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Invasive glioblastoma cells acquire stemness and increased Akt activation.

ABSTRACT: Glioblastoma multiforme (GBM) is the most frequent and most aggressive brain tumor in adults. The dismal prognosis is due to postsurgery recurrences arising from escaped invasive tumor cells. The signaling pathways activated in invasive cells are under investigation, and models are currently designed in search for therapeutic targets. We developed here an in vivo model of human invasive GBM in mouse brain from a GBM cell line with moderate tumorigenicity that allowed simultaneous primary tumor growth and dispersal of tumor cells in the brain parenchyma. This strategy allowed for the first time the isolation and characterization of matched sets of tumor mass (Core) and invasive (Inv) cells. Both cell populations, but more markedly Inv cells, acquired stem cell markers, neurosphere renewal ability, and resistance to rapamycin-induced apoptosis relative to parental cells. The comparative phenotypic analysis between Inv and Core cells showed significantly increased tumorigenicity in vivo and increased invasion with decreased proliferation in vitro for Inv cells. Examination of a large array of signaling pathways revealed extracellular signal-regulated kinase (Erk) down-modulation and Akt activation in Inv cells and an opposite profile in Core cells. Akt activation correlated with the increased tumorigenicity, stemness, and invasiveness, whereas Erk activation correlated with the proliferation of the cells. These results underscore complementary roles of the Erk and Akt pathways for GBM proliferation and dispersal and raise important implications for a concurrent inhibitory therapy.

SUBMITTER: Molina JR

PROVIDER: S-EPMC2887086 | biostudies-other | 2010 Jun

REPOSITORIES: biostudies-other

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Invasive glioblastoma cells acquire stemness and increased Akt activation.

Molina Jennifer R JR Hayashi Yuho Y Stephens Clifton C Georgescu Maria-Magdalena MM

Neoplasia (New York, N.Y.) 20100601 6

Glioblastoma multiforme (GBM) is the most frequent and most aggressive brain tumor in adults. The dismal prognosis is due to postsurgery recurrences arising from escaped invasive tumor cells. The signaling pathways activated in invasive cells are under investigation, and models are currently designed in search for therapeutic targets. We developed here an in vivo model of human invasive GBM in mouse brain from a GBM cell line with moderate tumorigenicity that allowed simultaneous primary tumor g ...[more]

PMID: 20563248

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Project description:Despite the functional role of serglycin as an intracellular proteoglycan, a variety of malignant cells depends on its expression and constitutive secretion to advance their aggressive behavior. Serglycin arose to be a biomarker for glioblastoma, which is the deadliest and most treatment-resistant form of brain tumor, but its role in this disease is not fully elucidated. In our study we suppressed the endogenous levels of serglycin in LN-18 glioblastoma cells to decipher its involvement in their malignant phenotype. Serglycin suppressed LN-18 (LN-18shSRGN) glioblastoma cells underwent astrocytic differentiation characterized by induced expression of GFAP, SPARCL-1 and SNAIL, with simultaneous loss of their stemness capacity. In particular, LN-18shSRGN cells presented decreased expression of glioma stem cell-related genes and ALDH1 activity, accompanied by reduced colony formation ability. Moreover, the suppression of serglycin in LN-18shSRGN cells retarded the proliferative and migratory rate, the invasive potential in vitro and the tumor burden in vivo. The lack of serglycin in LN-18shSRGN cells was followed by G2 arrest, with subsequent reduction of the expression of cell-cycle regulators. LN-18shSRGN cells also exhibited impaired expression and activity of proteolytic enzymes such as MMPs, TIMPs and uPA, both in vitro and in vivo. Moreover, suppression of serglycin in LN-18shSRGN cells eliminated the activation of pro-tumorigenic signal transduction. Of note, LN-18shSRGN cells displayed lower expression and secretion levels of IL-6, IL-8 and CXCR-2. Concomitant, serglycin suppressed LN-18shSRGN cells demonstrated repressed phosphorylation of ERK1/2, p38, SRC and STAT-3, which together with PI3K/AKT and IL-8/CXCR-2 signaling control LN-18 glioblastoma cell aggressiveness. Collectively, the absence of serglycin favors an astrocytic fate switch and a less aggressive phenotype, characterized by loss of pluripotency, block of the cell cycle, reduced ability for ECM proteolysis and pro-tumorigenic signaling attenuation.

Project description:BackgroundGlioblastoma is devastating cancer with a high frequency of occurrence and poor survival rate and it is urgent to discover novel glioblastoma-specific antigens for the therapy. Cancer-germline genes are known to be related to the formation and progression of several cancer types by promoting tumor transformation. Dazl is one such germline gene and is up-regulated in a few germ cell cancers. In this study, we analyzed the expression of Dazl in human glioblastoma tissues and cells, and investigated its significance in proliferation, migration, invasion and chemoresistance of the glioblastoma cell lines.MethodsWe evaluated the expression of Dazl in different pathologic grades of glioblastoma tissues by immunohistochemistry. We assessed the expression of Dazl in glioblastoma cells and normal human astrocytes (NHA) cells by western blotting and RT-qPCR. Then we generated Dazl knockout glioblastoma cell lines using the CRISPR/Cas9 gene-editing technology to explore the cellular function of Dazl. We detected the proliferation and germline traits via CCK-8 assays and alkaline phosphatase staining, respectively. Boyden chamber assays were performed to measure glioblastoma cell migration and invasion. Crystal violet staining was used to determine the number of viable cells after the treatment of Doxorubicin and Temozolomide. Finally, we used subcutaneous xenograft studies to measure the growth of tumors in vivo.ResultsWe found that Dazl was upregulated in glioblastoma tissues and glioblastoma cell lines. Dazl knockdown glioblastoma cells showed decreased cellular proliferation, migration, invasion, and resistance in vitro, and inhibited the initiation of glioblastoma in vivo. The glioblastoma cell lines A172, U251, and LN229 were found to express stem cell markers CD133, Oct4, Nanog, and Sox2. The expression of these markers was downregulated in Dazl-deficient cells.ConclusionsOur results indicated that Dazl contributes to the tumorigenicity of glioblastoma via reducing cell stemness. Therefore, cancer-germline genes might represent a new paradigm of glioblastoma-initiating cells in the treatment of malignant tumors.

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Invasive glioblastoma cells acquire stemness and increased Akt activation.

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Invasive glioblastoma cells acquire stemness and increased Akt activation.

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