The role of emergency granulopoiesis in the pathogenesis of Fanconi Anemia

Fanconi途径通过同源的DNA复制叉修复崩溃和停滞的DNA复制叉。 重组修复（HRR）和跨损伤合成（TLS）;从而在S 细胞周期的阶段。先天性缺乏任何Fanconi蛋白导致Fanconi贫血（FA）; 其特征在于早期阶段的骨髓衰竭（BMF）。假设BMF发生为 未修复的DNA损伤触发FA造血干细胞（HSC）和祖细胞的凋亡。 在FA的BMF期存活的患者有发生骨髓发育不良的趋势， 进展这被假设是由累积突变引起的，这些突变诱导细胞周期抵抗 检查点和/或对DNA损伤的凋亡反应。 FA患者表现出稳态粒细胞减少症和感染易感性。我们的研究表明， 在感染性攻击期间受损的“紧急粒细胞生成”（EG）也有助于免疫缺陷， FA.在正常情况下，EG相关的细胞因子刺激粒细胞立即释放， 骨髓，然后扩增HSC和粒细胞/单核细胞祖细胞（GMP）。这种增殖 阶段涉及S阶段缩短。我们发现Fanconi C和F在原发性肝癌中表达增加， 用IL 1和其他介导EG反应的细胞因子处理的鼠GMP。我们确定， FancC缺陷型小鼠不能产生体内EG应答。相反，我们发现， 在大多数FancC-/-小鼠中，EG-刺激导致全血细胞减少、BMF和死亡。我们还发现 在FancC-/-小鼠中EG刺激的副作用被IL 1-R拮抗剂阻断。 我们假设，反复的，失败的紧急粒细胞生成事件加速骨髓 失败在FA而且，不成功的EG事件为导致克隆的突变提供了机会。 进展这一假设将通过三个目标来实现： 目的1：明确EG过程中Fanconi通路激活和DNA修复的机制。重量，FancC- 缺乏或FancA缺乏的鼠骨髓细胞将用EG相关细胞因子处理并分析 Fanconi途径激活和DNA修复。将使用各种EG刺激进行体内研究。 目的2：明确FA中EG相关骨髓衰竭的分子机制。我们还将 使用上述模型来确定EG相关的细胞因子是否在Fan缺陷的骨中诱导细胞凋亡 骨髓与细胞周期检查点激活相关。 目的3：确定多次失败的EG发作是否促进FA的克隆进展：我们将使用这些 模型，以确定是否阻断特定的细胞因子防止重复EG发作期间的克隆进展。 这些研究的目的是确定无效EG发作在BMF中的作用， FA.这些研究可能会提出可以迅速转化为临床的治疗方法。

The Fanconi pathway repairs collapsed and stalled DNA-replication forks through homologous recombination repair (HRR) and translesional synthesis (TLS); thereby maintaining genomic integrity during S phase of the cell cycle. Congenital absence of any Fanconi protein results in Fanconi Anemia (FA); a disorder that is characterized during early stages by bone marrow failure (BMF). BMF is hypothesized to occur as unrepaired DNA damage triggers apoptosis in FA hematopoietic stem cells (HSC) and progenitor cells. Patients who survive the BMF stage of FA have a tendency to develop bone marrow dysplasia with clonal progression. This is hypothesized to be caused by accumulating mutations that induce resistance to cell cycle checkpoints and/or the apoptotic response to DNA-damage. FA patients exhibit steady state granulocytopenia and susceptibility to infection. Our studies suggest that impaired "emergency granulopoiesis" (EG) during infectious challenge also contributes to immuno-deficiency in FA. Under normal circumstances, EG-related cytokines stimulate immediate granulocyte release from the bone marrow, followed by expansion of HSC and granulocyte/monocyte progenitors (GMP). This proliferative phase involves S phase-shortening. We found that expression of Fanconi C and F increased in primary murine GMP treated with IL1¿ and other cytokines that mediate the EG response. And, we determined that FancC deficient mice are unable to mount an in vivo EG-response. Instead, we found that repeated episodes of EG-stimulation result in pancytopenia, BMF, and death in the majority of FancC-/- mice. We also found that the adverse effects of EG stimulation in FancC-/- mice are blocked by an IL1-R antagonist. We hypothesize that repeated, failed episodes of emergency granulopoiesis accelerate bone marrow failure in FA. And, that unsuccessful EG episodes provide opportunity for mutations that result in clonal progression. This hypothesis will be pursued through three aims: AIM 1: Define mechanisms of Fanconi pathway activation and DNA-repair during EG. Wt, FancC- deficient, or FancA-deficient murine bone marrow cells will be treated with EG-related cytokines and analyzed for Fanconi pathway activation and DNA-repair. In vivo studies will be performed with various EG stimuli. AIM 2: Identify molecular mechanisms involved in EG-related bone marrow failure in FA. We will also use the models described above to determine if EG-related cytokines induce apoptosis in Fanc-deficient bone marrow in association with cell cycle checkpoint activation. AIM 3: Determine if multiple failed episodes of EG facilitate clonal progression in FA: We will use these models to determine if blocking specific cytokines prevents clonal progression during repeated EG episodes. The goal of these studies is to define the role of ineffective EG episodes in BMF and clonal progression in FA. These studies may suggest therapeutic approaches that could be rapidly translated to the clinic.