The relationship between decreased BMD and increased risk of fractures is well established

genesis and the expression of angiogenesis regulators: CXCL1, and THBS1 were both altered by co-culture. Thus we evaluated expression changes for CXCL1 and THBS1 and as well as for CXCL6, another angiogenesis related chemokine. In the main computational analysis, changes in expression for CXCL6 did not pass multiple hypothesis correction, but based on its relevance to angiogenesis and nearly significant pvalue of 0.053 it was included in the analysis. PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19672638 For all three genes, conditioned media treatments recapitulated the gene expression changes predicted from the microarray results indicating that their regulation was through the actions of secreted factors. CXCL6 expression was observed in both HBMECs PDE7B is frequently upregulated in GBM and has prognostic significance To establish overall PDE7B expression in GBM, we first examined two independent publically accessible gene expression datasets. Using Oncomine we found datasets by both Murat et al and Sun et al showing that PED7B expression was significantly greater in GBM compared to normal brain controls. We next verified PDE7B over expression by qRTPCR in 18 of the 22 primary GBM specimens. Fold change was calculated as: for genes with increased expression upon co-culture, or as: 2) for genes downregulated. doi:10.1371/journal.pone.0107397.t001 expression using data from The Cancer Genome Atlas and molecular subtype-characteristic centroid expression profiles as previously described by us. PDE7B expression was significantly greater in the Classical, compared to Mesenchymal or Proneural subtypes of GBM. There was also a significant difference between PDE7B expression in Mesenchymal and Neural subtypes of PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19672212 GBM. Together these data indicate that PDE7B mRNA expression is increased in GBM and that PDE7B expression may distinguish between subtypes of disease. To determine whether DMXB-A levels of PDE7B expression are prognostic in GBM we first examined the National Cancer Institute’s Rembrandt Database for all grades of astrocytoma. Stratification of all astrocytomas based on PDE7B expression two-fold above or below the median level of expression revealed a striking correlation with survival. While too few cases with PDE7B expression greater than two-fold above the median were available for survival analysis, PDE7B expression 2-fold lower than the median was correlated with significantly increased survival compared to median levels of expression . The effect of PDE7B on survival was distinct from that of two other cAMP specific phosphodiesterases evaluable in the Rembrandt Database. Expression of PDE4A and PDE8B at two-fold lower than the median was correlated with worse survival, while expression of PDE8B at two-fold higher than the median was associated with better survival, compared to cases with median levels of expression. This analysis suggests there may be some specificity to the actions of PDE7B in GBM. We confirmed the effect of PDE7B expression on survival in the Murat et al dataset where patients with GBM alive at 5-years showed significantly lower levels of expression than patients who had died of disease. Overexpression of PDE7B stimulates in vitro stemness and in vivo GBM growth and tumorigenicity To directly test whether the level of PDE7B expression affects GBM biology, we cloned and overexpressed PDE7B in U87-GL cells and a low passage primary GBM cell line, G144. As a control, we engineered a catalytically inactive form of PDE7B. Catalytically inactive forms of other cAMP specifi