Abstract
BackgroundA functional blood supply is essential for tumor growth and proliferation. However, the mechanism of blood vessel recruitment to the tumor is still poorly understood. Ideally, a thorough molecular assessment of blood vessel cells would be critical in our comprehension of this process. Yet, to date, there is little known about the molecular makeup of the endothelial cells of tumor-associated blood vessels, due in part to the difficulty of isolating a pure population of endothelial cells from the heterogeneous tissue environment.MethodsHere we describe the use of a recently developed technique, Expression Microdissection, to isolate endothelial cells from the tumor microenvironment. The methylation status of the dissected samples was evaluated for GSTP1 and RARβ2 promoters via the QMS-PCR method.ResultsComparing GSTP1 and RARβ2 promoter methylation data, we show that 100% and 88% methylation is detected, respectively, in the tumor areas, both in epithelium and endothelium. Little to no methylation is observed in non-tumor tissue areas.ConclusionWe applied an accurate microdissection technique to isolate endothelial cells from tissues, enabling DNA analysis such as promoter methylation status. The observations suggest that epigenetic alterations may play a role in determining the phenotype of tumor-associated vasculature.
Highlights
A functional blood supply is essential for tumor growth and proliferation
The molecular study of the tumor microenvironment has taken a dramatic turn. It has been shown in three different tissue types, breast, colon and prostate, that the cellular microenvironment surrounding the tumor epithelium demonstrates similar epigenetic changes to those found in the tumor cells [1,2,3]
While epigenetic changes have been identified in stroma, fibroblasts, and the colorectal mucosa, there has not been an evaluation of tumor-associated endothelial cells [1,2,3]
Summary
A functional blood supply is essential for tumor growth and proliferation. the mechanism of blood vessel recruitment to the tumor is still poorly understood. While epigenetic changes have been identified in stroma, fibroblasts, and the colorectal mucosa, there has not been an evaluation of tumor-associated endothelial cells [1,2,3] This is due in part to the difficulty in obtaining a pure population of endothelial cells from tissues for analysis. Other methods for isolating endothelial cells, such as antibody labeled magnetic beads and fluorescence activated cell sorting (FACS), require large amounts of fresh tissue and long processing times [4]. Such approaches may in themselves alter the endothelial cells, making it difficult to assess the baseline characteristics that were present in the tumor microenvironment
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