Novel approaches for quantifying protein biomarkers in gliomas: benefits and pitfalls

Dahlrot, Rikke Hedegaard; Sørensen, Mads D.; Rosager, Ann Mari; Hellwege, Sofie; Bangsø, Julie A; Rosenberg, Tine; Petterson, Stine Asferg; Klitkou, J.; Fosmark, Sigurd; Hansen, Steinbjørn; Kristensen, Bjarne Winther

doi:10.2217/cns.14.30

Cited by 11 publications

(10 citation statements)

References 52 publications

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“…However, in both studies semi-quantitative pathologist-based scoring systems was used and no statistical comparison was made. To provide a more unbiased quantification approach we used blinded quantitative stereology and systematic random sampling [29]. Moreover, we focused on and compared astrocytic tumors only.…”

Section: Discussionmentioning

confidence: 99%

Expression and Prognostic Value of Oct-4 in Astrocytic Brain Tumors

et al. 2016

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BackgroundGlioblastomas are the most frequent type of malignant primary brain tumor with a median overall survival less than 15 months. Therapy resistance of glioblastomas has been attributed to the presence of tumor initiating stem-like cells (TSCs). TSC-related markers have therefore been suggested to have promising potentials as prognostic markers in gliomas.Methodology/Principal FindingsThe aim of the present study was to investigate the expression and prognostic impact of the TSC-related marker Oct-4 in astrocytic brain tumors of increasing grade. In total 114 grade II, III and IV astrocytic brain tumors were immunohistochemically stained for Oct-4, and the fraction and intensity of Oct-4 positive cells were determined by morphometric analysis of full tumor sections. Oct-4 was expressed in all tumors, and the Oct-4 positive cell fraction increased with tumor grade (p = 0.045). There was no association between survival and Oct-4 positive cell fraction, neither when combining all tumor grades nor in analysis of individual grades. Oct-4 intensity was not associated with grade, but taking IDH1 status into account we found a tendency for high Oct-4 intensity to be associated with poor prognosis in anaplastic astrocytomas. Double immunofluorescence stainings showed co-localization in the perivascular niches of Oct-4 and two other TSC markers CD133 and nestin in glioblastomas. In some areas Oct-4 was expressed independently of CD133 and nestin.ConclusionsIn conclusion, high Oct-4 fraction was associated with tumor malignancy, but seemed to be without independent prognostic influence in glioblastomas. Identification of a potential prognostic value in anaplastic astrocytomas requires additional studies using larger patient cohorts.

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Section: Discussionmentioning

confidence: 99%

Expression and Prognostic Value of Oct-4 in Astrocytic Brain Tumors

et al. 2016

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“…Omission of primary antibodies served as negative controls as well as controls for nonspecific staining induced by the detection systems alone. Reactions were evaluated digitally by measuring the staining intensities of the spheroids using the VIS software (Visiopharm, Hørsholm, Denmark) [ 46 , 47 , 52 ]. Briefly, 20 randomly selected spheroids were marked as regions of interest.…”

Section: Methodsmentioning

confidence: 99%

“…To monitor stem cell differentiation, we analyzed the expression of three stem cell markers, CD133 [ 32–38 ], nestin [ 33 , 37–43 ] and musashi-1 [ 35 , 38 , 42 , 44 , 45 ]. Expression of all markers was investigated by chromogenic immunohistochemical staining and quantified digitally in order to obtain precise less observer-dependent measurements [ 46 , 47 ].…”

mentioning

confidence: 99%

Heterogenic expression of stem cell markers in patient-derived glioblastoma spheroid cultures exposed to long-term hypoxia

et al. 2018

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Aim:To investigate the time profile of hypoxia and stem cell markers in glioblastoma spheroids of known molecular subtype.Materials & methods:Patient-derived glioblastoma spheroids were cultured up to 7 days in either 2% or 21% oxygen. Levels of proliferation (Ki-67), hypoxia (HIF-1α, CA9 and VEGF) and stem cell markers (CD133, nestin and musashi-1) were investigated by immunohistochemistry.Results:Hypoxia markers as well as CD133 and partially nestin increased in long-term hypoxia. The proliferation rate and spheroid size were highest in normoxia.Conclusion:We found differences in hypoxia and stem cell marker profiles between the patient-derived glioblastoma cultures. This heterogeneity should be taken into consideration in development of future therapeutic strategies.

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“…Sections stained with IgG or sections where primary antibodies were omitted were devoid of staining. IHC double staining was performed using the BenchMark fully automated staining instrument and the Vectastain Universal Elite ABC kit (Vector Labs, USA) [ 14 ]. Sections were counterstained with Mayer’s hematoxylin (Bie & Berntsen, Herlev, Denmark).…”

Section: Methodsmentioning

confidence: 99%

Cell therapy centered on IL-1Ra is neuroprotective in experimental stroke

et al. 2016

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Cell-based therapies are emerging as new promising treatments in stroke. However, their functional mechanism and therapeutic potential during early infarct maturation has so far received little attention. Here, we asked if cell-based delivery of the interleukin-1 receptor antagonist (IL-1Ra), a known neuroprotectant in stroke, can promote neuroprotection, by modulating the detrimental inflammatory response in the tissue at risk. We show by the use of IL-1Ra-overexpressing and IL-1Ra-deficient mice that IL-1Ra is neuroprotective in stroke. Characterization of the cellular and spatiotemporal production of IL-1Ra and IL-1α/β identifies microglia, not infiltrating leukocytes, as the major sources of IL-1Ra after experimental stroke, and shows IL-1Ra and IL-1β to be produced by segregated subsets of microglia with a small proportion of these cells co-expressing IL-1α. Reconstitution of whole body irradiated mice with IL-1Ra-producing bone marrow cells is associated with neuroprotection and recruitment of IL-1Ra-producing leukocytes after stroke. Neuroprotection is also achieved by therapeutic injection of IL-1Ra-producing bone marrow cells 30 min after stroke onset, additionally improving the functional outcome in two different stroke models. The IL-1Ra-producing bone marrow cells increase the number of IL-1Ra-producing microglia, reduce the availability of IL-1β, and modulate mitogen-activated protein kinase (MAPK) signaling in the ischemic cortex. The importance of these results is underlined by demonstration of IL-1Ra-producing cells in the human cortex early after ischemic stroke. Taken together, our results attribute distinct neuroprotective or neurotoxic functions to segregated subsets of microglia and suggest that treatment strategies increasing the production of IL-1Ra by infiltrating leukocytes or microglia may also be neuroprotective if applied early after stroke onset in patients.Electronic supplementary materialThe online version of this article (doi:10.1007/s00401-016-1541-5) contains supplementary material, which is available to authorized users.

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Novel approaches for quantifying protein biomarkers in gliomas: benefits and pitfalls

Cited by 11 publications

References 52 publications

Expression and Prognostic Value of Oct-4 in Astrocytic Brain Tumors

Expression and Prognostic Value of Oct-4 in Astrocytic Brain Tumors

Heterogenic expression of stem cell markers in patient-derived glioblastoma spheroid cultures exposed to long-term hypoxia

Cell therapy centered on IL-1Ra is neuroprotective in experimental stroke

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