Lactose supports Muller cell protein expression patterns in the absence of the retinal pigment epithelium.
OBJECTIVE
We have previously shown that lactose, but not mannose, promotes the assembly of nascent photoreceptor outer segments in the absence of the retinal pigment epithelium (RPE). The purpose of the present study was to determine if, in addition to the improved outer segment assembly observed in the presence of lactose, biosynthetic changes in Muller cells could also be detected.


METHODS
The RPE was removed from intact isolated Xenopus embryonic eyes that were allowed to complete differentiation in Niu-Twitty medium, Niu-Twitty with mannose, or Niu-Twitty with lactose. Control retinas matured in vitro with an adherent RPE. Retinal morphology was evaluated for organized folding of outer segment membranes and cell loss. In addition, the expression of three Muller cell proteins, glial fibrillary acidic protein (GFAP), cellular retinaldehyde-binding protein (CRALBP), and glutamine synthetase, was examined.


RESULTS
In control retinas, GFAP is undetectable, CRALBP heavily immunolabels Muller cells, and radial patterns of glutamine synthetase immunoreactivity are present. In the absence of the RPE, Muller cells upregulate GFAP expression, CRALBP labeling is present at a slightly reduced level, and glutamine synthetase immunolabeling is negligible. Neither mannose nor lactose modify significantly the expression of CRALBP. Similarly, both compounds completely prevent the upregulation of GFAP. However, normal glutamine synthetase expression was observed only in the presence of lactose, but not in the presence of mannose. Statistical analyses of slot blot-based protein quantification confirmed our immunochemical results.


CONCLUSIONS
In RPE-deprived retinas supplemented with lactose, Muller cells were morphologically normal. The proper photoreceptor outer segment morphogenesis observed under these conditions was uniquely associated with normal levels of glutamine synthetase expression. The exact significance of this finding with respect to photoreceptor outer segment morphogenesis is unknown. We suggest that glutamine synthetase may be a marker of Muller cell metabolic or structural integrity that may reflect the enhanced ability of these cells, in the presence of lactose, to support photoreceptor outer segment morphogenesis.
Retinal Degeneration Research Center, Department of Ophthalmology, The University of Tennessee Health Science Center at Memphis, Memphis, TN 38163, USA. mjablonski@mail.eye.utmem.edu
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Iannaccone
D000818:Animals; D002352:Carrier Proteins / Q000096:biosynthesis*; D002478:Cells, Cultured; D018920:Coculture Techniques; D005904:Glial Fibrillary Acidic Protein / Q000096:biosynthesis*; D005974:Glutamate-Ammonia Ligase / Q000096:biosynthesis*; D007150:Immunohistochemistry; D007785:Lactose / Q000494:pharmacology*; D008358:Mannose / Q000494:pharmacology; D009024:Morphogenesis; D009457:Neuroglia / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000502:physiology*; D012160:Retina / Q000196:embryology; D015854:Up-Regulation; D014982:Xenopus laevis
C020932:11-cis-retinal-binding protein; D002352:Carrier Proteins; D005904:Glial Fibrillary Acidic Protein; D005974:Glutamate-Ammonia Ligase; D007785:Lactose; D008358:Mannose