Paracrine IGF/GFBP Interactions In Vivo

描述(申请人提供)：胰岛素样生长因子(IGFS)调节细胞的增殖、存活和分化。这些行动由当地生产的政府间气候变化框架以及从流通中衍生的政府间气候变化框架来执行。IGF在血浆和细胞间质之间的分配可能部分是由高亲和力IGF结合蛋白家族的成员完成的。我们认为，旁分泌IGF-I和IGFBPs的功能最好是在体内组织特异性扰动后探索，因为它们的活性似乎是在损伤或其他触发组织重塑的因素之后被招募的。当受到特殊的病理生理挑战时，血管壁、肠和膀胱的SMC保持着增殖和重塑细胞外基质的能力。其中IGF-I最为突出，因为它能在体外和体内刺激SMC的生长和存活。球囊损伤后，IGF-I基因表达明显上调。此外，在SMC中靶向过表达IGF-I的转基因小鼠在颈动脉损伤后通过促进增殖和细胞迁移促进了新生内膜的形成。尽管如此，作为损伤反应中介物的IGF-I产生旁分泌的要求还没有最终确定，决定局部IGF-I生物利用度的其他因素的作用也不是决定性的。在这项提案中，我们将使用老鼠遗传策略来解决其中一些问题。我们将致力于以下目标：1)确定IGF受体在血管损伤后的反应中是否需要IGFFR：我们将首先研究IGF I型受体(IGF)特异性缺失对动脉损伤后血管生长、迁移和新生内膜形成的影响。2)确定血管损伤对LGF-1产生旁分泌的需求：我们将在SMP8-cre-ERT2小鼠和纯合子loxP-Igf1小鼠杂交中探讨这一问题。损伤实验将在他莫昔芬导致IGFI等位基因缺失和局部IGF-I组织存储耗尽后进行。3)确定局部IGFBP在血管损伤反应中的表达需求：我们将首先通过以他莫昔芬依赖的方式靶向删除平滑肌细胞中的lGFBP-4来探讨这一点。我们将测试SMP8-cre-ERT2/loxP-lgfBP-4小鼠的假设，即SMP8-cre-ERT2/loxP-lgfBP-4小鼠的IGF-I的平滑肌组织储存减少，这将与对颈动脉损伤的反应受损有关。4)确定锌金属蛋白酶PAPP-A在损伤反应中局部表达的必要性：在此，我们将检验一种假说，即在他莫昔芬诱导的FLOFLED PAPP-A重组后，IGFBP蛋白酶活性将被取消或抑制，IGFBP-4稳定，导致IGF-I封闭，SMC迁移和增殖受损。

DESCRIPTION (provided by applicant): Insulin-like growth factors (IGFs) regulate cell proliferation, survival and differentiation. These actions are carried out by locally produced IGFs as well as IGFs derived from the circulation. IGF partition between plasma and cellular interstitial compartments is likely accomplished in part by members of a family of high affinity IGF binding proteins. We believe the function of paracrine IGF-I and of the IGFBPs is best explored after tissue-specific perturbations in vivo, since their activity appears to be recruited after injury or other triggers for tissue remodeling. SMC of the vascular wall, intestine and urinary bladder retain the ability to proliferate and to remodel the extracellular matrix when subjected to particular pathophysiological challenges. IGF-I is prominent among them, as it stimulates SMC growth and survival in vitro and in vivo. After balloon endoarterial injury, there is a marked induction of IGF-I gene expression. Moreover, transgenic mice with targeted overexpression of IGF-I in SMC have enhanced neointimal formation through increased proliferation and cell migration after carotid artery injury. Despite this, the requirement for paracrine production of IGF-I as a mediator of injury responses has not been conclusively established, nor is the role of other factors determining local IGF-I bioavailability. In this proposal we will address some of these questions using mouse genetic strategies. We will pursue the following aims: 1) Determine whether lgflr is required for the smooth muscle response to injury: We will first characterize the effects of smooth muscle-specific deletion of the IGF type I receptor (Igflr) on growth, migration and neointimal formation after arterial injury. 2) Determine the requirement of paracrine production of lGF-1 in response to vascular injury: We will explore this question in SMP8-cre-ERT2 mice crossed with homozygous loxP-Igfl mice. Injury experiments will be done after tamoxifen-induced deletion of the IgfI alleles and depletion of local IGF-I tissue stores. 3) Determine the requirement for local IGFBP expression in smooth muscle response to injury: We will first explore this by targeted deletion of lGFBP-4 in smooth muscle cells in a tamoxifen-dependent manner. We will test the hypothesis that SMP8-cre-ERT2/loxP-lgfbp-4 mice will have diminished smooth muscle tissue stores of IGF-I, and that this will be associated with impaired responses to carotid artery injury. 4) Determine the requirement for local expression of the zinc metalloproteinase PAPP-A in smooth muscle responses to injury: Here we will test the hypothesis that after tamoxifen-induced recombination of floxed PAPP-A, IGFBP protease activity will be abolished or dampened, IGFBP-4 stabilized, resulting in IGF-I sequestration and impaired SMC migration and proliferation.