EXPERIMENTAL CELL RESEARCH 241, 210–221 (1998) ARTICLE NO. EX984048 Suppression of Fibroblast Growth Factors 1 and 2 by Antisense Oligonucleotides in Embryonic Chick Retinal Cells in Vitro Inhibits Neuronal Differentiation and Survival Laurent De ´sire ´, Yves Courtois, and Jean-Claude Jeanny 1 De ´veloppement, vieillissement et pathologie de la re ´tine, INSERM U. 450, affilie ´e CNRS, Association Claude Bernard, 29, rue Wilhem, 75016, Paris, France in the appropriate innervation and connectivity of the As retinal histogenesis proceeds there is a pro- mature CNS [3]. nounced increase in the expression of fibroblast One of the biological models available to study the growth factor (FGF ), reaching its maximum in the ma- action of neurotrophic factors on neurogenesis in the ture retina and largely in terminal differentiated reti- CNS is the avian retina. This organ is derived from an nal neurons. Recent in vivo evidence suggests that ex- invagination of the optic vesicle leading, after waves of ogenous FGF functions as a differentiation and sur- proliferation, differentiation, and cell death [4, 5], to vival factor for a wide variety of cell types including a well-defined structure of concentric neuronal layers. CNS neurons and that endogenous FGF may perform Neurotrophic factors, such as fibroblast growth factor similar functions. We have examined the consequences 1 (FGF1) and FGF2 (acidic and basic fibroblast growth of selectively and independently inhibiting FGF1 or factor, respectively), are thought to play an important FGF2 expression using antisense oligonucleotides in role in retinal development and maintenance. embryonic chick retinal cells, differentiating in vitro. In vitro and in vivo, the neurotrophic activity of FGF Whether FGF1 or FGF2 expression was inhibited the in the retina has been studied mostly by examining the results were the same: a marked reduction in neuronal action of exogenous FGF1 or FGF2 or by spatiotempo- photoreceptor cells differentiation, an increase in pro- ral correlations of expression. In vitro, FGF1 and FGF2 grammed cell death, but no effects on cell prolifera- increase proliferation of glial cells and differentiation tion. Even although these two related factors promote the same final effect on retinal cells, namely, neuronal and survival of several types of retinal neurons [6–9]. differentiation and survival, their normal combined In the chick embryo, endogenous FGF1 and FGF2 are activities or levels appear to be important in achieving present at very low levels in early development, and this effect. Stimulation with either exogenous FGF1 their expression strongly increases in all retinal neu- or FGF2 served to increase endogenous levels of both rons, following the sequential appearance of the neu- FGF1 and FGF2 and reversed the effects of antisense ronal layers [10 – 14]. Additionally, these factors exhibit blockade of either FGF1 or FGF2. Our data suggest important neuroprotective effects in animal models of that although other sources of FGF exist within the retinal neurodegeneration [6, 15 – 18]. This suggests a eye, the function of endogenous FGF in differentiating direct role for endogenous FGF1 and FGF2 in prolifera- retinal neurons may be to stimulate their differentia- tion, migration, early and terminal differentiation of tion and promote their survival.  1998 Academic Press retinal neurons, and in retinal maintenance. FGF1 and 2 (but not NGF, TGFb, and EGF) trigger chick neural retina regeneration from the pigmented epi- INTRODUCTION thelium [19 – 22]. A recent study [23] suggested that in early development, FGF2 acts as an extrinsic factor in Developing neurons of the central nervous system the induction of the presumptive neural retina from the (CNS) compete for disposable neurotrophic factors es- pigmented epithelium. This pathway can be blocked by sential for their differentiation, maintenance, and sur- anti-FGF2 neutralizing antibodies and is fully consistent vival [1, 2] delivered by their targets, innervating neu- with the distribution of FGF receptors in the developing rons, or neighboring glial cells. A consequence is the eye cup [13, 24 – 26]. However, this blockade is efficient subsequent developmental neuronal death that results during the first events of ganglion cells differentiation and not later in development. Alternatively, blockade of FGF signaling during early ganglion cell differentiation 1 To whom correspondence and reprint requests should be ad- dressed. Fax: 33 1 40 50 01 95. E-mail: jcjeanny@infobiogen.fr. using dominant negative FGF-R [27 – 29] does not lead 210 0014-4827/98 $25.00 Copyright  1998 by Academic Press All rights of reproduction in any form reserved.