Glucocorticoids and glucocorticoid receptor translational isoforms

说明(申请人提供)：糖皮质激素在治疗炎症中是不可或缺的，尽管它们在治疗部分患者的中性粒细胞炎症方面无效。我们的长期目标是了解糖皮质激素受体(GR)介导细胞特异性功能的机制，因为这些信息可能有助于开发具有改善疗效/风险比的抗炎治疗。本研究的目的是确定GR翻译异构体在中性粒细胞存活和功能中的作用。我们最近发现GR有八种翻译异构体，它们对基因表达和细胞功能有非常不同的影响。我们假设选择性GR翻译异构体介导中性粒细胞对糖皮质激素诱导的细胞凋亡的抵抗。我们建议通过改变GR亚型来提高糖皮质激素对抗中性粒细胞炎症的疗效。为了验证我们的假设，我们将在小鼠哮喘模型中检测Lenti病毒转导的人CD34细胞来源的中性粒细胞、小鼠骨髓来源的中性粒细胞和中性粒细胞。目标1的研究将确定GR-A亚型是否会增加中性粒细胞对糖皮质激素诱导的凋亡的敏感性。我们发现初级中性粒细胞主要有GR-D亚型。在多个白细胞中，GR-A，而不是GR-D，是促凋亡的。我们的初步数据还表明，在中性粒细胞中，维甲酸(RA)将GR-D转换为-A亚型。如果表达GR-A的中性粒细胞对糖皮质激素敏感，则糖皮质激素可能有效地抑制中性粒细胞的气道炎症。目的2确定翻译因子如异源核糖核蛋白L(hnRNP-L)是否调节中性粒细胞GR异构体的选择性表达和糖皮质激素敏感性。我们已经确定hnRNP-L是GR-D相关的翻译因子。在早幼粒细胞HL-60细胞中，hnRNP-L被敲除后，GR-D亚型转变为-A亚型。我们将在体外和体内测试在中性粒细胞中敲除hnRNP-L是否会改变中性粒细胞的糖皮质激素反应。这些研究将提高我们对GR翻译异构体在中性粒细胞炎症中的作用的理解。GR-A亚型有望增强糖皮质激素诱导中性粒细胞凋亡和抑制中性粒细胞炎症成分的能力。

DESCRIPTION (provided by applicant): Glucocorticoids are indispensable in the treatment of inflammation although they are not effective in treating neutrophilic inflammation in subsets of patients. Our long-term goal is to understand the mechanisms by which the glucocorticoid receptor (GR) mediates cell-specific functions, as this information may be useful in the development of anti-inflammatory treatments with improved efficacy/risk ratios. The goal of this proposal is to determine the role of GR translational isoforms in neutrophil survival and functions. We have recently discovered that the GR has eight translational isoforms that have profoundly different effects on gene expression and cell functions. We hypothesize that selective GR translational isoforms mediate neutrophil resistance to glucocorticoid-induced apoptosis. We propose to increase the efficacy of glucocorticoids in circumventing neutrophilic inflammation by altering GR isoforms. To test our hypotheses, we will examine lenti virus-transduced human CD34+ cell-derived neutrophils, mouse bone marrow-derived neutrophils, and neutrophils in a murine asthma model. Studies in Aim 1 will determine whether the GR-A isoform will increase the sensitivity of neutrophils to glucocorticoid induced-apoptosis. We have found that primary neutrophils have predominantly the GR-D isoforms. In multiple leukocytes, the GR-A, but not the GR-D, isoform is proapoptotic. Our preliminary data also show that retinoic acid (RA) switches the GR-D to the -A isoform in neutrophils. If GR-A expressing neutrophils are sensitive to glucocorticoids, glucocorticoids may be effective in inhibiting neutrophilic airway inflammation. Aim 2 will determine whether translation factors such as heterogenous nuclear ribonucleoprotein L (hnRNP-L) regulate the selective expression of GR isoforms and glucocorticoid sensitivity of neutrophils. We have identified hnRNP-L as a GR-D associated translation factor. Knockdown of hnRNP-L in promyelocytic HL-60 cells switched the GR-D to -A isoform. We will test whether knockdown of hnRNP-L in neutrophils alter neutrophil glucocorticoid responses in vitro and in vivo. These studies will improve our understanding of the role of GR translational isoforms in neutrophilc inflammation. The GR-A isoform is anticipated to increase the ability of glucocorticoids to induce neutrophil apoptosis and to inhibi components of neutrophilic inflammation.