TNF Alpha in PKC Epsilon Signaling to Carcinogenesis

PKC Epsilon 信号传导中的 TNF Alpha 致癌作用

基本信息

批准号：
8893904
负责人：
AJIT K VERMA
金额：
$ 26.33万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-04-12 至 2016-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8893904
关键词：
Acetates Affect Alleles Biological Assay Bromodeoxyuridine CD34 gene Cell Proliferation Cell Separation Cell surface Cells Chronic Colony-Stimulating Factor Gene Crossbreeding Cutaneous Development EMSA Family Gene Expression Gene Silencing Genes Genetic Transcription Granulocyte Colony-Stimulating Factor Granulocyte-Macrophage Colony-Stimulating Factor Hair follicle structure Human Inflammation Inflammatory Integrins Knock-out Knockout Mice Label Link Luciferases MAP Kinase Gene Malignant - descriptor Mediating Mus Neoplasm Metastasis Papilloma Phospholipids Play Predisposition Protein Isoforms Protein-Serine-Threonine Kinases Proteins Protocols documentation Radiation therapy Reporter Genes Reporting Role Signal Transduction Signal Transduction Pathway Skin Skin Cancer Skin Carcinogenesis Skin Neoplasms Squamous cell carcinoma Stem cells Stress TNF gene Techniques Testing Transgenic Mice Tumor Necrosis Factor-alpha Tumor Promoters Tumor Promotion UV Radiation Exposure Ultraviolet Rays carcinogenesis cytokine dimethylbenzanthracene human TNF protein in vivo knockout gene novel overexpression precursor cell protein kinase C epsilon receptor transcription factor

项目摘要

DESCRIPTION (provided by applicant): The objectives of this revised renewal proposal (RO1 CA 102431A2) are to determine whether protein kinase C epsilon (PKC5) signals ultraviolet radiation (UVR)-induced development of squamous cell carcinoma (SCC) via increased expression of the pro-inflammatory cytokines tumor necrosis factor alpha (TNF1) and granulocyte colony-stimulating factor (G-CSF) which may directly promote the proliferation of hair follicle stem cells, the targets of SCC. Chronic exposure to UVR is the most common etiological factor linked to the development of SCC, a nonmelanoma form of skin cancer that can metastasize. PKC5 is among the six isoforms (1, 4, 5, 7, 5, 6) expressed in both human and mouse skin. To determine the in vivo functional role of PKC5 in mouse skin carcinogenesis, we generated PKC5 transgenic mouse (FVB/N) lines 224 and 215 that overexpress approximately 8- and 18-fold respectively PKC5 protein over endogenous levels in basal epidermal cells and cells of the hair follicle. We reported that epidermal PKC5 level dictates the susceptibility of transgenic mice to the development of SCC elicited either by the repeated exposure to UVR or by the DMBA - TPA tumor promotion protocol. Our studies to elucidate mechanisms of PKC5-mediated development of SCC, either by DMBA-TPA or UVR, indicated a common converging point- elevated release of TNF1 and G-CSF. To conclusively determine whether TNF1 is essential for the development of SCC in PKC5 transgenic mice, we generated TNF1-knockout//PKC5 transgenic mice by crossbreeding TNF1 knockout mice with PKC5 transgenic mice. Deletion of both TNF1 alleles in PKC5 transgenic mice only partially inhibited (50%, p = 0.0136) the development of SCC by the DMBA-TPA protocol. Deletion of the TNF1 gene in PKC5 transgenic mice also partially suppressed UVR-induced skin damage and inflammation. These results indicate that, in addition to TNF1, other UVR-induced cytokines such as G-CSF may play essential roles in the development of SCC. We hypothesize that: 1) TNF1 cooperates with G-CSF in PKC5 signal transduction pathways to the development of SCC by UVR, 2) PKC5-induced TNF1 and G-CSF mediate induction of SCC through direct effects on stem cells in the mouse hair follicle and 3) PKC5 interacts with MAPK cascade to regulate UVR- induced TNF1 and G-CSF gene transcription. We propose three specific aims to test these hypotheses: Specific aim #1: To determine, using gene knockout approach, whether both TNF1 and G-CSF contribute to the development of SCC in PKC5 transgenic mice by UVR. Specific Aim# 2: To define, in intact mouse skin in vivo, whether the roles of TNF1 and G-CSF in UVR-induced development of SCC involve the activation and proliferation of putative hair follicle stem cell. Hair follicle stem cell proliferation will be determined by quantitations of label retaining cells (LRCs) using BrdU and FACS analysis using cell surface markers (CD34, 16-integrin). Specific Aim #3: To define the mechanism, using gene-silencing techniques, by which PKC5 signals UVR-induced TNF1 and G-CSF gene expression.

描述（由申请人提供）：此修订后的更新建议的目标（RO1 CA 102431A2）是确定蛋白激酶C epsilon（PKC5）是否信号是否通过增加鳞状细胞癌（SCC）的发育来确定紫罗兰辐射（UVR）的发育是否通过增加pro-inflamm inflamm inflamm andn andn andn andn andn andn andn andn andn andn andn andn andn andn andn andn andn。粒细胞刺激因子（G-CSF），可以直接促进毛囊干细胞的增殖，即SCC的靶标。长期暴露于UVR是与SCC发展的最常见病因因素，SCC的发展是一种可以转移的皮肤癌。 PKC5是在人和小鼠皮肤中表达的六种同工型（1、4、5、7、5、6）。为了确定PKC5在小鼠皮肤癌发生中的体内功能作用，我们生成了PKC5转基因小鼠（FVB/N）线224和215，在基底表皮细胞和毛状细胞的基础表皮细胞和细胞中，在内源性水平上过度表达了大约8倍和18倍PKC5蛋白。我们报告说，表皮PKC5水平决定了转基因小鼠对通过反复接触UVR或DMBA -TPA肿瘤促进方案而引起的SCC发展的敏感性。我们的研究阐明了通过DMBA-TPA或UVR的PKC5介导的SCC发育的机制，表明TNF1和G-CSF的常见融合点升高。为了最终确定TNF1是否对于PKC5转基因小鼠的SCC是否至关重要，我们通过用PKC5转基因小鼠通过杂交TNF1敲除小鼠产生了TNF1-KNOCKOUT // PKC5转基因小鼠。 PKC5转基因小鼠中两个TNF1等位基因的删除仅部分抑制了DMBA-TPA方案的SCC的发展（50％，P = 0.0136）。 PKC5转基因小鼠中TNF1基因的缺失也部分抑制了UVR诱导的皮肤损伤和炎症。这些结果表明，除TNF1外，其他UVR诱导的细胞因子（例如G-CSF）可能在SCC的发展中起重要作用。我们假设：1）TNF1在PKC5信号转导途径中与G-CSF合作，通过UVR开发SCC的发展，2）PKC5诱导的TNF1和G-CSF通过对小鼠毛囊中的干细胞和3）PKC Casce cascce casce uvr-casce uvr-tone to s介导SCC介导SCC的诱导，并介导了SCC，并介导了SCC。转录。我们提出了三个特定的目的来检验这些假设：具体目的＃1：使用基因敲除方法确定TNF1和G-CSF是否有助于通过UVR在PKC5转基因小鼠中的SCC发展。特定目的＃2：在体内完整的小鼠皮肤中定义，TNF1和G-CSF在UVR诱导的SCC发育中的作用是否涉及假定的毛囊干细胞的激活和增殖。毛囊干细胞增殖将通过使用BRDU和FACS分析使用细胞表面标记（CD34，16-整合素）来确定标记固定细胞（LRC）的定量。特定目的＃3：使用基因分解技术定义机制，PKC5信号UVR诱导的TNF1和G-CSF基因表达。