Glial subcellular re-sealed particles (referred to as gliosomes here) were purified from rat cerebral cortex and investigated for their ability to release glutamate. Confocal microscopy showed that the glia-specific proteins glial fibrillary acidic protein (GFAP) and S-100, but not the neuronal proteins 95-kDa postsynaptic density protein (PSD-95), microtubule-associated protein 2 (MAP-2) and b-tubulin III, were enriched in purified gliosomes. Furthermore, gliosomes exhibited labelling neither for integrin-aM nor for myelin basic protein, which are specific for microglia and oligodendrocytes respectively. The Ca 2+ ionophore ionomycin (0. The role of glia in the brain is an area of intense investigation. In the past decade, exciting results in this field have led to dramatic conceptual changes about the role of glial cells, which were formerly thought to provide only structural and trophic support to neurones. An increasing number of papers have suggested that glia share at least some of the features typical of neurones, particularly those concerned with excitatory neurotransmission (for a review see Haydon 2001). In fact, glial cells are

ClinicalTrials.gov Identifier: NCT01397136.

BackgroundNeuroblastoma is the most common, pediatric, extra-cranial, malignant solid tumor. Despite multimodal therapeutic protocols, outcome for children with a high-risk clinical phenotype remains poor, with long-term survival still less than 40%. Hereby, we evaluated the potential of non-coding RNA expression to predict outcome in high-risk, stage 4 neuroblastoma.MethodsWe analyzed expression of 481 Ultra Conserved Regions (UCRs) by reverse transcription-quantitative real-time PCR and of 723 microRNAs by microarrays in 34 high-risk, stage 4 neuroblastoma patients.ResultsFirst, the comparison of 8 short- versus 12 long-term survivors showed that 54 UCRs were significantly (P < 0.0491) over-expressed in the former group. For 48 Ultra Conserved Region (UCRs) the expression levels above the cut-off values defined by ROC curves were strongly associated with good-outcome (OS: 0.0001

In assisted reproduction technology, embryo competence is routinely evaluated on morphological criteria. Over the last decade, efforts in improving non-invasive embryo assessment have looked into the secretome of embryos. Human embryos release genomic DNA (gDNA) and mitochondrial DNA (mtDNA) into the culture medium, and the mtDNA/gDNA ratio is significantly correlated with embryo fragmentation. Here, we investigate whether mtDNA/gDNA ratio in embryo spent medium is correlated with blastulation potential and implantation. The mtDNA/gDNA ratio was assessed in 699 Day 3 culture media by quantitative polymerase chain reaction (qPCR) to investigate its correlation with embryo morphology, blastocyst development and implantation. A logistic regression model evaluated whether mtDNA/gDNA ratio in the secretome may improve the prediction of blastulation. We found that embryos that successfully developed into blastocysts exhibited a significantly higher mtDNA/gDNA ratio in the culture medium compared with those that arrest (P = 0.0251), and mtDNA/gDNA, combined with morphological grading, has the potential to predict blastulation better than morphology alone (P = 0.02). Moreover, mtDNA/gDNA ratio was higher in the media from good-quality embryos that reached the full blastocyst stage on Day 5 compared with those that developed more slowly (P < 0.0001). With respect to blastocyst morphology, higher trophectoderm quality was associated with a higher mtDNA/gDNA ratio in the culture medium. Finally, a high mtDNA/gDNA ratio in spent medium was associated with successful implantation outcome (P = 0.0452) of good-quality embryos. In summary, the mtDNA/gDNA ratio in the Day 3 embryo secretome, in combination with morphological grading, may be a novel, non-invasive, early biomarker to improve identification of viable embryos with high developmental potential.

The oncogenic Bcr-Abl tyrosine kinase activates various signaling pathways including phosphoinositide 3-kinase/ Akt and nuclear factor-KB that mediate proliferation, transformation, and apoptosis resistance in Bcr-Abl(+) myeloid leukemia cells. The hop flavonoid xanthohumol inhibits tumor growth by targeting the nuclear factor-KB and Akt pathways and angiogenesis. Here, we show that xanthohumol has in vitro activity against Bcr-Abl(+) cells and clinical samples and retained its cytotoxicity when imatinib mesylate -resistant K562 cells were examined. Xanthohumol inhibition of K562 cell viability was associated with induction of apoptosis, increased p21 and p53 expression, and decreased survivin levels. We show that xanthohumol strongly inhibited Bcr-Abl expression at both mRNA and protein levels and show that xanthohumol caused elevation of intracellular reactive oxygen species and that the antioxidant N-acetylcysteine blunted xanthohumol-induced events. Further, we observed that xanthohumol inhibits leukemia cell invasion, metalloprotease production, and adhesion to endothelial cells, potentially preventing in vivo life-threatening complications of leukostasis and tissue infiltration by leukemic cells. As structural mutations and/or gene amplification in Bcr-Abl can circumvent an otherwise potent anticancer drug such as imatinib, targeting Bcr-Abl expression as well as its kinase activity could be a novel additional therapeutic approach for the treatment of Bcr-Abl(+) myeloid leukemia. [Mol Cancer Ther 2008;7(9):2692 -702]

About 50% of children with neuroblastoma (NB) show a metastatic disease and have a poor prognosis. However, disease progression is greatly variable and depends on patients' age and MYCN oncogene amplification. To investigate the role of patients' age in tumor aggressiveness, we performed array-CGH and gene expression profiles of three groups (G) of metastatic NBs: G1, stage 4S patients and MYCN single copy (MYCN-) tumors; G2, stage 4 patients, ≤18 months of age, MYCN- tumors and favorable outcome and G3, Stage 4 patients, a19 months with unfavorable outcome. G1 was characterized by numerical aberrations prevalently; on the contrary, all G3 tumors had structural rearrangements, whereas G2 showed an intermediate pattern. The average of numerical alterations decreased significantly from G1 to G2 to G3 (p < 0.01). Contrarily, the number of structural aberrations increased from G1 to G2 to G3 (p< 2.35 E-05). Noteworthy, G3/MYCN- NBs were characterized by several complex intrachromosome rearrangements. Expression analysis of the three groups showed significant differences in genes of Rho and Ras signaling pathways, development and adhesion, cell cycle regulation and telomerase activity. Accumulation of structural alterations increased with patients' age and was associated with a more aggressive disease. Abnormal expression of genes involved in cell cycle and telomerase in G3 may be responsible for the genomic instability in this cohort of patients. The higher DNA instability observed in G3/MYCN- NBs than in MYCN-amplified G3 may also explain why patients a19 months have a poor outcome independently by MYCN status. Copyright © 2012 UICC

Metastases in the bone marrow (BM) are grim prognostic factors in patients with neuroblastoma (NB). In spite of extensive analysis of primary tumor cells from high- and low-risk NB patients, a characterization of freshly isolated BM-infiltrating metastatic NB cells is still lacking. Our aim was to identify proteins specifically expressed by metastatic NB cells, that may be relevant for prognostic and therapeutic purposes. Sixty-six Italian children over 18 months of age, diagnosed with stage 4 NB, were included in the study. Metastatic NB cells were freshly isolated from patients' BM by positive immunomagnetic bead manipulation using anti-GD2 monoclonal antibody. Gene expression profiles were compared with those obtained from archived NB primary tumors from patients with 5y-follow-up. After validation by RT-qPCR, expression/secretion of the proteins encoded by the up-regulated genes in the BM-infiltrating NB cells was evaluated by flow cytometry and ELISA. Compared to primary tumor cells, BM-infiltrating NB cells down-modulated the expression of CX3CL1, AGT, ATP1A2 mRNAs, whereas they up-regulated several genes commonly expressed by various lineages of BM resident cells. BM-infiltrating NB cells expressed indeed the proteins encoded by the top-ranked genes, S100A8 and A9 (calprotectin), CD177 and CD3, and secreted the CXCL7 chemokine. BM-infiltrating NB cells also expressed CD271 and HLA-G. We have identified proteins specifically expressed by BM-infiltrating NB cells. Among them, calprotectin, a potent inflammatory protein, and HLA-G, endowed with tolerogenic properties facilitating tumor escape from host immune response, may represent novel biomarkers and/or targets for therapeutic intervention in high-risk NB patients.

BACKGROUND.Neuroblastic tumors (NTs) are largely comprised of neuroblastic (Nb) cells with various quantities of Schwannian stromal (SS) cells. NTs show a variable genetic heterogeneity. NT gene expression profiles reported so far have not taken into account the cellular components. The authors reported the genome‐wide expression analysis of whole tumors and microdissected Nb and SS cells.METHODS.The authors analyzed gene expression profiles of 10 stroma‐poor NTs (NTs‐SP) and 9 stroma‐rich NTs (NTs‐SR) by microarray technology. Nb and SS cells were isolated by laser microdissection from NTs‐SP and NTs‐SR and probed with microarrays. Gene expression data were analyzed by the Significance Analysis of Microarrays (SAM) and Game Theory (GT) methods, the latter applied for the first time to microarray data evaluation.RESULTS.SAM identified 84 genes differentially expressed between NTs‐SP and NTs‐SR, whereas 50 were found by GT. NTs‐SP mainly express genes associated with cell replication, nervous system development, and antiapoptotic pathways, whereas NTs‐SR express genes of cell‐cell communication and apoptosis. Combining SAM and GT, the authors found 16 common genes driving the separation between NTs‐SP and NTs‐SR. Five genes overexpressed in NTs‐SP encode for nuclear proteins (CENPF, EYA1, PBK, TOP2A, TFAP2B), whereas only 1 of 11 highly expressed genes in NTs‐SR encodes for a nuclear receptor (NR4A2).CONCLUSIONS.The results showed that NT‐SP and NT‐SR gene signatures differ for a set of genes involved in distinct pathways, and the authors demonstrated a low intratumoral heterogeneity at the mRNA level in both NTs‐SP and NTs‐SR. The combination of SAM and GT methods may help to better identify gene expression profiling in NTs. Cancer 2008. © 2008 American Cancer Society.