Wnt-3a-activated human fibroblasts promote human keratinocyte proliferation and matrix destruction.
Aberrant Wnt regulation, detectable by nuclear translocation of beta-catenin, is a hallmark of many cancers including skin squamous cell carcinomas (SCCs). By analyzing primary human skin SCCs, we demonstrate that nuclear beta-catenin is not restricted to SCC cells but also detected in stromal fibroblasts, suggesting an important role for aberrant Wnt regulation also in the tumor microenvironment. When human keratinocytes and fibroblasts were treated with Wnt-3a, fibroblasts proved to be more responsive. Accordingly, Wnt-3a did not alter HaCaT cell functions in a cell-autonomous manner. However, when organotypic cultures (OTCs) were treated with Wnt-3a, HaCaT keratinocytes responded with increased proliferation. As nuclear beta-catenin was induced only in the fibroblasts, this argued for a Wnt-dependent, paracrine keratinocyte stimulation. Global gene expression analysis of Wnt-3a-stimulated fibroblasts identified genes encoding interleukin-8 (IL-8) and C-C motif chemokine 2 (CCL-2) as well as matrix metalloproteinase-1 (MMP-1) as Wnt-3a targets. In agreement, we show that IL-8 and CCL-2 were secreted in high amounts by Wnt-3a-stimulated fibroblasts also in OTCs. The functional role of IL-8 and CCL-2 as keratinocyte growth regulators was confirmed by directly stimulating HaCaT cell proliferation in conventional cultures. Most important, neutralizing antibodies against IL-8 and CCL-2 abolished the Wnt-dependent HaCaT cell hyperproliferation in OTCs. Additionally, MMP-1 was expressed in high amounts in Wnt-3a-stimulated OTCs and degraded the stromal matrix. Thus, our data show that Wnt-3a stimulates fibroblasts to secrete both keratinocyte proliferation-inducing cytokines and stroma-degrading metalloproteinases, thereby providing evidence for a novel Wnt deregulation in the tumor-stroma directly contributing to skin cancer progression.
Genetics of Skin Carcinogenesis, German Cancer Research Center (DKFZ), Im Neuenheimer Feld 280, 69120, Heidelberg, Germany.
Katrin
K
Sobel
Marius
M
Tham
Hans-Jürgen
HJ
Stark
Hermann
H
Stammer
Silke
S
Prätzel-Wunder
Jackie R
JR
Bickenbach
Petra
P
Boukamp
D002294:Carcinoma, Squamous Cell / Q000235:genetics / Q000473:pathology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D018932:Chemokine CCL2 / Q000235:genetics / Q000378:metabolism; D005109:Extracellular Matrix / Q000187:drug effects* / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D015151:Immunoblotting; D007376:Interleukin-2 / Q000235:genetics / Q000378:metabolism; D015603:Keratinocytes / Q000187:drug effects* / Q000378:metabolism; D020781:Matrix Metalloproteinase 1 / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009924:Organ Culture Techniques; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012878:Skin Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D060449:Wnt Signaling Pathway / Q000187:drug effects / Q000235:genetics; D060509:Wnt3A Protein / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*
D018932:Chemokine CCL2; D007376:Interleukin-2; D060509:Wnt3A Protein; D020781:Matrix Metalloproteinase 1
Wnt/beta-catenin signaling; organotypic cultures; squamous cell carcinoma; tumor microenvironment