Stromal cells are increasingly recognized as playing an essential role in tissue regenerative and reparative processes. In the intestine, a rich population of connective tissue and immune cells supports the function and architecture of the continuously renewing epithelium. Disruption of epithelial homeostasis, such as occurs with tissue injury or in cancer, leads to the recruitment and activation of myeloid cells of the innate immune system. These play diverse roles in tissue defense and repair and in certain instances contribute to disease pathogenesis. This is particularly true in colorectal cancer. Recent work has shown that the transition of benign adenomatous polyps to adenocarcinoma is accompanied by an increase in the number of infiltrating immune cells. Furthermore, colorectal cancer survival has been linked to both macrophage and neutrophil density within the tumors. How immune cells regulate cancer progression is poorly understood, but there is good evidence that these cells can contribute to cancer cell growth, invasion and metastasis. The goal of this proposal is to study immune cell function in an in vivo, zebrafish cell invasion model that shares important features with currently accepted models of human cancer cell invasion that were developed using in vitro systems. This proposal consists of two specific aims. The goal of the first aim is to define the innate immune cell population in the intestine of wild type zebrafish larvae and myh11-meltdown mutants that develop epithelial cell invasion in response to unregulated smooth muscle myosin function. Immune cells will be localized in vivo using transgenic lines that express fluorescent reporter genes under the control of established myeloid cell promoters. Additionally, the use of fluorescent reporters and specific labeling dyes will be used to assess the activation pathways upregulated in polarized immune cells. Time-lapse recordings will be used to study interactions between the immune cells and the invasive epithelial cells. Genetic loss-of-function studies will be used to assess what role if any these cells play in the cell invasion phenotype. Comparable studies will be performed in a transgenic line that drives formation of epithelial cell invadopodia but that does not support stromal invasion of the epithelial cells, thus allowing immune cells analyses at specific stages of the invasive process. The goal of the second aim will be to perform transcriptional profiling of innate immune cells in the two zebrafish models, and to correlate changes in gene expression with myeloid cell subtypes, particularly those linked to cancer progression as well as functionally assay the importance of differentially regulated genes to the invasion process. Collectively, these studies will enhance our understanding of how immune cells regulate tissue architecture in the intestine. Furthermore, they may lead to the identification of markers of novel innate immune cell subtypes that have prognostic significance in colorectal cancer.

Public Health Relevance

Colorectal cancer is the third leading cause of cancer-related deaths in the U.S. A key stage in tumor progression, invasion, is poorly understood, but may be promoted by cells of the innate immune system. The goal of this proposal is to identify the innate immune cell types that infiltrate sites of invasion, and the evolving functional and molecular phenotypes that occur in these cells to promote invasiveness.

Agency
National Institute of Health (NIH)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
1F32DK101166-01A1
Application #
8716846
Study Section
Special Emphasis Panel (ZDK1)
Program Officer
Podskalny, Judith M,
Project Start
Project End
Budget Start
Budget End
Support Year
1
Fiscal Year
2014
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
City
Philadelphia
State
PA
Country
United States
Zip Code
19104