats given low-dose insulin treatment tended to exhibit enhanced retinal Akt phosphorylation. We found that in STZ-diabetic rats, short-term insulin treatment can ameliorate endothelial dysfunction by inducing overexpressions of eNOS and/or VEGF mRNAs, possibly via IGF-1 receptors (Kobayashi and Kamata, 2002a).…”

The Increased Transforming Growth Factor-β Signaling Induced by Diabetes Protects Retinal Vessels

“…To capture the earliest effects of diabetes on the retinal vessels and the impact of TGF-b inhibition, we profiled gene expression in rats with 3.5 months of diabetes, a duration insufficient to cause histopathology 16 or even to accelerate apoptosis 18 in rat retinal vessels. A group of diabetic rats and a group of control rats received SM16 for 3 weeks; the drug did not affect body weight, blood glucose, or the levels of HbA 1c (Table 1).…”

“…The RNA was reverse-transcribed into cDNA, as previously described. 18 Gene-expression profiling was performed using the RT 2 Profiler Endothelial Cell Biology PCR Array (PARN-015; SABiosciences/Qiagen). The array plates included a panel of housekeeping genes used for normalizing the PCR array data, and the appropriate controls for genomic DNA contamination, RNA quality, and PCR performance.…”