ENTEROCYTE APOPTOSIS AFTER INTESTINAL RESECTION

描述(申请人提供)：肠适应是大量小肠切除(SBR)后的一种关键的代偿性反应，其特征是以增殖和凋亡的升高来衡量肠细胞的周转。在这笔赠款的前一个资金周期中，当细胞凋亡被主动抑制时，观察到了放大的适应，表明了细胞凋亡对适应程度的重要性。虽然SBR后细胞凋亡增加的机制(S)目前尚不清楚，但我们已经确定这种反应受表皮生长因子受体(EGFR)信号调节，需要细胞凋亡原Bcl02家族成员BAX和转录因子信号转导和转录激活因子(STAT)-1的表达。作为这些关键观察的延伸，我们提出了一个全球假设，即EGFR信号调节Bax的表达和活性，以调节切除诱导的细胞凋亡。为了验证这一假说，我们的目的是：1)确定STAT-1在肠道适应过程中调节Bax表达的作用。STAT-1的表达和活性将在SBR后的回肠和诱导细胞凋亡后的细胞培养中进行测定。将检测STAT-1缺陷对Bax表达和细胞凋亡的影响，并将研究Bax启动子上可能的STAT-1结合位点。2)确定p38α丝裂原活化蛋白激酶(MAPK)在手术诱导适应过程中对Bax活性的调节作用。在小鼠SBR后，p38表达/活性和Bax激活的时空分布将被记录下来，并补充体外细胞凋亡模型。SBR后将确定有条件的、肠道特异性的p38表达缺失对Bax活性和细胞凋亡的影响。3)确定EGFR调节Bax表达和活性的机制。增强或干扰EGFR信号对STAT-1和p38激活和表达的影响将被记录下来。在EGFR抑制的背景下，减弱的STAT-1或p38的表达对细胞凋亡和Bax的活性和表达的影响将被确定。这项应用中提出的研究将确定大规模SBR后最相关的直接细胞凋亡的信号通路。彻底了解诱导细胞凋亡的精确机制是将治疗靶点转移到床边的基础，这些治疗靶点旨在最大限度地刺激肠粘膜的再生，以应对大量的肠道丢失。与公共卫生相关：在大量肠道丢失后，剩余的肠道试图重新生长以补偿。如果这种反应不完全，患者将接受终生静脉营养和所有相关并发症。该项目旨在了解粘膜细胞死亡在肠道再生过程中的作用。了解这一重要反应的确切分子调控可能会导致旨在改善肠道灾难性丢失后肠道再生长的创新治疗。

DESCRIPTION (provided by applicant): Intestinal adaptation is a critical, compensatory response to massive small bowel resection (SBR) and characterized by increased enterocyte turnover as gauged by elevated rages of both proliferation and apoptosis. The significance of apoptosis to the magnitude of adaptation was revealed during the previous funding cycle of this grant as amplified adaptation was observed when apoptosis was actively inhibited. While the mechanisms(s) for elevated apoptosis after SBR is presently unknown, we have established that this response is regulated by epidermal growth factor receptor (EGFR) signaling and requires expression of the proapototic Bcl02 family member Bax and the transcription factor signal transducer and activator of transcription (STAT)-1. As an extension of these key observations, we propose the global hypothesis that EGFR signaling modulates the expression and activity of Bax to regulate resection-induced apoptosis. To test this hypothesis our aims are: 1) Determine the role for STAT-1 in the regulation of Bax expression during intestinal adaptation. STAT-1 expression and activity will be determined in the ileum after SBR as well as in cell culture following induction of apoptosis. The effect of STAT-1 deficiency on Bax expression and apoptosis will be measured and putative STAT-1 binding sites on Bax promoter will be investigated. 2) Determine the role of p38alpha mitogen-activated protein kinase (MAPK) as modulator of Bax activity during resection induced adaptation. A temporal and spatial profile of p38 expression/activity and Bax activation will be recorded after SBR in mice and complementary in vitro models of apoptosis. The effect of conditional, intestine-specific deletion of p38 expression on Bax activity and apoptosis will be determined after SBR. 3) Determine the mechanism for EGFR regulation of Bax expression and activity. The effect of enhanced or disrupted EGFR signaling on STAT-1 and p38 activation and expression will be recorded. The effect of attenuated STAT-1 or p38 expression in the context of EGFR inhibition on apoptosis and Bax activity and expression will be determined. The proposed studies in this application will identify the most relevant signaling pathway to direct apoptosis after massive SBR. A thorough understanding of the precise mechanism for induction of apoptosis is fundamental for bench-to-bedside translation of therapeutic targets intended to maximally stimulate regrowth of the intestinal mucosa in response to massive intestinal loss. PUBLIC HEALTH RELEVANCE: Following massive intestinal loss, the remaining bowel attempts to grow back to compensate. If this response is incomplete, the patient will be subjected to a lifetime of nutrition by vein and all the associated complications. This project is designed to understand the contribution of mucosal cell death to the process of intestinal regrowth. Understanding the exact molecular regulation of this important response may lead to innovative therapy intended to improve intestinal regrowth following a catastrophic loss of the intestine.