Enterocyte Apoptosis after Intestinal Resection

在大量小肠切除（SBR）后，剩余的肠道通过一个称为适应的关键过程进行补偿，其中包括肠细胞增殖、绒毛高度、肠口径和长度的增加。也许是为了抵消这种有丝分裂反应，也观察到肠细胞凋亡率增加。初步研究表明，促凋亡因子bax的表达增加和促生存因子bcl-w的表达减少参与了肿瘤切除术后细胞凋亡的调控。表皮生长因子（EGF）通过增加肠细胞增殖来增强适应性。虽然其意义尚不清楚，但EGF也能抑制切除后肠细胞凋亡。本项目将验证EGF通过抑制肠细胞凋亡增强大面积小肠切除后肠道适应性的整体假设。该项目的长期目标是通过描述肠细胞凋亡在肠道适应中的意义来优化短肠综合征患者的治疗。我们的具体目标是：1)确定EGF受体刺激和抑制对适应过程中bax和bcl-w基因表达和凋亡的影响。为了验证EGF受体信号调节bax和bcl-w的表达以指导肠细胞在适应过程中凋亡的假设，我们将使用激光捕获显微解剖（LCM）显微镜来建立SBR后肠隐窝-绒毛轴上bax和bcl-w mRNA和蛋白在时间和区域上的肠细胞特异性表达。然后在EGF受体刺激和抑制条件下，在SBR后LCM提取的肠细胞中测定肠细胞中bax和bcl-w的表达。2)确定EGF减少肠细胞凋亡的机制。为了验证EGF需要bax和bcl-w调控术后凋亡的假设，我们将对bax和bcl-w敲除小鼠进行SBR或假手术。杂交实验将允许直接测试在EGF受体导向的扩增或受损适应条件下缺乏bax或bcl-w表达的影响。3)通过EGFR信号转导确定Bax和Bcl-w表达的机制。我们将开发一个体外模型系统，使用肠上皮细胞来测试相关的EGFR信号转导途径，以激活bax和bcl-w的表达。这些研究将极大地促进对适应的遗传调控以及细胞凋亡与这一过程的相关性的理解。这一信息对于未来安全有效的临床治疗的发展是至关重要的，旨在扩大这种重要的代偿反应在遭受大量肠道损失的患者中。

After massive small bowel resection (SBR), the remaining intestine compensates by a critical process termed adaptation, which includes increases in enterocyte proliferation, villus height, bowel caliber and length. Perhaps to counterbalance this mitogenic response, increased rates of enterocyte apoptosis are also observed. Preliminary studies suggest that increased expression of the pro-apoptosis factor bax and reduced expression of the prosurvival factor bcl-w are involved in the regulation of postresection apoptosis. Epidermal growth factor (EGF) enhances adaptation by increasing enterocyte proliferation. While its significance is not known, EGF also inhibits postresection enterocyte apoptosis. This project will test that overall hypothesis that EGF enhances intestinal adaptation after massive small bowel resection by inhibiting enterocyte apoptosis. The long-term goal of this project is to optimize therapy for patients with the short bowel syndrome by delineating the significance of enterocyte apoptosis during intestinal adaptation. Our specific aims are: 1) Determine the effect of EGF receptor stimulation and inhibition on bax and bcl-w gene expression and apoptosis during adaptation. To test the hypothesis that EGF receptor signaling regulates the expression of bax and bcl-w to direct enterocyte apoptosis during adaptation, laser capture microdissection (LCM) microscopy will be used to establish a temporal and regional, enterocyte-specific expression of bax and bcl-w mRNA and protein along the crypt- villus axis after SBR. The expression of bax and bcl-w in enterocytes will then be determined in LCM extracted enterocytes after SBR during conditions of EGF receptor stimulation and inhibition. 2) Determine the mechanism for reduced enterocyte apoptosis by EGF. To test the hypothesis that bax and bcl-w are required for the regulation of postresection apoptosis by EGF, SBR or sham operations will be performed in bax and bcl-w knockout mice. Crossbreeding experiments will permit direct testing of the effect of absent bax or bcl-w expression during conditions of EGF receptor-directed amplified or impaired adaptation. 3) Determine the mechanism for Bax and Bcl-w expression by EGFR signal transduction. We will develop an in vitro model system using intestinal epithelial cells to test relevant EGFR signal transduction pathways for the activation of bax and bcl-w expression. These studies will substantially contribute toward an enhanced understanding of the genetic regulation of adaptation and the relevance of apoptosis to this process. This information is vital for the future development of safe and effective clinical therapy designed to amplify this important compensatory response in patients suffering massive intestinal loss.