Maternal diabetes pre-dating pregnancy significantly increases the risk for congenital malformations. The malformations occur very early during gestation, at the beginning of organogenesis. In an effort to understand the molecular mechanisms by which maternal diabetes disturbs embryonic development, previous research from the Principal Investigator's laboratory has shown that, in a mouse model of diabetic pregnancy, excess glucose metabolism by the embryo inhibits expression of Pax3, a gene that encodes a transcription factor required for development of neuroepithelium and neural crest. Reduced expression of Pax3 is associated with increased neural tube and cardiac outflow tract defects, two of the most common defects that occur in diabetic pregnancy. As a result of insufficient Pax3 expression, cells forming the neural tube or neural crest undergo p53-dependent apoptosis, and consequently, these structures fail to form properly. Recent research using embryonic stem cells (ESC) and cancer cells has demonstrated that self-renewability and pluripotentiality are coupled to a high rate of glycolysis. As ESC start to differentiated, p53 becomes activated and regulates expression of genes and enzyme activity that promote differentiation, senescence, and oxidative metabolism. Pax3-expressing cells are progenitors that must be able to proliferate and maintain plasticity to develop into multiple different specialized cell types until the stages of development when cells should stop proliferating and terminally differentiate. The overall hypothesis to be tested is that Pax3 is required during early development of neuroepithelial and neural crest cells because it must inhibit the thrust of p53 to prematurely cause senescence, differentiation, and high rates of oxidative metabolism. Evidence acquired during the previous funding period using mouse ESC indicates that Pax3 inhibits p53 stability, and that Pax3 and p53 are found in a complex with each other. However, it is not known how Pax3 destabilizes p53, nor whether degradation of p53 in response to Pax3 prevents the loss of stem cell characteristics as embryo cells start to differentiate.
The Specific Aims of this proposal are to: (1) Determine the mechanism by which Pax3 induces p53 degradation. (2) Test whether down regulation of p53 by Pax3 prevents loss of stem cell characteristics by supporting expression of genes required for self-renewal and inhibition of differentiation. (3) Test whether down regulation of p53 by Pax3 prevents loss of the balance of anaerobic: aerobic glucose metabolism that is characteristic of undifferentiated cells. These experiments will provide insight into the mechanisms by which malformations occur during diabetic pregnancy and other conditions when Pax3 expression is impaired. Furthermore, they will provide new information into the regulation of early embryo and stem cell fate by glycolytic and oxidative energy metabolism.

Public Health Relevance

Diabetes existing before pregnancy significantly increases risk for birth defects;the incidence of birth defects in the offspring of diabetic women is 2-5 times that in nondiabetic pregnancy. This diabetic complication is known as, diabetic embryopathy. The birth defects occur at the very earliest stages of embryonic development, when the organ systems are just starting to form. This project will investigate how altered expression of a gene that is required during early embryo development, resulting from diabetic pregnancy, leads to birth defects.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK052865-13
Application #
8265893
Study Section
Pregnancy and Neonatology Study Section (PN)
Program Officer
Jones, Teresa L Z
Project Start
1998-08-01
Project End
2014-02-28
Budget Start
2012-03-01
Budget End
2013-02-28
Support Year
13
Fiscal Year
2012
Total Cost
$348,840
Indirect Cost
$132,840
Name
Joslin Diabetes Center
Department
Type
DUNS #
071723084
City
Boston
State
MA
Country
United States
Zip Code
02215
Loeken, Mary R (2014) Intersection of complex genetic traits affecting maternal metabolism, fetal metabolism, and neural tube defect risk: looking for needles in multiple haystacks. Mol Genet Metab 111:415-7
Sanders, Kaitlyn; Jung, Jin Hyuk; Loeken, Mary R (2014) Use of a murine embryonic stem cell line that is sensitive to high glucose environment to model neural tube development in diabetic pregnancy. Birth Defects Res A Clin Mol Teratol 100:584-91
Jung, Jin Hyuk; Wang, Xiao Dan; Loeken, Mary R (2013) Mouse embryonic stem cells established in physiological-glucose media express the high KM Glut2 glucose transporter expressed by normal embryos. Stem Cells Transl Med 2:929-34
Wang, Xiao Dan; Morgan, Sarah C; Loeken, Mary R (2011) Pax3 stimulates p53 ubiquitination and degradation independent of transcription. PLoS One 6:e29379
Zabihi, Sheller; Loeken, Mary R (2010) Understanding diabetic teratogenesis: where are we now and where are we going? Birth Defects Res A Clin Mol Teratol 88:779-90
Chappell Jr, James H; Wang, Xiao Dan; Loeken, Mary R (2009) Diabetes and apoptosis: neural crest cells and neural tube. Apoptosis 14:1472-83
Morgan, Sarah C; Lee, Hyung-Yul; Relaix, Frederic et al. (2008) Cardiac outflow tract septation failure in Pax3-deficient embryos is due to p53-dependent regulation of migrating cardiac neural crest. Mech Dev 125:757-67
Morgan, Sarah C; Relaix, Frederic; Sandell, Lisa L et al. (2008) Oxidative stress during diabetic pregnancy disrupts cardiac neural crest migration and causes outflow tract defects. Birth Defects Res A Clin Mol Teratol 82:453-63
Li, R; Thorens, B; Loeken, M R (2007) Expression of the gene encoding the high-Km glucose transporter 2 by the early postimplantation mouse embryo is essential for neural tube defects associated with diabetic embryopathy. Diabetologia 50:682-9
Chang, T I; Horal, M; Jain, S K et al. (2003) Oxidant regulation of gene expression and neural tube development: Insights gained from diabetic pregnancy on molecular causes of neural tube defects. Diabetologia 46:538-45

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