Abstract
CXCL12 is a CXC-type chemokine that plays important roles in hematopoiesis, development, and organization of the immune system and supports the survival or growth of a variety of normal or malignant cell types. Our laboratory recently showed that CXCL12 is secreted by aging stromal fibroblast cells and is a major paracrine factor that specifically stimulates the proliferation of prostate epithelial cells. The current study shows that this CXCL12-mediated proliferative response may be either ERK-dependent or ERK-independent. Moreover, CXCL12 initiates a previously undefined and complex global transcriptional response in prostate epithelial cells. This CXCL12-mediated transcriptional response directly stimulates the expression of genes encoding proteins that are involved in the promotion of cellular proliferation and progression through the cell cycle, tumor metastasis, and cellular motility, and directly represses the transcription of genes encoding proteins involved in cell-cell adhesion and resistance to apoptosis. Thus, CXCL12 may play a major role in the etiology of benign proliferative disease in the context of an aging tissue microenvironment.
Highlights
Cells plated in triplicate at 50,000 cells/well were pre-treated with 0.05% Me2SO or 0.05% Me2SO and 5 M U0126 (Cell Signaling) for 2 h, stimulated with CXCL12 at concentrations shown previously to induce EGR1 gene transcription, e.g. 10 pM CXCL12 for N15C6 cells and 50 pM CXCL12 for LNCaP cells
These results suggest that CXCL12-mediated EGR1 gene transcription is more ERKdependent in N15C6 than LNCaP cells
CXCL12 Stimulates a Global Transcriptional Response in Prostate Epithelial Cells That Is Partially ERK-dependent—We had previously reported that both non-transformed N15C6 and transformed LNCaP prostate epithelial cells respond proliferatively to low, picomolar levels of CXCL12, similar to those secreted by aging human prostate stromal fibroblasts [6]
Summary
Cell Cultures—N15C6 cells, developed as described previously, were used at passages 35– 45 and were maintained in 5% HIE media (Ham’s F-12 (Mediatech Inc., Herndon, VA) with. Experiments utilizing U0126 were performed as above, except that cells were pre-treated for 2 h with either 0.05% Me2SO (vehicle) or 0.05% Me2SO plus 5 M U0126 before the addition of CXCL12. Cells were maintained in media containing appropriate antibodies for the entirety of the experiment, and cells were counted at 24 and 96 h as above. Normalized array-acquired transcript expression values were analyzed using a t-statistic test and by calculation of -fold change between data sets. Genes that exhibited both a large t-statistic (Ͼ10.0) and a large -fold change (Ͼ2.0) were considered to be differentially expressed. All other data were assessed by t test or analysis of variance with p Ͻ 0.05 considered statistically significant
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