Citations

Abstract Primary central nervous system (CNS) lymphoma (PCNSL) is a diffuse large B-cell lymphoma (DLBCL) confined to the CNS. A genome-wide gene expression comparison between PCNSL and non-CNS DLBCL was performed, the latter consisting of both nodal and extranodal DLBCL (nDLBCL and enDLBCL), to identify a “CNS signature.” Pathway analysis with the program SigPathway revealed that PCNSL is characterized notably by significant differential expression of multiple extracellular matrix (ECM) and adhesion-related pathways. The most significantly up-regulated gene is the ECM-related osteopontin (SPP1). Expression at the protein level of ECM-related SPP1 and CHI3L1 in PCNSL cells was demonstrated by immunohistochemistry. The alterations in gene expression can be interpreted within several biologic contexts with implications for PCNSL, including CNS tropism (ECM and adhesion-related pathways, SPP1, DDR1), B-cell migration (CXCL13, SPP1), activated B-cell subtype (MUM1), lymphoproliferation (SPP1, TCL1A, CHI3L1), aggressive clinical behavior (SPP1, CHI3L1, MUM1), and aggressive metastatic cancer phenotype (SPP1, CHI3L1). The gene expression signature discovered in our study may represent a true “CNS signature” because we contrasted PCNSL with wide-spectrum non-CNS DLBCL on a genomic scale and performed an in-depth bioinformatic analysis.

Acetylation of histone H3 lysine 56 (H3-K56) occurs in S phase, and cells lacking H3-K56 acetylation are sensitive to DNA-damaging agents. However, the histone acetyltransferase (HAT) that catalyzes global H3-K56 acetylation has not been found. Here we show that regulation of Ty1 transposition gene product 109 (Rtt109) is an H3-K56 HAT. Cells lacking Rtt109 or expressing rtt109 mutants with alterations at a conserved aspartate residue lose H3-K56 acetylation and exhibit increased sensitivity toward genotoxic agents, as well as elevated levels of spontaneous chromosome breaks. Thus, Rtt109, which shares no sequence homology with any other known HATs, is a unique HAT that acetylates H3-K56.

Abstract Background: The genetic heterogeneity of sporadic colorectal cancer (CRC) makes the choice of genetic markers and sequence variation–detection technologies critical to the performance of screening assays. We have previously described the effectiveness of a CRC assay composed of 22 known variants in KRAS, APC, TP53, and BAT-26 (V1). We introduce a new marker formulation (V2) that includes detection of de novo variation in APC, PIK3CA, and CTNNB1, hypermethylated sequences within SMARCA3 and VIM, and a single-base variation within BRAF. We compared the abilities of the V1 and V2 markers to detect aberrant DNA in colorectal neoplasias. Methods: V1 and V2 marker formulations were used to analyze 144 colorectal tissue samples comprising 50 precancerous adenomas, 94 carcinomas, and 11 nonpathologic tissues. V1 analysis consisted of single-base extension analysis of the 22 V1 variants. V2 analysis consisted of DNA scanning of the APC mutation cluster region, PIK3CA exons 9 and 20, CTNNB1 exon 3, analysis for the BRAF Val600Glu substitution, and methylation-specific PCR analysis of VIM and SMARCA3. Results: The V2 marker formulation had significantly higher sensitivity than the V1 markers for carcinomas (93.6% and 72.3%, respectively; P = 0.0002) and adenomas (92.0% and 62.0%, respectively; P = 0.0006). None of the nonpathologic samples were positive for any marker. Conclusions: We demonstrate improved sensitivity of a new marker formulation (V2) to detect aberrant DNA in CRC and precancerous adenoma tumor tissues.

AbstractOBJECTIVE:Olfaction is often sacrificed to gain access to the cranial base in anterior craniofacial surgery. We describe the long-term results of olfactory function in patients who underwent anterior craniofacial surgery and a cribriform plate osteotomy to preserve olfaction.METHODS:Between 1992 and 2004, 28 patients underwent 29 cribriform plate osteotomies in an attempt to preserve olfaction during anterior craniofacial surgery performed through modified extended transbasal approaches. Patients' charts and office notes were reviewed retrospectively. Formal olfactory testing was available in 5 patients, but most data were based on patients' subjective reports of olfaction. Olfactory preservation was defined by the subjective ability to detect fumes such as coffee, chocolate, roses, and orange juice regardless of the intensity of the sensation. Follow-up was based on phone calls to patients.RESULTS:Four patients were lost to follow-up and excluded. Therefore, follow-up was available in 24 patients after 25 procedures. On the basis of patients' subjective reports, olfaction was spared in 22 patients after 23 procedures (92%) and was confirmed objectively in the five patients formally tested. After surgery, only two patients were anosmic.CONCLUSION:Olfaction can be preserved in selected patients undergoing anterior craniofacial surgery. At least 1 cm of nasal mucosa should remain attached to the cribriform plate, which can be achieved by including the nasal bone in the osteotomy of the orbital bar. A medial orbital canthopexy is therefore necessary after these procedures.

Recent evidence suggests that alveolar epithelial cells (AECs) may contribute to the development, propagation, and resolution of acute lung injury (ALI)/acute respiratory distress syndrome (ARDS). Proinflammatory cytokines, pathogen products, and injurious mechanical ventilation are important contributors of excessive inflammatory responses in the lung. In the present study, we used cDNA microarrays to define the gene expression patterns of A549 cells (an AEC line) in the early stages of three models of pulmonary parenchymal cell activation: cells treated with tumor necrosis factor-α (TNFα) (20 ng/ml), lipopolysaccharide (LPS, 1 μg/ml), or cyclic stretch (20% elongation) for either 1 h or 4 h. Differential gene expression profiles were determined by gene array analysis. TNFα induced an inflammatory response pattern, including induction of genes for chemokines, inflammatory mediators, and cell surface membrane proteins. TNFα also increased genes related to pro- and anti-apoptotic proteins, signal transduction proteins, and transcriptional factors. TNFα further induced a group of genes that may form a negative feedback loop to silence the NFκB pathway. Stimulation of AECs with mechanical stretch changed cell morphology and activated Src protein tyrosine kinase. The combination of TNFα plus stretch enhanced or attenuated expression of multiple genes. LPS decreased microfilament polymerization but had less impact on NFκB translocation and gene expression. Results from this study indicate that AECs can tailor their response to different stimuli or/and combination of stimuli and subsequently play an important role in acute inflammatory responses in the lung.

Mechanical unloading of the heart with a left ventricular assist device (LVAD) significantly decreases mortality in patients with heart failure. Moreover, it provides a human model to define the critical regulatory genes governing myocardial remodeling in response to significant reductions in wall stress. Statistical analysis of a gene expression library of 19 paired human heart samples harvested at the time of LVAD implant and again at explant revealed a set of 22 genes that were downregulated and 85 genes that were upregulated in response to mechanical unloading with a false discovery rate of less than 1%. The analysis revealed a high percentage of genes involved in the regulation of vascular networks including neuropilin-1 (a VEGF receptor), FGF9, Sprouty1, stromal-derived factor 1, and endomucin. Taken together these findings suggest that mechanical unloading alters the regulation of vascular organization and migration in the heart. In addition to vascular signaling networks, GATA-4 binding protein, a critical mediator of myocyte hypertrophy, was significantly downregulated following mechanical unloading. In summary, these findings may have important implications for defining the role of mechanical stretch and load on autocrine/paracrine signals directing vascular organization in the failing human heart and the role of GATA-4 in orchestrating reverse myocardial remodeling. This unbiased gene discovery approach in paired human heart samples has the potential to provide critical clues to the next generation of therapeutic treatments aimed at heart failure.

AbstractDT388–GM-CSF, a targeted fusion toxin constructed by conjugation of human granulocyte-macrophage colony-stimulating factor (GM-CSF) with the catalytic and translocation domains of diphtheria toxin, is presently in phase I trials for patients with resistant acute myeloid leukemia. HL-60/VCR, a multidrug-resistant human myeloid leukemia cell line, and wild-type HL-60 cells were used to study the impact of DT388–GM-CSF on metabolism of ceramide, a modulator of apoptosis. After 48 hours with DT388–GM-CSF (10 nM), ceramide levels in HL-60/VCR cells rose 6-fold and viability fell to 10%, whereas GM-CSF alone was without influence. Similar results were obtained in HL-60 cells. Examination of the time course revealed that protein synthesis decreased by about 50% and cellular ceramide levels increased by about 80% between 4 and 6 hours after addition of DT388–GM-CSF. By 6 hours this was accompanied by activation of caspase-9, followed by activation of caspase-3, cleavage of caspase substrates, and chromatin fragmentation. Hygromycin B and emetine failed to elevate ceramide levels or induce apoptosis at concentrations that inhibited protein synthesis by 50%. Exposure to C6-ceramide inhibited protein synthesis (EC50∼5 μM) and decreased viability (EC50 ∼6 μM). Sphingomyelinase treatment depleted sphingomyelin by about 10%, while increasing ceramide levels and inhibiting protein synthesis. Diphtheria toxin increased ceramide and decreased sphingomyelin in U-937 cells, a cell line extremely sensitive to diphtheria toxin; exposure to DT388–GM-CSF showed sensitivity at less than 1.0 pM. Diphtheria toxin and conjugate trigger ceramide formation that contributes to apoptosis in human leukemia cells through caspase activation and inhibition of protein synthesis.