SEARCH FOR THE MOLECULAR CAUSES OF DIABETIC EMBRYOPATHY

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Diabetic embryopathy is a well recognized, but poorly understood, complication of diabetes in which the early embryo of a diabetic method develops congenital malformations. The objective of the work proposed here is to study the regulation by diabetes of a critical development control gene, Pax-3, during the development of one of the most common diabetes- associated malformations, neural tube defects (NTD). Using a new mouse model that was developed in my laboratory, we have observed that the rate of NTD is about three-fold higher in the embryos of diabetic mice than in the embryos of non-diabetic mice. The high rate of NTD is correlated with reduced expression of Pax-3, an embryonic gene which encodes a DNA-binding transcription factor that is required for formation of the brain and spinal chord. Subsequent to the reduction in expression of Pax-3, and apparently as a consequence, cells forming the neural tube are seen to undergo unscheduled apoptosis. There are three specific aims of this proposal. In the first aim, we will test the hypothesis that glucose toxicity associated with maternal diabetes is responsible for abnormalities in embryonic gene expression, apoptosis, and NTD. This will be accomplished by (1) attempting to prevent glucose toxicity by lowering glucose levels in pregnant diabetic mice with insulin or phlorizin administration, (2) attempting to induce glucose toxicity in pregnant non-diabetic mice by a hyperglycemic glucose clamp procedure, (3) attempting to induce glucose toxicity during culture of post-implementation mouse embryos in media containing elevated levels of glucose, and (4) testing whether expression of the high Km GLUT-2 glucose transporter renders the early embryos sensitive to glucose toxicity during diabetic pregnancy. In the second aim, we will determine whether the reduction in Pax-3 mRNA observed in embryos of diabetic and non-diabetic mice is due to transcriptional or post-transcriptional control, and identify the Pax-3 control elements involved in inhibition by diabetes. In the third aim, we will test the hypothesis that mouse strains which fail to develop NTD during diabetic pregnancy are resistant to the inhibition of Pax-3 expression that occurs in strains which are susceptible to NTD. These experiments will reveal, in a way that has not been done in the past, the molecular mechanisms by which embryonic development is during diabetic pregnancy.