Regulation of Lipid Droplets and FOXO3 in Intestinal Epithelial Cell Proliferatio

描述(申请人提供)：肥胖和超重在过去20年里一直是发达国家的大流行病，与一系列疾病有关，包括结肠癌等不同器官的癌症。到目前为止，人们还不完全了解这些关联的基本机制。一种新出现的可能性是，肥胖者组织中积累的过量脂肪直接刺激结肠肿瘤的进展。细胞内的脂质储存在脂肪组织和许多人类肿瘤中发现的动态细胞器--脂滴中。人们认为LDS是驱动增殖的能量来源，而增殖是肿瘤进展的标志；然而，他们的调节和功能还不是很清楚。我们的初步数据显示，在肥胖小鼠的结肠以及发炎和肿瘤的结肠组织中，LD的积累增加。此外，LDS受到肿瘤抑制因子FOXO_3的负调控，而刺激LDS积聚会导致FOXO_3活性丧失。因此，我们提出了一个LD-FOXO_3网络的存在，它还包括PI3K和SIRT6(甘油三酯(TGS)的负调控因子)。此外，初步数据表明，LD-FOXO_3网络在结肠癌细胞的增殖中起关键作用，也在肥胖影响结肠上皮细胞中发挥作用。 支持LD-FOX03网络上游受EGFR和/或PGE2控制，而下游调节细胞周期调节和甘油三酯合成。该方案的中心假设是，在LD-FOXO_3网络控制下的结肠细胞的增殖导致协调LD的积累，依赖FOXO_3缺失的细胞周期进程是肥胖相关的结肠肿瘤进展的机制之一。目的1.评估LD诱导的结肠癌细胞增殖反应是否依赖于FOXO_3的活性：(A)EGFR的激活以及这是否依赖于PGE_2，(B)依赖PI3K的FOXO_3细胞周期停滞的丧失，以及(C)结肠癌细胞中TGS的负调控因子SIRT6依赖。目的2.确定在体外，肿瘤坏死因子诱导的细胞增殖和随之而来的LD积聚是否是由于EGFR、PGE2的激活以及FOXO_3细胞周期停滞和SIRT6的丢失所致，以及(B)确定在经肿瘤坏死因子处理的FOXO_3中是否选择性地阻断LD 或EGFR缺陷小鼠通过促进Foxo3依赖的SIRT6、细胞周期停滞和TGS而抑制结肠上皮细胞的增殖。目的3.评估肥胖和Foxo3缺乏是否会加剧DSS/AOM模型中腺瘤的进展，导致G0-G1检查点和TGS调节因子的改变，(B)选择性阻断LD、PGE2或TNF是否抑制腺瘤进展，以及(C)确定调节脂代谢的Foxo3依赖基因。拟议研究的结果将对人类健康产生重大的积极影响，因为LD-FOXO_3网络将成为一种新的治疗靶点。虽然结肠癌进展的机制很复杂，但在肥胖促进结肠癌进展的患者中，LDS的抑制可能提供一种有效的肿瘤进展阻断。

DESCRIPTION (provided by applicant): Obesity and overweight, which have been immense epidemics in developed countries for the last two decades, are associated with a wide array of diseases including cancer of different organs such as the colon. To date, the underlying mechanisms for these associations are not completely understood. One of emerging possibilities is that excess lipids accumulated in the tissue of obese individuals directly stimulates progression of colonic tumors. Intracellular lipids are stored in lipid droplets (LDs), dynamic organelles found in adipose tissue, as well as in many human tumors. It is believed that LDs are the source of energy that drives proliferation, a hallmark of tumor progression; however, their regulation and function are not well understood. Our preliminary data reveals that LD accumulation is increased in the colon of obese mice as well as in inflamed and neoplastic colonic tissue. Moreover, LDs are negatively regulated by tumor suppressor FOXO3, while stimulating LDs accumulation leads to a loss of FOXO3 activity. Thus we propose the existence of an LD-FOXO3 network that also includes PI3K and SIRT6 (a negative regulator of triglycerides (TGs). Moreover, preliminary data suggested that the LD-FOXO3 network is critical in proliferation of colonic cancer cells and also operates in obesity influence colonic epithelium. There is support that the LD-FOXO3 network is upstream control by EGFR and/or PGE2, while downstream modifies cell cycle regulators and triglyceride synthesis. The central hypothesis of this proposal is that proliferation in colonic cells controlled by LD-FOXO3 network leads to coordinate LD accumulation and cell cycle progression dependent on loss of FOXO3 is one mechanism responsible for obesity related colon tumor progression. The following Specific Aims are proposed: Aim 1. To assess if the LD induced proliferative response in colonic cancer cells depends on FOXO3 activity via (a) activation of EGFR and whether this is dependent on PGE2, (b) PI3K dependent loss of FOXO3 cell cycle arrest, and (c) SIRT6 dependent, a negative regulator of TGs in colon cancer cells. Aim 2. To determine if TNF-induced proliferation and consequent LD accumulation are due to activation of EGFR, PGE2 and a loss of FOXO3 cell cycle arrest and SIRT6 (in vitro), and (b) to find if selective blockade of LD in TNF treated Foxo3 or EGFR deficient mice suppresses proliferation of colonic epithelia by promoting Foxo3 dependent SIRT6 and cell cycle arrest and TGs. Aim 3. To assess if obesity and a deficiency in Foxo3 (a) exacerbated adenoma progression in DSS/AOM model leading to alterations in regulators of G0-G1 checkpoint and TGs, (b) if adenoma progression is suppressed by selective blockade of LD, PGE2, or TNF, and (c) to identify the Foxo3 dependent genes regulating lipid metabolism. Findings from the proposed study would have significant positive effects on human health by presenting the LD-FOXO3 network as a novel therapeutic target. Although mechanisms of colon cancer progression are complex, the inhibition of LDs may provide an effective blockade to tumor progression in patients with obesity promoted colonic tumor progression.