Elevated expression of TGF‐β1 in head and neck cancer–associated fibroblasts

Abstract

Head and neck cancers are characterized by a vigorous desmoplastic response, but the contribution of stromal-derived growth factors to the tumor microenvironment is poorly understood. We evaluated the expression of stromal growth factor expression in head and neck squamous cell carcinoma (HNSCC) in normal and tumor-associated stromal cells. Stromal tissue was isolated from epithelial cells with laser capture microdissection (LCMD) and analyzed by cDNA array for the expression of TGFalpha, TGF-beta1, HGF, PDGF-alpha, IGFII, bFGF, aFGF, VEGFC, and VEGF. Primary fibroblasts were isolated in vitro from HNSCC tumors, adjacent histologically normal mucosa, and skin in vitro. Fibroblast populations were assessed for TGF-beta1 expression by ELISA and luciferase reporter assay to assess protein expression. We identified TGF-beta1 and IGFII overexpression in normal and tumor-associated stromal cells; however, only TGF-beta1 was significantly overexpressed (3.4-fold) in tumor-associated stroma. Assessment of carcinoma-associated fibroblasts (CAFs), normal dermal fibroblasts (NDFs), and normal mucosal fibroblasts (NMFs) in propagated fibroblasts demonstrated persistently elevated levels of TGF-beta1 in CAFs compared to NMF and NDF populations. Elevated levels of TGF-beta1 were identified in the stromal compartment of HNSCC tumors compared to normal mucosa by immunohistochemical analysis. These results suggest that TGF-beta1 mRNA and protein is specifically upregulated in CAFs in vitro and in vivo.

Examination of Oral Cancer Biomarkers by Tissue Microarray Analysis

“…42 The situation in OSCC is less well understood, but studies of carcinoma-associated fi- broblasts, extracellular matrix turnover, and tumor cell motility have begun to delineate the role of desmoplastic stroma in OSCC carcinogenesis. [43][44][45] Recently, Weber et al 46 performed genome-wide analysis of loss of heterozygosity and allelic imbalance on specimens of tumor stroma and tumor epithelium isolated by laser capture microdissection from more than 120 patients with OSCC and a history of smoking. They discovered more than 40 hot spots of loss of heterozygosity/allelic imbalance within the stroma, nearly twice as many as they found in the epithelium, and subsequently identified 3 stroma-specific loci that were significantly associated with tumor size and cervical lymph node metastasis.…”

Identification of Two Distinct Carcinoma-Associated Fibroblast Subtypes with Differential Tumor-Promoting Abilities in Oral Squamous Cell Carcinoma

“…The importance of TGF-b1 for CAF activation was confirmed by demonstration of significant deeper invasion occurring when TGF-b1-pretreated NOF were embedded in 3D constructs and seeded on top with transformed oral keratinocytes, as compared with matched controls. Most likely, inflammatory cells that are present in vivo in the tumor stroma provide TGF-b1 that initiates this process (42), but over time, the change in the transcriptome, with increased expression of many TGF-b1 target genes, leads to emergence of the nonmotile (41), high TGF-b1-secreting fibroblast subtype (14). By correlating Transwell motility assay with secretome analysis, we found this subset of high TGF-b1 secretors to be present at significantly higher proportions in CAF-D strains.…”

“…However, CAF were shown to show phenotypic and genotypic diversity (4,10,11), as well as complex changes in their secretory activity (12)(13)(14), but the functional significance of this heterogeneity has been poorly addressed so far. Only recently, the essential role of this heterogeneity and the cooperation between different subpopulations was shown for prostate tumorigenesis, with the differential TGF-b signaling in tumor stromal cells indicated to be pivotal (15).…”

Emdogain® in carcinogenesis: a systematic review of in vitro studies

“…Furthermore, EMD enhances the production of TGF-β1 in many different cell lines, including osteoblasts, fibroblasts, and malignant osteosarcoma cells (39). TGF-β1 has broad regulatory functions, but it has also become clear that TGF-β1 can act as one of the principal cytokines in carcinogenesis (90,91). During tumor progression, TGF-β1 and other cytokines (those in the epidermal growth factor (EGF) families, for example) enhance the morphological and invasive phases of the EMT phenotype (92).…”