q 2001 International Society for Neurochemistry, Journal of Neurochemistry, 77, 1157±1167 1157 Journal of Neurochemistry, 2001, 77, 1157±1167 Effects of insulin-like growth factor-1 on retinal endothelial cell glucose transport and proliferation Brian J. DeBosch,* Elisabeth Baur,* Baljit K. Deo,* Miki Hiraoka* and Arno K. Kumagai* , ² *Department of Internal Medicine, Michigan Diabetes Research and Training Center, Ann Arbor, Michigan, USA ²JDRF Center for Complications in Diabetes, University of Michigan Medical School, Ann Arbor, Michigan, USA Abstract Insulin-like growth factor-1 (IGF-1) plays important roles in the developing and mature retina and in pathological states characterized by retinal neovascularization, such as diabetic retinopathy. The effects of IGF-1 on glucose transport and proliferation and the signal transduction pathways underlying these effects were studied in a primary bovine retinal endothelial cell (BREC) culture model. IGF-1 stimulated uptake of the glucose analog 2-deoxyglucose in a dose- dependent manner, with a maximal uptake at 25 ng/mL (3.3 nM) after 24 h. Increased transport occurred in the absence of an increase in total cellular GLUT1 transcript or protein. IGF-1 stimulated activity of both protein kinase C (PKC) and phosphatidylinositol-3 kinase (PI3 kinase), and both pathways were required for IGF-1-mediated BREC glucose transport and thymidine incorporation. Use of a selective inhibitor of the b isoform of PKC, LY379196, revealed that IGF-1 stimulation of glucose transport was mediated by PKC-b; however, inhibition of PKC-b had no effect on BREC proliferation. Taken together, these data suggest that the actions of IGF-1 in retinal endothelial cells couple proliferation with delivery of glucose, an essential metabolic substrate. The present studies extend our general understanding of the effects of IGF-1 on vital cellular activities within the retina in normal physiology and in pathological states such as diabetic retinopathy. Keywords: diabetic retinopathy, glucose transport, GLUT1, inner blood±retinal barrier, insulin-like growth factor, retinal endothelial cells. J. Neurochem. (2001) 77, 1157±1167. Insulin-like growth factor-1 (IGF-1), a polypeptide hormone that mediates the systemic actions of growth hormone (GH) (Jones and Clemmons 1995), is synthesized by a variety of cell types in the retina, including microvascular endothelial cells, pericytes, Mu Èller cells, ganglion cells, and retinal pigment epithelium (RPE) (Lee et al. 1992). In normal retinal physiology, IGF-1 is thought to play a role in the development of the neuroretina in different species (Ocrant et al. 1989; Calvaruso et al. 1996), and the expression of IGF-1, IGF-1 binding proteins and the IGF-1 receptor in the adult mammalian retina (Waldbillig et al. 1988, 1991; Ocrant et al. 1989) suggests that IGF-1 participates in processes in the mature retina as well. These processes include interactions between the RPE and photoreceptor in the vision signal transduction cascade (Waldbillig et al. 1988, 1991), and analogous to its role in the brain, IGF-1 also appears to act as an anti-apoptotic factor in an experimental model of retinal injury (Kermer et al. 2000). In pathological states characterized by retinal neovascular- ization, such as diabetic retinopathy (DR), accumulating evidence suggests that IGF-1, along with vascular endothelial growth factor (VEGF), serves as a key mediator in disease progression. Elevated vitreal and serum concen- trations of IGF-1 have been documented in patients with proliferative diabetic retinopathy (PDR) (Merimee et al. 1983; Grant et al. 1986; Meyer-Schwickerath et al. 1993; Burgos et al. 2000) and are in the range known to stimulate Received December 8, 2000; revised manuscript received February 23, 2001; accepted February 26, 2001. Address correspondence and reprint requests to Dr Arno K. Kumagai, Department of Internal Medicine, 5570 MSRB-2, University of Michigan Medical School, Ann Arbor, MI 48109±0678, USA. E-mail: akumagai@umich.edu Abbreviations used: BRB, blood±retinal barrier; BREC, bovine retinal endothelial cells; 2DG, 2-deoxyglucose; DMEM, Dulbecco's modi®ed Eagle's medium; DR, diabetic retinopathy; EGS, endothelial growth supplement; FCS, fetal calf serum; GH, growth hormone; IGF-1, insulin-like growth factor-1; KRP-BSA, Kreb's Ringers phosphate buffer containing 1% bovine serum albumin; 3MG, 3-O-methylglucose; PKC, protein kinase C; PI3 kinase, phosphatidylinositol-3 kinase; PDR, proliferative diabetic retinopathy; PMSF, phenylmethylsulfonyl ¯uoride; RPE, retinal pigment epithelium; SDS, sodium dodecyl sulfate; VEGF, vascular endothelial growth factor.