Mapping of oxidative stress responses of human tumor cells following photodynamic therapy using hexaminolevulinate

<p>Abstract</p> <p>Background</p> <p>Photodynamic therapy (PDT) involves systemic or topical administration of a lesion-localizing photosensitizer or its precursor, followed by irradiation of visible light to cause singlet oxygen-induced damage to the affected tissue. A number of mechanisms seem to be involved in the protective responses to PDT, including activation of transcription factors, heat shock proteins, antioxidant enzymes and apoptotic pathways.</p> <p>Results</p> <p>In this study, we address the effects of a destructive/lethal hexaminolevulinate (HAL) mediated PDT dose on the transcriptome by using transcriptional exon evidence oligo microarrays. Here, we confirm deviations in the steady state expression levels of previously identified early defence response genes and extend this to include unreported PDT inducible gene groups, most notably the metallothioneins and histones. HAL-PDT mediated stress also altered expression of genes encoded by mitochondrial DNA (mtDNA). Further, we report PDT stress induced alternative splicing. Specifically, the ATF3 alternative isoform (deltaZip2) was up-regulated, while the full-length variant was not changed by the treatment. Results were independently verified by two different technological microarray platforms. Good microarray, RT-PCR and Western immunoblotting correlation for selected genes support these findings.</p> <p>Conclusion</p> <p>Here, we report new insights into how destructive/lethal PDT alters the transcriptome not only at the transcriptional level but also at post-transcriptional level via alternative splicing.</p

Phospholipase C Isozymes Are Deregulated in Colorectal Cancer – Insights Gained from Gene Set Enrichment Analysis of the Transcriptome

Colorectal cancer (CRC) is one of the most common cancer types in developed countries. To identify molecular networks and biological processes that are deregulated in CRC compared to normal colonic mucosa, we applied Gene Set Enrichment Analysis to two independent transcriptome datasets, including a total of 137 CRC and ten normal colonic mucosa samples. Eighty-two gene sets as described by the Kyoto Encyclopedia of Genes and Genomes database had significantly altered gene expression in both datasets. These included networks associated with cell division, DNA maintenance, and metabolism. Among signaling pathways with known changes in key genes, the “Phosphatidylinositol signaling network”, comprising part of the PI3K pathway, was found deregulated. The downregulated genes in this pathway included several members of the Phospholipase C protein family, and the reduced expression of two of these, PLCD1 and PLCE1, were successfully validated in CRC biopsies (n = 70) and cell lines (n = 19) by quantitative analyses. The repression of both genes was found associated with KRAS mutations (P = 0.005 and 0.006, respectively), and we observed that microsatellite stable carcinomas with reduced PLCD1 expression more frequently had TP53 mutations (P = 0.002). Promoter methylation analyses of PLCD1 and PLCE1 performed in cell lines and tumor biopsies revealed that methylation of PLCD1 can contribute to reduced expression in 40% of the microsatellite instable carcinomas. In conclusion, we have identified significantly deregulated pathways in CRC, and validated repression of PLCD1 and PLCE1 expression. This illustrates that the GSEA approach may guide discovery of novel biomarkers in cancer

Cluster of spliceosome complex genes that were significantly altered in response to HAL-PDT

<p><b>Copyright information:</b></p><p>Taken from "Mapping of oxidative stress responses of human tumor cells following photodynamic therapy using hexaminolevulinate"</p><p>http://www.biomedcentral.com/1471-2164/8/273</p><p>BMC Genomics 2007;8():273-273.</p><p>Published online 13 Aug 2007</p><p>PMCID:PMC2045114.</p><p></p> Horizontal stripes represent genes and columns show treatment protocols. The log2-fold changes of gene ratios are color coded as shown in the bar. The three different columns represent repeated microarray experiments

Verification of MYC, ATF3 and JUN expression after HAL-PDT by Western blotting

<p><b>Copyright information:</b></p><p>Taken from "Mapping of oxidative stress responses of human tumor cells following photodynamic therapy using hexaminolevulinate"</p><p>http://www.biomedcentral.com/1471-2164/8/273</p><p>BMC Genomics 2007;8():273-273.</p><p>Published online 13 Aug 2007</p><p>PMCID:PMC2045114.</p><p></p> Cell extracts were prepared at at 1 h, 2 h and 4 h after HAL-PDT (A). Since PDT affected the expression of well-known house keeping proteins such actin (induced) and tubulin (repressed) the total protein content was measured and the gels were stained after electrophoresis, thus normalize against total protein content. The fold-changes obtained by the microarray analysis for the selected genes are presented for comparison. Fold-change of three repeated experiments presented as the mean ± St. Dev (B)

Venn diagrams of differentially altered genes/probes in response to HAL-PDT

<p><b>Copyright information:</b></p><p>Taken from "Mapping of oxidative stress responses of human tumor cells following photodynamic therapy using hexaminolevulinate"</p><p>http://www.biomedcentral.com/1471-2164/8/273</p><p>BMC Genomics 2007;8():273-273.</p><p>Published online 13 Aug 2007</p><p>PMCID:PMC2045114.</p><p></p> A, includes up-regulated and down-regulated; B, only up-regulated; C, only down-regulated genes/probes at 1, 2 and 4 h time course. Each circle represents one time point as indicated. A total number of differentially expressed genes in the group are given in the parentheses. The intersections indicate numbers of genes/probe that are shared between the different time points, i.e. the genes that were steady induced. The number in the right bottom of each square indicates a total post processed and analyzed number of genes/probes minus the number of differentially expressed genes presented in circles

Improved prognostication of glioblastoma beyond molecular subtyping by transcriptional profiling of the tumor microenvironment

Glioblastoma (GBM), the most aggressive form of brain cancer, is characterized by a high level of molecular heterogeneity, and infiltration by various immune and stromal cell populations. Important advances have been made in deciphering the microenvironment of GBMs, but its association with existing molecular subtypes and its potential prognostic role remain elusive. We have investigated the abundance of infiltrating immune and stromal cells in silico, from gene expression profiles. Two cohorts, including in-house normal brain and glioma samples (n = 70) and a large sample set from TCGA (n = 393), were combined into a single exploratory dataset. A third independent cohort (n = 124) was used for validation. Tumors were clustered based on their microenvironment infiltration profiles, and associations with known GBM molecular subtypes and patient outcome were tested a posteriori in a multivariable setting. We identified a subset of GBM samples with significantly higher abundances of most immune and stromal cell populations. This subset showed increased expression of both immune suppressor and immune effector genes compared to other GBMs and was enriched for the mesenchymal molecular subtype. Survival analyses suggested that tumor microenvironment infiltration pattern was an independent prognostic factor for GBM patients. Among all, patients with the mesenchymal subtype with low immune and stromal infiltration had the poorest survival. By combining molecular subtyping with gene expression measures of tumor infiltration, the present work contributes with improving prognostic models in GBM

Epigenetic disruption of miR-130a promotes prostate cancer by targeting SEC23B and DEPDC1

MicroRNAs (miRNAs) are small, non-coding RNAs that mediate post-transcriptional gene silencing, fine tuning gene expression. In an initial screen, miRNAs were found to be globally down-regulated in prostate cancer (PCa) cell lines and primary tumors. Exposure of PCa cell lines to a demethylating agent, 5-Aza-CdR resulted in an increase in the expression levels of miRNAs in general. Using stringent filtering criteria miR-130a was identified as the most promising candidate and selected for validation analyses in our patient series. Down-regulation of miR-130a was associated with promoter hypermethylation. MiR-130a methylation levels discriminated PCa from non-malignant tissues (AUC=0.956), and urine samples revealed high specificity for non-invasive detection of patients with PCa (AUC=0.89). Additionally, repressive histone marks were also found in the promoter of miR-130a. Over-expression of miR-130a in PCa cells reduced cell viability and invasion capability, and increased apoptosis. Putative targets of miR-130a were assessed by microarray expression profiling and DEPD1C and SEC23B were selected for validation. Silencing of both genes resembled the effect of over-expressing miR-130a in PCa cells. Our data indicate that miR-130a is an epigenetically regulated miRNA involved in regulation of key molecular and phenotypic features of prostate carcinogenesis, acting as a tumor suppressor miRNA.info:eu-repo/semantics/publishedVersio