Glial Cell Invasion Research Articles

HIF-1α facilitates glioma proliferation and invasion by activating pyroptosis signaling axis

BackgroundHIF-1α is thought to be a novel regulator which contributes to carcinogenesis. However, the mechanism underlying the effect of HIF-1α in gliomas remains largely unknown.MethodsIn the research, we demonstrate that HIF-lα mRNA and protein levels are elevated in glioma cells. The colony formation assays, transwell assays, and wound-healing assays showed that overexpression of HIF-1α promoted proliferation and invasion of glioma cells.ResultsOverexpression of HIF-lα also increased the expression of inflammatory factors related to pyrolysis (TNF-α, IL-10, and IL-1β) and protein related to pyrolysis signal pathway (NLRP3, ASC, caspase-1, GSDMD, and GSDME).ConclusionsTherefore, we speculate that HIF-1α promotes the proliferation and invasion of glial cells by regulating pyrolysis pathway. These results might provide a novel strategy and target for treatment of glioma.

Deletion of NADH oxidase in Listeria monocytogenes promotes the bacterial infection of brain

NADH oxidase (NOX) plays important roles in respiration and reactive oxygen species (ROS) generation in cells. In this study, we explored the function of NOX in Listeria monocytogenes by gene deletion. From our results, nox mutant strain (∆nox) had lower H2O2 level and showed no significant alteration in bacteria growth activity. But it had enhanced invasiveness during the invasion of glial cells and mice brain compared to wildtype strain. Furthermore, several virulence genes involved in invasion, such as inlA, inlB, vip and sigB, were upregulated in ∆nox, and the alterations could be restored by complementation. To explore if nox was involved in the interaction of pathogen and host, we examined the generation of host ROS including superoxide and H2O2 during infection, and found ∆nox invasion leading to less superoxide and H2O2 generation. Besides, the upregulation of pro-inflammatory factors in glial cells was restrained when invaded by ∆nox compared to wildtype and complementary strain. In conclusion, our study evaluated the function of nox in L. monocytogenes and indicated that nox could regulate the invasion of L. monocytogenes by regulating virulence genes expression and the interaction of host-and- pathogens.

Abstract 4102: FAT1: A novel regulator of cancer and inflammation.

Abstract FAT1, a member of cadherin superfamily is a large trans-membrane protein with 34 cadherin repeats. FAT1 was originally identified as a tumor suppressor gene in Drosophila but its role in human cancers is just beginning to unfold. Considering its tumor suppressive role in Drosophila, few studies have reported similar function of FAT1 in human cancers but on the contrary several studies have also demonstrated oncogenic role of FAT1. In this study we identified oncogenic role of FAT1 in human glioma where knockdown of FAT1 was found to inhibit migration and invasion of glial cells. Further investigation into molecular mechanism of FAT1 function revealed direct link to AP-1 mediated transcription via Programmed Cell Death 4 (PDCD4). Knockdown of FAT1 was found to inhibit AP-1 mediated transcription and expression of downstream target genes. AP-1 is a known mediator of cancer and inflammation and accordingly we found decreased expression of several pro-inflammatory markers like COX-2, cytokines like IL-6, IL-1β and chemokines like IL-8, MCP-1 and MIP-2 after FAT1 knockdown. Interestingly, the expression of anti-inflammatory cytokine IL-10 was found to be increased after FAT1 knockdown. Moreover FAT1 expression was also found to positively correalate with COX-2 and IL-6 expression in primary GBM tumors. Thus, in this study FAT1 was found to act as an oncogene as well as to orchestrate the inflammatory milieu of the glioma cells by regulating AP-1 mediated transcription via PDCD4. Citation Format: Bhawana Dikshit, Parthaprasad Chattopadhyay, Subrata Sinha, Kunzang Chosdol. FAT1: A novel regulator of cancer and inflammation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4102. doi:10.1158/1538-7445.AM2013-4102

Regulation of glial cell invasion growth by AS - miR - 21 via RECK in vitro and vivo

Objective To investigate the regulation of miR - 21 on invasion growth of human glioma cells by RECK.Method The human glioma LN229 cells were transfected with AS - miR - 21 or scrambled sequences by Lipofectamine2000.Real time PCR was conducted to detect the expression of miR-21.Luciferase experiment was performed to detect the relationship between miR-21 and RECK.The expression of RECK was evaluated by Western blot.The invasion ability was evaluated by transwell assay and subcutaneous models.Western Blot, ELISA and immunofluorescence were used to estimate the changes of MMP2/9.Results The expression of miR - 21 in LN229 cells decreased after transfection with AS-miR-21. It was proved that RECK was a direct target of miR -21 by luciferase experiment.Meanwhile, the high expression of RECK protein in AS - miR -21 group conformed its important function in this mechanism.Transwell assay demonstrated decreased invasion capability of LN229 cell lines transfected with AS- miR- 21.Western blot, ELISA, and immunofluorescence demonstrated the levels of MMP2/9 were down -regulated in AS -miR -21 group compared with control and scrambled group.Conclusions AS - miR -21 could depress the invasion of glioma cells owing to up - regulating the level of RECK which could inhibit MMP2/9 activities both in vitro and vivo. Key words: Glioma; Invasion; miR -21; RECK

Modeling Glioma Growth and Invasion in Drosophila melanogaster

Glioblastoma is the most common and most malignant intrinsic human brain tumor, characterized by extensive invasion and proliferation of glial (astrocytic) tumor cells, frequent activation of tyrosine kinase receptor signaling pathways, relative resistance to chemotherapy and radiotherapy, and poor prognosis. Using the Gal4-UAS system, we have produced glioma models in Drosophila by overexpressing homologs of human tyrosine kinase receptors under control of the glia-specific promoter reversed polarity (repo). Glial overexpression of activated epidermal growth factor receptor (EGFR) resulted in enhanced proliferation and migration of larval glial cells with increased numbers in the eye imaginal disc, diffuse tumor-like enlargement of the optic stalk, and marked ectopic invasion of glial cells along the optic nerve. Glial overexpression of the downstream kinase PI3K showed similar pathology. Overexpression of activated pvr (platelet-derived growth factor receptor/vascular endothelial growth factor receptor homolog) led to migration of glial cells along the optic nerve, whereas expression of activated htl (fibroblast growth factor receptor 1 homolog) and INR (insulin receptor) showed markedly elevated numbers of glial cells in the optic stalk. The EGFR/phosphatidylinositol 3-phosphate kinase (PI3K) phenotype was partly reverted by the administration of the EGFR tyrosine kinase inhibitor gefitinib and completely rescued by the PI3K inhibitor wortmannin and the Akt inhibitor triciribine. We suggest that Drosophila models will be useful for deciphering signaling cascades underlying abnormal behavior of glioma cells for genetic screens to reveal interacting genes involved in gliomagenesis and for experimental therapy approaches.

A microarray study to characterize the molecular mechanism of TIMP-3-mediated tumor rejection

Glial cell invasion is a multistep cellular process that involves a complex system of tightly regulated proteases (matrix metalloproteinases; MMPs) and their endogenous inhibitors (tissue inhibitors of metalloproteinases; TIMPs) to mediate the degradation of the basement membrane and extracellular matrix. Tissue inhibitor of metalloproteinases-3 (TIMP-3) is a matrix-bound inhibitor of MMPs. In the present study, we have overexpressed the TIMP3 gene in human glioma cells with a herpes simplex virus type 1 amplicon-based vector. Oligonucleotide DNA arrays were employed to identify genes that were differentially modulated by the overexpression of TIMP-3. Consistent with the function of TIMP-3, genes associated with angiogenesis, growth factors, cytokines, death receptors, and substrates of the various MMPs were found to be up-regulated. Furthermore, caspases are important in the signaling pathway of cellular apoptosis, and the overexpression of TIMP-3 in glioma cells is tightly associated with the activation of caspases, including caspase-1, at both the mRNA level (P=0.0371) and the protein level. Moreover, the activation of an apoptotic pathway via the overexpression of TIMP-3 induced apoptosis of transduced human glioma cells in vitro and the growth inhibition of human glioma tumor xenografts in immunodeficient mice.

Inhibition of Autotaxin by Lysophosphatidic Acid and Sphingosine 1-Phosphate

Autotaxin (ATX) or nucleotide pyrophosphatase/phosphodiesterase 2 (NPP2) is an NPP family member that promotes tumor cell motility, experimental metastasis, and angiogenesis. ATX primarily functions as a lysophospholipase D, generating the lipid mediator lysophosphatidic acid (LPA) from lysophosphatidylcholine. ATX uses a single catalytic site for the hydrolysis of both lipid and non-lipid phosphodiesters, but its regulation is not well understood. Using a new fluorescence resonance energy transfer-based phosphodiesterase sensor that reports ATX activity with high sensitivity, we show here that ATX is potently and specifically inhibited by LPA and sphingosine 1-phosphate (S1P) in a mixed-type manner (Ki approximately 10(-7) M). The homologous ecto-phosphodiesterase NPP1, which lacks lysophospholipase D activity, is insensitive to LPA and S1P. Our results suggest that, by repressing ATX activity, LPA can regulate its own biosynthesis in the extracellular environment, and they reveal a novel role for S1P as an inhibitor of ATX, in addition to its well established role as a receptor ligand.

Electrophysiological and ultrastructural changes in severed motor axons of the crayfish.

In many invertebrates, distal stumps of severed axons degenerate slowly and survive for long periods of time; this lengthy process allows the study of the physiological and structural changes involved in axonal degeneration processes. The following experiments demonstrate a reduction in EPSP amplitude, an increase in the distance between neighboring release sites, an extended duration of transmitter release, and a doubling in the average number of quanta released per stimulus at each release site. Ultrastructural examination of those stumps revealed various degrees of glial cell invasion. In the same distal stump, some axons were partially filled with glial cells, but adjacent axons could be completely filled by them. Glial cell invasion was greater at regions closer to the site of axotomy and increased as time progressed. The glia engulfing the stumps exhibited hypertrophy and changes in nuclear morphology. The nuclei of some of those glia cells were unusually close to the axonal membrane in the distal stumps. In spite of these severe morphological changes, the stumps were still capable of conducted action potentials and releasing transmitter at their synapses.

The role of integrin receptors in aspects of glioma invasion in vitro

Integrins are heterodimers consisting of non-covalently associated alpha and beta subunits. They mediate adherence of normal and tumour cells to the extracellular matrix, a property which is essential for migration of neoplastic astrocytes as they invade into the normal brain parenchyma. Flow cytometry and immunocytochemical analysis of cultured cells derived from 10 gliomas (1 pilocytic astrocytoma, 1 astrocytoma, 1 oligoastrocytoma, 1 anaplastic oligoastrocytoma, 4 anaplastic astrocytomas and 2 glioblastoma multiforme) revealed that the β1 integrin subunit was generally expressed more strongly than α4 or αv integrin subunits. Subsequent studies with function-blocking antibodies against the β1 subunit inhibited adhesion, motility and invasion of the gliomas in vitro, to varying degrees, on all extracellular matrix substrates investigated (laminin, collagen type IV, fibronectin and vitronectin), the inhibition by β1 subunit was greatest on collagen type IV. These studies therefore substantiate the case for a role of the β1 integrin subunit in neoplastic glial cell invasion of the brain.

Glial cell involvement in vascular occlusion of diabetic retinopathy.

Twenty areas of retinal vascular occlusion from ten eyes of 6 diabetic patients were studied by immunohistochemistry to type IV collagen (basement membranes), von Willebrand factor (endothelial cells), and to glial fibrillary acid protein (glial cells) on serial sections. In all studied lesions immunoreactivity to type IV collagen and von Willebrand factor was confined to the retinal vascular walls whereas the material accumulated to occlude the vascular lumens centrally displayed immunoreactivity to glial fibrillary acid protein. All arterioles observed in the lesions were occluded. These arterioles had retained their circular shape, and the intravascular glial protein immunoreactivity communicated with the extravascular glial tissue through localised breaks in the vascular wall. The intravascular immunoreactivity was found to continue inside the arteriole along its successive diminishing to reach the capillary level. The venules were only occluded in less than half of the studied lesions. These venules were collapsed to assume a bean-like shape, and sequences with total obliteration of the vascular lumen alternated with sequences where a residual space corresponding to the former lumen displayed immunoreactivity to glial protein. The paper suggests that glial cell invasion, but not endothelial cells or basement membrane thickening, occludes the vascular lumen in areas of retinal non-perfusion secondary to diabetic retinopathy.