HORMONAL REGULATION OF LUNG DEVELOPMENT; PTHrP BIOLOGY

描述（由申请方提供）：甲状旁腺激素相关蛋白（PTHrP）缺失小鼠肺无法肺泡化。我们推测PTHrP的表达和扩增是肺发育、稳定和修复所必需的。我们的长期目标是确定肺形态发生的细胞/分子基础。从机制上讲，在肺发育的前肺泡假腺阶段，上皮PTHrP上调成纤维细胞FGF-10，刺激PTHrP的上皮Sonic Hedgehog（Shh）扩增。PTHrP信号传导因此下调成纤维细胞Wnt信号传导（通过刺激蛋白激酶A），上调脂成纤维细胞（LIF）分化，特别是PTHrP受体表达，进一步放大用于II型细胞分化的PTHrP信号传导。在目的1中，我们假设删除PTHrP表达将阻止LIF促进肺泡发育所需的FGF-10和Shh的上调。我们进一步假设，PTHrP的条件性再表达，这决定了LIF的表型，将“拯救”空小鼠的肺泡化。在目标2中，我们假设在无效的成年小鼠肺中缺失PTHrP信号传导将诱导肌成纤维细胞（MYF）分化，引起间质纤维化。我们将确定PTHrP的条件性再表达是否可以防止MYF分化和肺间质纤维化。我们将使用一个层次的方法来验证这些假设，包括免疫组织学，原位杂交，真实的时间RT-PCR和Western印迹的整个肺;形态计量学结合激光捕获显微切割/真实的时间RT-PCR切片肺;和分离的肺细胞mRNA转染，微阵列和稳定同位素代谢组学。这种系统的方法将确定PTHrP是否通过Wnt或PKA途径的细胞间信号传导决定肺泡发育和病理。这些研究将为新生儿和成人慢性肺部疾病的病理生理学和治疗提供重要的见解。

DESCRIPTION (provided by applicant): The Parathyroid Hormone-related Protein (PTHrP)-null mouse lung fails to alveolize. We hypothesize that PTHrP expression and amplification are necessary for lung development, stability and repair. Our long-term goal is to determine the cell/molecular basis of lung morphogenesis. Mechanistically, during the pre-alveolar pseudoglandular phase of lung development epithelial PTHrP up-regulates flbroblast FGF-10, stimulating epithelial Sonic Hedgehog (Shh) amplification of PTHrP. PTHrP signaling consequently down-regulates fibroblast Wnt signaling (by stimulating Protein Kinase A), up-regulating lipofibroblast (LIF) differentiation particularly PTHrP receptor expression, further amplifying PTHrP signaling for type II cell differentiation. In Aim 1 we hypothesize that deleting PTHrP expression will prevent the up-regulation of FGF-10 and Shh needed for LIF promotion of alveolar development. We further hypothesize that conditional re-expression of PTHrP, which determines the LIF phenotype, will 'rescue' alveolization in the null mouse. In Aim 2 we hypothesize that deleting PTHrP signaling in the null adult mouse lung will induce myofibroblast (MYF) differentiation, causing interstitial fibrosis. We will determine if conditional re-expression of PTHrP can prevent MYF differentiation and interstitial lung fibrosis. We will use a hierarchical approach to test these hypotheses, including immunohistology, in situ hybridization, Real Time RT-PCR and Western Blotting of whole lung; morphometry combined with Laser Capture Microdissection/Real Time RT-PCR of sectioned lung; and isolated lung cell mRNA transfection, microarray and stable isotope metabolomics. This systematic approach will determine if PTHrP determines lung alveolar development and pathology through intercellular signaling via the Wnt or PKA pathway. These studies will provide important insights to the pathophysiology and treatment of chronic lung diseases of the newborn and adult alike.