Molecular Mechanisms in Diabetic Embryopathy

糖尿病孕妇出生缺陷的风险是糖尿病孕妇的3到10倍，先天畸形是糖尿病母亲所生婴儿死亡和发病的主要原因。为了制定预防糖尿病胚胎病变的策略，有必要在分子水平上了解其病因和发病机制。我们的假设是，糖尿病妊娠中的代谢失衡使发育中的胚胎中胰腺转录因子的表达失控，从而导致糖尿病胚胎病变。这一观点得到了转基因范例的支持。在小鼠中，胰腺转录因子ISL-1的转基因诱导出类似于人类糖尿病胚胎病变的表型，特别是神经管和尾部生长缺陷。在人类中，突变的HLXB9，一种位于ISL-1下游的胰腺转录因子，会导致骶骨发育不全。我们已经证明，尾部生长缺陷的基础是一种与剂量相关的对中胚层的侮辱。我们的模型指出，ISL-1的去调控导致其下游靶基因的去调控，而这些下游靶基因反过来是出生缺陷表型发病的效应因子。我们的假设提出了两个关键问题：(I)哪些因素调节胚胎中的ISL-1，以及ISL-1在糖尿病妊娠中的表达如何被解除调节？(Ii)哪些下游通路发生了改变，它们是否也与其他后部生长缺陷有关？在实验上，我们追求以下具体目标：(1)确定胚胎中正常ISL-1表达的调控元件，并确定糖尿病如何影响它们的活性。我们已经在ISL-1基因座上发现了一个尾部区域特异性增强子，它是代谢失衡去调节的候选控制元件。基因调控元件在糖尿病胚胎病变中的功能作用将在具有遗传或实验诱导糖尿病的小鼠妊娠中得到证实。(2)确定ISL-1的靶点，并探讨其在尾部生长缺陷发病机制中的作用。为了在我们的ISL-1转基因小鼠系统中识别ISL-1靶标，我们将使用基于微阵列的基因表达谱。初步的实时定量聚合酶链式反应结果表明，生长抑素和Wnt信号通路在孤立叶-L诱导的尾部生长缺陷中起重要作用。新的靶点，连同之前已确定的靶点PBX-1、PINC和Hlxb9，将在尾部缺陷和糖尿病妊娠后代的遗传模型中进行评估。这些实验将对胰腺转录因子的功能及其在代谢性疾病和胎儿发育中的致病潜力产生新的和全面的见解。

The risk for birth defects is 3 to 10-fold higher in diabetic pregnancy, and congenital malformations are the main cause of mortality and morbidity in infants born to mothers with diabetes. To devise preventive strategies for diabetic embryopathy, it is necessary to understand its etiology and pathogenesis at the molecular level. Our hypothesis is that metabolic imbalance in diabetic pregnancy de-regulates the expression of pancreatic transcription factors in the developing embryo, thus causing diabetic embryopathy. This idea is supported by transgenic paradigms. In mice, transgenes for the pancreatic transcription factor Isl-1 induce phenotypes that resemble human diabetic embryopathy, specifically neural tube and caudal growth defects. In humans, mutant HLXB9, a pancreatic transcription factor downstream of Isl-1, causes sacral agenesis. We have shown that the basis for caudal growth deficiency is a gone dosage-correlated insult to the mesoderm. Our model states that de-regulation of Isl-1 leads to de-regulation of its downstream target genes that, in turn, are the effectors for pathogenesis of the birth defect phenotype. Two key questions arise from our hypothesis: (i) Which factors regulate Isl-1 in the embryo, and how can Isl-1 expression become deregulated in diabetic pregnancy? (ii) Which downstream pathways are altered, and are they also involved in other posterior growth defects? Experimentally, we pursue the following specific aims: (1) To identify the regulatory elements for normal Isl-1 expression in the embryo, and determine how diabetes affects their activity. We have already identified a caudal region-specific enhancer in the Isl-1 locus, which is a candidate control element for de-regulation by metabolic imbalance. The functional role of gene regulatory elements in diabetic embryopathy will be established in murine pregnancies with genetic or experimentally induced diabetes. (2) To identify targets of Isl-1, and to investigate their role in the pathogenesis of caudal growth defects. To recognize Isl-1 targets in our Isl-1 transgenic mouse system, we will use microarray-based gene expression profiling. Preliminary quantitative real-time PCR results implicate the somatostatin and Wnt signaling pathways in Isl-l-induced caudal growth defects. Novel targets, together with the previously identified targets Pbx-1, Punc, and Hlxb9, will be evaluated in genetic models of caudal deficiencies, and progeny of diabetic pregnancies. These experiments will generate new and comprehensive insights into the function of pancreatic transcription factors and their pathogenic potential in metabolic disease and fetal development.