Suppression of Butyrylcholinesterase Induces Overexpression of Acetylcholinesterase and Increased Apoptosis in Embryonic Chick Retinospheroids

Abstract

We here report the generation of a reaggregation system of embryonic chick retinal cells by transfection with sense- and antisense-cholinesterase expression vectors. Reaggregation of embryonic retina cells leads to so-called retinospheroids, which show a high degree of structural development. The cholinesterases butyrylcholinesterase (BChE) as a proliferation marker and acetylcholinesterase (AChE) as a differentiation marker are expressed in embryonic retina in a mutually exclusive manner and seem to be coregulated (1). BChE is expressed before and during mitosis, whereas AChE is expressed about 10 to 15 hours after the last mitosis. During the proliferation period, inhibition of BChE transcription by an antisense strategy suggests that decrease of BChE mRNA promotes an increase of acetylcholinesterase. Moreover, a strong decrease of proliferation and growth of antisense 5′-BChE transfected retinospheroids was apparent (2). These facts demonstrate that AChE is expressed early in neuronal development, however, it is not localized to the neuronal cell membrane until later during the process of synaptogenesis. As detected by vital propidium iodide uptake, TdT-mediated dUTP nick end labeling (TUNEL) and DNA fragmentation assays, the suppression of BChE during the period of differentiation, resulted in an increase of apoptosis. Furthermore, the number of AChE-positive cells increased dramatically, whereas rod- and cone-specific photoreceptor precursor cells decreased. Thereby, red and green cones seem to be completely degenerated (3). These resuits suggest that the proliferation marker BChE determines an intracellular balance between proliferation, differentiation and apoptosis during retinogenesis. The inhibition of BChE induces a higher degree of differentiation of AChE-positive cells, and a higher rate of apoptotic photoreceptor precursors.