The Human Cutaneous Squamous Cell Carcinoma Microenvironment Is Characterized by Increased Lymphatic Density and Enhanced Expression of Macrophage-Derived VEGF-C

Abstract

Metastases from primary cutaneous squamous cell carcinoma (SCC) account for the majority of the ∼10,000 non-melanoma skin cancer deaths in the United States annually. We studied lymphangiogenesis in human SCC because of the potential link to metastasis. SCC samples were stained for lymphatic endothelial vessel marker LYVE-1 and positive cells were counted and compared with cells in normal skin. Gene set enrichment analysis and reverse transcription (RT)-PCR were performed on SCC, on adjacent non-tumor-bearing skin, and on normal skin to determine the differential expression of lymphangiogenesis-associated genes. Laser capture microdissection (LCM) was performed to isolate tumor cells and tumor-associated inflammatory cells for further gene expression analysis. Immunofluorescence was performed to determine the source of vascular endothelial growth factor-C (VEGF-C) in the tumor microenvironment. We found increased lymphatic density and reorganized lymphatic endothelial vessels in the dermis immediately adjacent to SCC nests. RT-PCR confirmed the presence of VEGF-C in skin immediately adjacent to SCC. LCM confirmed the increased expression of VEGF-C, the SCC inflammatory infiltrate. The presence of CD163(+)/CD68(+)/VEGFC(+) cells and absence of VEGF-C expression by CD3(+) or CD11C(+) cells suggested that VEGF-C is derived from tumor-associated macrophages. Clarification of mechanisms governing SCC-mediated lymphangiogenesis may identify potential targets for therapeutic intervention against aggressive or inoperable disease.

VEGF-C and VEGF-D Blockade Inhibits Inflammatory Skin Carcinogenesis

“…Activation of angiogenesis facilitates tumor growth by provision of oxygen and nutrients, whereas activation of lymphangiogenesis facilitates tumor dissemination (1). Angiogenesis and lymphangiogenesis are induced in human cutaneous SCCs (20,21). This is accompanied by increased expression of genes associated with lymphangiogenesis, such as VEGF-C, Nrp-2, and VEGFR-3, in peritumoral skin (20).…”

Interaction of tumor cells and lymphatic vessels in cancer progression

“…Recently, the role of the tumor microenvironment also has been emphasized by description of tumor-associated macrophages that may function as a second source of lymphangiogenic factors. 68,69 In the majority of clinical studies, a significant correlation has been observed between lymphatic vessel density and lymph node and organ metastasis (see meta-analyses in refs 70 --72). This has led to the concept that the denser the lymphatic vasculature is within or close to the tumor, the more potential entry sites the tumor cells have to vessels that could be used as highways for metastatic spread.…”

Tumour–stroma crosstalk in the development of squamous cell carcinoma

“…Tumour associated macrophages (TAMs) in turn express growth factors and matrix-proteases, promote angiogenesis and suppress adaptive immunity even while helping the host to inhibit tumour growth and destroy neoplastic cells (Mantovani, Bottazzi et al 1992;Sica, Allavena et al 2008). For instance, VEGF-C produced by TAMs in cSCC leads to lymphangiogenesis which then results in increased lymphatic vessel density (Belkin, Mitsui et al 2011), which in turn is linked to metastatic progression (Belkin, Mitsui et al 2011;Moussai, Mitsui et al 2011).…”