Novel chemotherapeutic agent FX-9 activates NF-κB signaling and induces G1 phase arrest by activating CDKN1A in a human prostate cancer cell line.

BACKGROUND: The aminoisoquinoline FX-9 shows pro-apoptotic and antimitotic effects against lymphoblastic leukemia cells and prostate adenocarcinoma cells. In contrast, decreased cytotoxic effects against non-neoplastic blood cells, chondrocytes, and fibroblasts were observed. However, the actual FX-9 molecular mode of action is currently not fully understood. METHODS: In this study, microarray gene expression analysis comparing FX-9 exposed and unexposed prostate cancer cells (PC-3 representing castration-resistant prostate cancer), followed by pathway analysis and gene annotation to functional processes were performed. Immunocytochemistry staining was performed with selected targets. RESULTS: Expression analysis revealed 0.83% of 21,448 differential expressed genes (DEGs) after 6-h exposure of FX-9 and 0.68% DEGs after 12-h exposure thereof. Functional annotation showed that FX-9 primarily caused an activation of inflammatory response by non-canonical nuclear factor-kappa B (NF-κB) signaling. The 6-h samples showed activation of the cell cycle inhibitor CDKN1A which might be involved in the secondary response in 12-h samples. This secondary response predominantly consisted of cell cycle-related changes, with further activation of CDKN1A and inhibition of the transcription factor E2F1, including downstream target genes, resulting in G1-phase arrest. Matching our previous observations on cellular level senescence signaling pathways were also found enriched. To verify these results immunocytochemical staining of p21 Waf1/Cip1 (CDKN1A), E2F1 (E2F1), PAI-1 (SERPNE1), and NFkB2/NFkB p 100 (NFKB2) was performed. Increased expression of p21 Waf1/Cip1 and NFkB2/NFkB p 100 after 24-h exposure to FX-9 was shown. E2F1 and PAI-1 showed no increased expression. CONCLUSIONS: FX-9 induced G1-phase arrest of PC-3 cells through activation of the cell cycle inhibitor CDKN1A, which was initiated by an inflammatory response of noncanonical NF-κB signaling.

Stratification of clear cell renal cell carcinoma (ccRCC) genomes by gene-directed copy number alteration (CNA) analysis.

Tumorigenic processes are understood to be driven by epi-/genetic and genomic alterations from single point mutations to chromosomal alterations such as insertions and deletions of nucleotides up to gains and losses of large chromosomal fragments including products of chromosomal rearrangements e.g. fusion genes and proteins. Overall comparisons of copy number alterations (CNAs) presented in 48 clear cell renal cell carcinoma (ccRCC) genomes resulted in ratios of gene losses versus gene gains between 26 ccRCC Fuhrman malignancy grades G1 (ratio 1.25) and 20 G3 (ratio 0.58). Gene losses and gains of 15762 CNA genes were mapped to 795 chromosomal cytoband loci including 280 KEGG pathways. CNAs were classified according to their contribution to Fuhrman tumour gradings G1 and G3. Gene gains and losses turned out to be highly structured processes in ccRCC genomes enabling the subclassification and stratification of ccRCC tumours in a genome-wide manner. CNAs of ccRCC seem to start with common tumour related gene losses flanked by CNAs specifying Fuhrman grade G1 losses and CNA gains favouring grade G3 tumours. The appearance of recurrent CNA signatures implies the presence of causal mechanisms most likely implicated in the pathogenesis and disease-outcome of ccRCC tumours distinguishing lower from higher malignant tumours. The diagnostic quality of initial 201 genes (108 genes supporting G1 and 93 genes G3 phenotypes) has been successfully validated on published Swiss data (GSE19949) leading to a restricted CNA gene set of 171 CNA genes of which 85 genes favour Fuhrman grade G1 and 86 genes Fuhrman grade G3. Regarding these gene sets overall survival decreased with the number of G3 related gene losses plus G3 related gene gains. CNA gene sets presented define an entry to a gene-directed and pathway-related functional understanding of ongoing copy number alterations within and between individual ccRCC tumours leading to CNA genes of prognostic and predictive value.

Network analysis of transcriptional regulation in response to intramuscular interferon-ß-1a multiple sclerosis treatment.

Interferon-ß (IFN-ß) is one of the major drugs for multiple sclerosis (MS) treatment. The purpose of this study was to characterize the transcriptional effects induced by intramuscular IFN-ß-1a therapy in patients with relapsing-remitting form of MS. By using Affymetrix DNA microarrays, we obtained genome-wide expression profiles of peripheral blood mononuclear cells of 24 MS patients within the first 4 weeks of IFN-ß administration. We identified 121 genes that were significantly up- or downregulated compared with baseline, with stronger changed expression at 1 week after start of therapy. Eleven transcription factor-binding sites (TFBS) are overrepresented in the regulatory regions of these genes, including those of IFN regulatory factors and NF-κB. We then applied TFBS-integrating least angle regression, a novel integrative algorithm for deriving gene regulatory networks from gene expression data and TFBS information, to reconstruct the underlying network of molecular interactions. An NF-κB-centered sub-network of genes was highly expressed in patients with IFN-ß-related side effects. Expression alterations were confirmed by real-time PCR and literature mining was applied to evaluate network inference accuracy.

Putative biomarker genes for grading clear cell renal cell carcinoma.

OBJECTIVE: The initial objective of this renal cancer study was to identify gene sets in clear cell renal cell carcinoma (ccRCC) to support grading of ccRCC histopathology. MATERIALS AND METHODS: Preselected ccRCC tumor tissues of grade 1 (G1, n = 14) and grade 3 (G3, n = 15) as well es 14 normal kidney tissues thereof were subjected to microarray expression analysis using Human Genome U133 Plus 2.0 Array. Event ratio scoring, hierarchical clustering and principal component analysis were used to determine gene sets that distinguish expression profiles from normal kidney tissue, G1 and G3 tumor tissues. RESULTS: An initial set of 73 genes provided seven gene subclusters (SC01 to SC07) that distinguish RNA expression profiles from G1, G3 tumor and normal kidney tissues. A ranked list of 24 genes was determined that separated G1 from G3 tumors in high concordance with histopathological grading confirmed by immunohistochemical analysis of ceruloplasmin protein expression. CONCLUSION: A final set of 24 genes has been determined awaiting further validation on the RNA as well as on the protein level by studying an additional cohort of ccRCC patients. A reliable separation of G1 and G3 tumor grades will be instrumental to foster and direct the administration of upcoming targeted therapeutics of ccRCC tumors in a more predictive and reliable manner.

Gene expression analysis in clear cell renal cell carcinoma using gene set enrichment analysis for biostatistical management.

OBJECTIVE: To improve the workflow for standardizing the statistical interpretation provides an opportunity for the analysis of gene expression in clear cell renal cell carcinoma (ccRCC). RCC as a solid tumour entity represents a very suitable tumour model for such investigations. Although it is possible to investigate expression profiles by microarray technologies, the main problem is how to adequately interpret the accumulated mass of data derived from microarray technologies. There is a clear lack of a defined, consistent and comparable biostatistical analysis system, with no specific biostatistical standard methodology being available to compare the results of microarray analyses. We used the gene set enrichment analysis (GSEA) method to analyze microarray data from RCC tissue. The present study aimed to analyze differential expression profiles and establish biomarkers suitable for prognostication at the time of renal surgery by comparing RCC patients with long-term survival data against RCC samples of patients with poorly differentiated (grade 3) RCC, concomitant metastatic disease and short survival. PATIENTS AND METHODS: In the present study, a total of 29 ccRCC fresh-frozen tissue samples were used; 14 samples from grade 1 (G1) RCC patients without metastatic disease and 15 from grade 3 (G3) RCC patients with synchronous metastatic disease. Expression profiling was performed with the Human Genome U133 Plus 2.0 Array (Affymetrix Corp., Santa Clara, CA, USA). Clinical data and long-term follow-up were obtained for all patients. The primary probe level analysis was performed using the Affymetrix MAS 5 algorithm. Further statistical processing was carried out by GSEA, using the Molecular Signatures Database, MSigDB (http://www.broad.mit.edu/gsea/msigdb/index.jsp). After selecting gene sets with the highest leading edge subsets, a cluster and a further analyses based on MSigDB data bank analysis was performed. RESULTS: In total, 15 poorly G3 ccRCC, 14 well differentiated G1 ccRCC and 14 normal renal tissue samples were analyzed for comparative gene expression profiling. There were 12 of 15 G3 ccRCC patients who had synchronous metastatic disease at the time of surgery (pN+ and/or distant metastases: pN+ only = 4, M+ only = 11 and pN+M+ = 3). The GSEA identified 700 gene sets. Out of these, 120 sets with the highest leading edge subset were selected monitored by hierarchical clustering G1 vs G3. Comparative analysis using the the MSigDB data bank for pathway network identified 16 gene sets that were differentially strongly over- or underexpressed in G3 vs G1 tumours and are involved in various aspects of tumour physiology, such as metastases and cell motility, signalling and cell proliferation, as well as gene products that are involved in the building of the extracellular matrix and as cell surface markers. CONCLUSIONS: We analyzed microarray data of gene expression in ccRCC comparing poorly differentiated and well differentiated tumour tissue samples. Using GSEA, we found a number of genes set candidates relevant to biological network processes with high complexity; conspicuously, these comprised members of the interleukin- and chemokine-family, cyclin-dependent kinases, angiogenic growth factors and transcriptional factors. This suggests that, in poorly differentiated aggressive ccRCC, there may be a limited number of gene sets that are responsible for the very aggressive biological behaviour. This comparison performed at a gene set level enables the identification of such congruency between different gene sets and whole data sets with respect to a specific biological question. GSEA embedded in the statistical workflow procedure for the suitable preparation of expression data may improve the analysis and avoid missing changes at the molecular level. A systematic approach such as GSEA is clearly needed to analyze raw data from microarray analyses, although these data can only be descriptive and the mass of raw data is derived from a relatively small number of tissue samples. However, consistent alterations of gene expression found in specific tumour entities may allow a better understanding of certain aspects of specific tumour biology. Therefore, the molecular characterization of individual tumours may potentially be useful for the better individual assessment of prognosis and, finally, the identification of biomarkers and targets of specific treatments may eventually help to improve treatment.

Molecular signatures and new candidates to target the pathogenesis of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic, inflammatory joint disease of unknown etiology and pronounced interpatient heterogeneity. To characterize RA at the molecular level and to uncover pathomechanisms, we performed genome-wide gene expression analysis. We identified a set of 1,054 genes significantly deregulated in pair-wise comparisons between RA and osteoarthritis (OA) patients, RA and normal donors (ND), or OA and ND. Correlation analysis revealed gene sets regulated identically in all three groups. As a prominent example secreted phosphoprotein 1 (SPP1) was identified to be significantly upregulated in RA compared with both OA and ND. SPP1 expression was found to correlate with genes expressed during an inflammatory response, T-cell activation and apoptosis, suggesting common underlying regulatory networks. A subclassification of RA patients was achieved on the basis of proteoglycan 4 (PRG4) expression, distinguishing PRG4 high and low expressors and reflecting the heterogeneity of the disease. In addition, we found that low PRG4 expression was associated with a more aggressive disease stage, which is in accordance with PRG4 loss-of-function mutations causing camptodactyly-arthropathy-coxa vara-pericarditis syndrome. Altogether we provide evidence for molecular signatures of RA and RA subclasses, sets of new candidate genes as well as for candidate gene networks, which extend our understanding of disease mechanisms and may lead to an improved diagnosis.

Mass spectrometric and peptide chip epitope analysis on the RA33 autoantigen with sera from rheumatoid arthritis patients.

As the potential of epitope chips for routine application in diagnostics relies on the careful selection of peptides, reliable epitope mapping results are of utmost interest to the medical community. Mass spectrometric epitope mapping in combination with peptide chip analysis showed that autoantibodies from patients who suffered from rheumatoid arthritis (RA) were directed against distinct surface structures on the full-length human autoantigen RA33 as well as against partial sequences. Using the combined mass spectrometric epitope extraction and peptide chip analysis approach, four sequence motifs on RA33 emerged as immuno-positive, showing that epitopes were not randomly distributed on the entire RA33 amino acid sequence. A sequential epitope motif ((245)GYGGG(249)) was determined on the C-terminal part of RA33 which matched with the Western blot patient screening results using the full-length protein and, thus, was regarded as a disease-associated epitope. Other epitope motifs were found on N-terminal partial sequences ((59)RSRGFGF(65), (111)KKLFVG(116)) and again on the C-terminal part ((266)NQQPSNYG(273)) of RA33. As recognition of these latter three motifs was also recorded by peptide chip analysis using control samples which were negative in the Western blot screening, these latter motifs were regarded as "cryptic epitopes". Knowledge of disease-associated epitopes is crucial for improving the design of a customized epitope peptide chip for RA and mass spectrometric epitope mapping pivotally assisted with selecting the most informative peptide(s) to be used for future diagnostic purposes.

Time course transcriptomics of IFNB1b drug therapy in multiple sclerosis.

Despite its generalized use as drug therapy for multiple sclerosis (MS), the molecular mechanisms of action of interferon beta (IFNB) are still poorly understood. IFNB therapy is long-termed and clinical effects are not immediate, therefore reliable early biomarkers for IFNB activity should maintain a differential expression over time, but longitudinal studies at a transcriptional level have been rare. Microarrays were used to monitor 18 IFNB1b treated MS patients at four time points spanning a period of 1 year. Genes showing in the majority of patients the greatest and most consistent changes in their expression levels were studied. Interferon regulated genes were significantly overrepresented. Fifteen markers were differentially expressed during all three time points and followed a consistent time course pattern: EIF2AK2, IFI6, IFI44, IFI44L, IFIH1, IFIT1, IFIT2, IFIT3, ISG15, MX1, OASL, RSAD2, SN, XAF1 and the marker 238704_at. Except for the last one, these biomarkers were all formerly identified as being indicative for IFNB activity. Expression changes were both early detectable and long lasting and could thus be optimal biomarkers for IFNB activity in long-term studies. Other known biomarkers of IFNB activity were found to be differentially expressed just for certain periods after therapy onset: Interleukin-8 was a short lasting marker and changes in STAT1 were detected with delay.

Transcriptome analysis in mouse tumors induced by Ret-MEN2/FMTC mutations reveals subtype-specific role in survival and interference with immune surveillance.

Activating mutations in the Ret proto-oncogene are responsible for occurrence of multiple endocrine neoplasia (MEN) type 2A and 2B, and familial medullary thyroid carcinoma (FMTC). A striking genotype-phenotype correlation between the mutated RET codon and clinical manifestation implies that tumorigenesis is conditioned by the type of mutation. We investigated gene expression profiles between and within distinct MEN2 subtypes through whole-genome microarray analysis in tumors induced by NIH-3T3 cells transformed with defined RET-MEN2A (C609Y, C634R), MEN2B, (A883F, M918T), and FMTC (Y791F) mutations. Expression profiling identified a statistically significant modification of 1494 genes, 628 down- and 866 upregulated in MEN2B compared with MEN2A/FMTC tumors. By contrast, no obvious alterations were observed among individual MEN2B and MEN2A type mutations, or between MEN2A and FMTC. Functional clustering of differential genes revealed RET-MEN2B specific upregulation of genes associated with novel growth and survival pathways. Intriguingly, RET-MEN2A/FMTC-specific tumors were characterized by a considerable number of genes involved in the host antitumor immune response via stimulation of natural killer/T-cell proliferation, migration, and cytotoxicity, which were completely absent in RET-MEN2B related cancers. QPCR on tumors versus cultured NIH-RET cell lines demonstrated that they are largely attributed to the host innate immune system, whereas expression of CX3CL1 involved in leukocyte recruitment is exclusively RET-MEN2A/FMTC tumor cell dependent. In correlation, massive inflammatory infiltrates were apparent only in tumors carrying MEN type 2A/FMTC mutations, suggesting that RET-MEN2B receptors specifically counteract immune infiltration by preventing chemokine expression, which may contribute to the different clinical outcome of both subtypes.

Gene expression profiling of arthritis using a QTL chip reveals a complex gene regulation of the Cia5 region in mice.

One of the major quantitative trait loci (QTLs) associated with arthritis in crosses between B10.RIII and RIIIS/J mice is the Cia5 on chromosome 3. Early in the congenic mapping process it was clear that the locus was complex, consisting of several subloci with small effects. Therefore, we developed two novel strategies to dissect a QTL: the partial advanced inter-cross (PAI) strategy, with which we recently found the Cia5 region to consist of three loci, Cia5, Cia21 and Cia22, and now we introduce the QTL-chip strategy, where we have combined congenic mapping with a QTL-restricted expression profiling using a novel microarray design. The expression of QTL genes was compared between parental and congenic mice in lymph node, spleen and paw samples in five biological replicates and in dye-swapped experiments at three time points during the induction phase of arthritis. The QTL chip approach revealed 4 genes located in Cia21, differently expressed in lymph nodes, and 14 genes in Cia22, located within two clusters. One cluster contains six genes, differently expressed in spleen, and the second cluster contains eight genes, differently expressed in paws. We conclude the QTL-chip strategy to be valuable in the selection of candidate genes to be prioritized for further investigation.

Protective effect of endotoxin preconditioning in pancreas: analysis with DNA chip arrays.

Recently we demonstrated a protective effect of endotoxin preconditioning 24 hours before pancreatic ischemia-reperfusion injury, which has also been described for other organs. The mechanisms underlying this phenomenon, such as differential gene expression, are poorly investigated. We chose to approach this question by investigating differential gene expression in the rat pancreas over the time course of endotoxin pretreatment. Male Wistar rats (5 groups, 5 animals per group) were pretreated with endotoxin intraperitoneally (1 mg/kg of body weight). After treatment at 30 minutes, and at 3 and 24 hours the pancreas was removed. Untreated animals and animals with injection of saline solution served as controls. After RNA isolation, RNA was pooled and hybridized to Affymetrix chips to measure the relative mRNA levels of 7000 genes and 1000 expression sequence tags. Three hours after administration of endotoxin there was an activation of proinflammatory transcription factors and other proinflammatory genes. After 24 hours there was a clear decrease of these proinflammatory genes, but a remaining and increasing upregulation of important antiapoptotic genes, antiproteases, and other probably protective genes. There was also a significant upregulation of complement factors. It was surprising that heat-shock proteins and other typical immediate early genes of the AP-1 complex were not upregulated. Our data show that 24 hours after endotoxin stress there is a regulation of a network of genes that represents a multifaceted preconditioning. As most important factors, inhibition of apoptosis and antiproteatic strategies are identified. Heat-shock proteins seem to play no important role in the mechanism of endotoxin preconditioning.

DNA microarray technology and its applications in dermatology.

The use of DNA microarray technology in biomedical research has dramatically increased during the past years. In the present report, we provide an overview on the basic DNA microarray technology and biostatistical methods for gene expression analysis. A focus is then put on its applications in dermatological research. In recent years, a series of gene expression studies have been performed for various dermatological diseases, such as malignant melanoma, psoriasis and lupus erythematosus. These analyses have identified interesting target genes as well as putative disease susceptibility loci. However, further functional studies will be needed for a more complete understanding of the pathogenesis of these diseases. This may be performed by means of the recently developed RNA interference technology. Besides its role in large-scale gene expression studies, DNA microarray technology has proved to be a valuable tool for genomic screens of genetic alterations, e.g. single nucleotide polymorphisms. These play a role in tumour development and progression, and also function as genetic markers for disease susceptibility. Taken together, DNA microarray technology opens enormous perspectives for dermatologists. It may help us understand the complex pathogenesis of a wide variety of dermatologic diseases and identify their genetic background.

[The extracellular matrix structure in keratoconus]. / Die Struktur der Extrazellulärmatrix bei Keratokonus.

PURPOSE: Keratoconus is a non-inflammatory disease characterized by progressive thinning of the central cornea. There are indications for an autosomal dominant heredity. Our purpose was to find correlations between gene expression and structural changes in extracellular matrix components. METHODS: Stromal RNA (keratoconus and comparison) was isolated from corneas, transformed to cRNA and analysed using biochips (Affymetrix). Structural investigations were performed by laser scanning and transmission electron microscopy. RESULTS: In keratoconus corneas there was an upregulation of different extracellular matrix components (collagen XV, metalloproteases) and a down-regulation of collagen IV (alpha1, alpha3) and versican. The morphological changes correlated to genetic obtained data. The orthogonal arrangement of the collagen fibrills (anteriorly and central) was altered in the collagen matrix of keratoconus corneas. CONCLUSIONS: The changes point on deregulation of the matrix arrangement. The interest in the cause of the disease is focused on the interfibrillar arrangement, the interaction between collagen and proteoglucanes.

[Gene expression in keratoconus. Initial results using DNA microarrays]. / Genexpression bei Keratokonus. Erster Einsatz von DNA-Microarrays.

INTRODUCTION: Keratoconus is a non-inflammatory ocular disease characterised by conical deformation, progressive thinning and scarring of the central cornea. Despite intensive investigations, the exact cause of the disease still remains unclear. Clinical studies provide strong indications of a major genetic role in the aetiology. We set out to examine the involvement in the manifestation of keratoconus of any of the 5,600 gene specificities available on the Affymetrix GeneChip HuGeneFL. METHODS: After examination of two corneas they were stored in RNAlater, RNA was extracted and hybridised on the chips. Using a combination of dyes it was possible to read the chips with laser detection and to visualise the gene expression pattern. RESULTS: We found an upregulation of collagens, versican, metalloproteinases and cell adhesion proteins. A downregulation was observed for TIGR protein, cytokeratins, eyes absent homologue (Eab1) and the proteins for radical treatment selenoprotein P and monooxygenase. CONCLUSIONS: Our results indicate that keratoconus is a process in which repair and scar-formation mechanisms operate at the same time. As candidate genes for this mechanism, collagen IV and related proteoglycans were favoured.

Differential expression of thrombospondin 2 in primary and metastatic malignant melanoma.

In the present report we used oligonucleotide microarray analysis for the identification of genes characterizing the late-stage metastatic phenotype of malignant melanoma. A panel of 5,600 genes was analysed in a low aggressive and a highly aggressive (metastatic) human malignant melanoma cell line, respectively. More than 300 differentially regulated genes were identified. High metastatic potential correlated with upregulated mRNA expression in the groups of cytoskeletal proteins, apoptosis and cell cycle proteins, GTP binding proteins and oncogenes, extracellular ligands and receptors, transcription and translation factors. In contrast, most angiogenesis factors, extracellular matrix molecules, and melanoma-specific antigens were downregulated. Particular target genes were further analysed by in situ hybridization and immunohistochemical staining of primary malignant melanomas and melanoma metastases. Here, we show that thrombospondin 2, an extracellular matrix molecule which was differentially regulated in the microarray analysis, was strongly expressed in melanoma metastases, but not in primary tumours. The identification of thrombospondin 2 as a target molecule emphasizes the importance of cell-matrix interactions for the pathogenesis of malignant melanoma metastasis and may open future perspectives for treatment of this tumour.

Microarray analysis of differentially expressed genes in placental tissue of pre-eclampsia: up-regulation of obesity-related genes.

Susceptibility genes present in both mother and fetus most likely contribute to the risk of pre-eclampsia. Placental biopsies were therefore investigated by high-density DNA microarray analysis to determine genes differentially regulated within chorionic villous tissue in pre-eclampsia and normal pregnancy. The pooled RNAs of pre-eclamptic and normotensive subjects were hybridized to the HuGeneFL array representing sequences from approximately 5600 full-length human cDNAs. The differentially expressed genes that were detected could be categorized into nine groups: adhesion molecules, obesity-related genes, transcription factors/signalling molecules, immunological factors, neuromediators, oncogenic factors, protease inhibitors, hormones and growth factor-binding proteins. Among those, the obesity-related genes included putative candidate genes associated with the pathogenesis of pre-eclampsia. One of the most up-regulated transcripts was the obese gene (43.6-fold change), and this was reflected by elevated leptin protein levels. In the case of feto-maternal contribution of polymorphic genes to pre-eclampsia, expression analysis of placental tissue has lead to numerous target genes waiting for large scale genetic linkage analyses.

The KOX zinc finger genes: genome wide mapping of 368 ZNF PAC clones with zinc finger gene clusters predominantly in 23 chromosomal loci are confirmed by human sequences annotated in EnsEMBL.

The chromosome locations of 368 human Kruppel-type zinc finger (ZNF) PAC clones were physically mapped by FISH to human chromosomes in support of recent efforts of assigning KOX cDNAs (KOX1-KOX32) to zinc finger gene clusters. Recent mapping results were validated and confirmed by sequence comparisons to zinc finger gene sequences automatically annotated in EnsEMBL. In toto, 799 Kruppel-type zinc finger genes have been annotated in EnsEMBL of which 290 genes are found to encode KRAB domains. Sequence homologies of the zinc finger domains were used to establish phylogenic trees of KOX zinc finger genes as well as of all KRAB containing human zinc finger and KOX genes documenting the evolution of KRAB zinc finger genes late in primate evolution. A list of 368 assigned ZNF PAC clones is available under http://www.pzr.uni-rostock.de/supplements.

Gene expression profiling of the nervous system in murine experimental autoimmune encephalomyelitis.

Multiple sclerosis is thought to be a polygenic disease driven by dysregulation of the immune system leading to an autoimmune response against one or several antigens of cerebral white matter tissue. Experimental autoimmune encephalomyelitis (EAE) is a mouse model that is used to study the aetiology and pathogenesis of multiple sclerosis and new therapeutic approaches. We used oligonucleotide microarrays to determine gene expression profiles of the inflamed spinal cords of EAE mice at the onset and at the peak of the disease. Of the approximately 11 000 genes studied, 213 were regulated differentially and 100 showed consistent differential regulation throughout the disease. Inflammation resulted in a profile of increased gene expression of immune-related molecules, extracellular matrix and cell adhesion molecules and molecules involved in cell division and transcription, and differential regulation of molecules involved in signal transduction, protein synthesis and metabolism. Of the 104 genes with defined chromosomal locations, 51 mapped to known EAE-linked quantitative trait loci and as such are putative candidate genes for susceptibility to EAE.