Integrin-linked kinase is required for laminin-2–induced oligodendrocyte cell spreading and CNS myelination

Early steps in myelination in the central nervous system (CNS) include a specialized and extreme form of cell spreading in which oligodendrocytes extend large lamellae that spiral around axons to form myelin. Recent studies have demonstrated that laminin-2 (LN-2; α2β1γ1) stimulates oligodendrocytes to extend elaborate membrane sheets in vitro (cell spreading), mediated by integrin α6β1. Although a congenital LN-2 deficiency in humans is associated with CNS white matter changes, LN-2–deficient (dy/dy) mice have shown abnormalities primarily within the peripheral nervous system. Here, we demonstrate a critical role for LN-2 in CNS myelination by showing that dy/dy mice have quantitative and morphologic defects in CNS myelin. We have defined the molecular pathway through which LN-2 signals oligodendrocyte cell spreading by demonstrating requirements for phosphoinositide 3-kinase activity and integrin-linked kinase (ILK). Interaction of oligodendrocytes with LN-2 stimulates ILK activity. A dominant negative ILK inhibits LN-2–induced myelinlike membrane formation. A critical component of the myelination signaling cascade includes LN-2 and integrin signals through ILK

Factors associated with month 2 smear non-conversion among category 1 tuberculosis patients in Karachi, Pakistan

Predictors of smear non-conversion at baseline can help identify cases at risk for failure of tuberculosis treatment. Retrospective data for smear-positive Category 1 patients in Karachi, Pakistan, was analyzed. Predictors of sputum conversion were determined using multiple logistic regression with sputum conversion as outcome variable and patient demographics, baseline weight, baseline sputum smear grade, case-finding approach as explanatory variables. Age ≥35 years, baseline sputum grade of 3+ were significantly associated with predicting sputum smear positivity at month 2 of treatment. Monitoring compliance to TB treatment should be considered amongst older patients and those with a high sputum grade at baseline

Dynamic Near-Infrared Optical Imaging of 2-Deoxyglucose Uptake by Intracranial Glioma of Athymic Mice

BACKGROUND: It is recognized that cancer cells exhibit highly elevated glucose metabolism compared to non-tumor cells. We have applied in vivo optical imaging to study dynamic uptake of a near-infrared dye-labeled glucose analogue, 2-deoxyglucose (2-DG) by orthotopic glioma in a mouse model. METHODOLOGY AND PRINCIPAL FINDINGS: The orthotopic glioma model was established by surgically implanting U87-luc glioma cells into the right caudal nuclear area of nude mice. Intracranial tumor growth was monitored longitudinally by bioluminescence imaging and MRI. When tumor size reached >4 mm diameter, dynamic fluorescence imaging was performed after an injection of the NIR labeled 2-DG, IRDye800CW 2-DG. Real-time whole body images acquired immediately after i.v. infusion clearly visualized the near-infrared dye circulating into various internal organs sequentially. Dynamic fluorescence imaging revealed significantly higher signal intensity in the tumor side of the brain than the contralateral normal brain 24 h after injection (tumor/normal ratio, TNR = 2.8+/-0.7). Even stronger contrast was achieved by removing the scalp (TNR = 3.7+/-1.1) and skull (TNR = 4.2+/-1.1) of the mice. In contrast, a control dye, IRDye800CW carboxylate, showed little difference (1.1+/-0.2). Ex vivo fluorescence imaging performed on ultrathin cryosections (20 microm) of tumor bearing whole brain revealed distinct tumor margins. Microscopic imaging identified cytoplasmic locations of the 2-DG dye in tumor cells. CONCLUSION AND SIGNIFICANCE: Our results suggest that the near-infrared dye labeled 2-DG may serve as a useful fluorescence imaging probe to noninvasively assess intracranial tumor burden in preclinical animal models

Improved protein arrays for quantitative systems analysis of the dynamics of signaling pathway interactions

An improved version of quantitative protein array platform utilizing linear Quantum dot signaling for systematically measuring protein levels and phosphorylation states is presented. The signals are amplified linearly by a confocal laser Quantum dot scanner resulting in ~1000-fold more sensitivity than traditional Western blots, but are not linear by the enzyme-based amplification. Software is developed to facilitate the quantitative readouts of signaling network activities. Kinetics of EGFRvIII mutant signaling was analyzed to quantify cross-talks between EGFR and other signaling pathways

A TNF-JNK-Axl-ERK signaling axis mediates primary resistance to EGFR inhibition in glioblastoma.

Aberrant epidermal growth factor receptor (EGFR) signaling is widespread in cancer, making the EGFR an important target for therapy. EGFR gene amplification and mutation are common in glioblastoma (GBM), but EGFR inhibition has not been effective in treating this tumor. Here we propose that primary resistance to EGFR inhibition in glioma cells results from a rapid compensatory response to EGFR inhibition that mediates cell survival. We show that in glioma cells expressing either EGFR wild type or the mutant EGFRvIII, EGFR inhibition triggers a rapid adaptive response driven by increased tumor necrosis factor (TNF) secretion, which leads to activation in turn of c-Jun N-terminal kinase (JNK), the Axl receptor tyrosine kinase and extracellular signal-regulated kinases (ERK). Inhibition of this adaptive axis at multiple nodes rendered glioma cells with primary resistance sensitive to EGFR inhibition. Our findings provide a possible explanation for the failures of anti-EGFR therapy in GBM and suggest a new approach to the treatment of EGFR-expressing GBM using a combination of EGFR and TNF inhibition

SNS-032 Prevents Tumor Cell-Induced Angiogenesis By Inhibiting Vascular Endothelial Growth Factor1

Cell proliferation, migration, and capillary network formation of endothelial cells are the fundamental steps for angiogenesis, which involves the formation of new blood vessels. The purpose of this study is to investigate the effect of a novel aminothiazole SNS-032 on these critical steps for in vitro angiogenesis using a coculture system consisting of human umbilical vein endothelial cells (HUVECs) and human glioblastoma cells (U87MG). SNS-032 is a potent selective inhibitor of cyclin-dependent kinases 2, 7, and 9, and inhibits both transcription and cell cycle. In this study, we examined the proliferation and viability of HUVECs and U87MG cells in the presence of SNS-032 and observed a dose-dependent inhibition of cellular proliferation in both cell lines. SNS-032 inhibited threedimensional capillary network formations of endothelial cells. In a coculture study, SNS-032 completely prevented U87MG cell-mediated capillary formation of HUVECs. This inhibitor also prevented the migration of HUVECs when cultured alone or cocultured with U87MG cells. In addition, SNS-032 significantly prevented the production of vascular endothelial growth factor (VEGF) in both cell lines, whereas SNS-032 was less effective in preventing capillary network formation and migration of endothelial cells when an active recombinant VEGF was added to the medium. In conclusion, SNS-032 prevents in vitro angiogenesis, and this action is attributable to blocking of VEGF

Epidermal Growth Factor Receptor in Glioma: Signal Transduction, Neuropathology, Imaging, and Radioresistance

Aberrant epidermal growth factor receptor (EGFR) signaling is common in cancer. Increased expression of wild type and mutant EGFR is a widespread feature of diverse types of cancer. EGFR signaling in cancer has been the focus of intense investigation for decades primarily for two reasons. First, aberrant EGFR signaling is likely to play an important role in the pathogenesis of cancer, and therefore, the mechanisms of EGFR-mediated oncogenic signaling are of interest. Second, the EGFR signaling system is an attractive target for therapeutic intervention. EGFR gene amplification and overexpression are a particularly striking feature of glioblastoma (GBM), observed in approximately 40% of tumors. GBM is the most common primary malignant tumor of the central nervous system in adults. In approximately 50% of tumors with EGFR amplification, a specific EGFR mutant (EGFRvIII, also known as EGFR type III, de2-7, ΔEGFR) can be detected. This mutant is highly oncogenic and is generated from a deletion of exons 2 to 7 of the EGFR gene, which results in an in-frame deletion of 267 amino acids from the extracellular domain of the receptor. EGFRvIII is unable to bind ligand, and it signals constitutively. Although EGFRvIII has the same signaling domain as the wild type receptor, it seems to generate a distinct set of downstream signals that may contribute to an increased tumorigenicity. In this review, we discuss recent progress in key aspects of EGFR signaling in GBM, focusing on neuropathology, signal transduction, imaging of the EGFR, and the role of the EGFR in mediating resistance to radiation therapy in GBM