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. 2018 Oct 31;9(23):4496-4502.
doi: 10.7150/jca.26769. eCollection 2018.

GFAP Expression Is Influenced by Astrocytoma Grade and rs2070935 Polymorphism

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Free PMC article

GFAP Expression Is Influenced by Astrocytoma Grade and rs2070935 Polymorphism

Mantas Sereika et al. J Cancer. .
Free PMC article

Abstract

Glial fibrillary acidic protein (GFAP) is an intermediate filament that provides mechanical support to astrocytes. Rs2070935 is a single nucleotide polymorphism (SNP) located in the promoter region of the GFAP gene. The aim of this pilot study is to investigate GFAP expression at mRNA, protein levels and rs2070935 polymorphism in 50 different grade human astrocytoma samples. GFAP expression at mRNA level was measured using quantitative reverse transcription polymerase chain reaction (qRT-PCR) with SYBR Green dye, whereas the translational activity of the following gene was detected using western blot assay. Furthermore, genotypes of rs2070935 were identified using qPCR with TaqMan probes. As a result, GFAP mRNA and protein expression was found to be declining with increasing astrocytoma grade (p < 0.05). A tendency was observed between increased GFAP mRNA expression and shorter grade IV astrocytoma patient survival (p = 0.2117). The rs2070935 CC genotype was found to be associated with increased GFAP translational activity in grade II astrocytoma (p = 0.0238). Possible links between rs2070935 genotypes and alternative splicing of GFAP were also observed. The rs2070935 AA genotype was found to be associated with poor clinical outcome for grade IV astrocytoma patients (p = 0.0007), although the following data should be checked in a larger sample size of astrocytoma patients.

Keywords: GFAP; glioblastoma; rs2070935.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interest exists.

Figures

Figure 1
Figure 1
GFAP expression varies in different grades of human astrocytoma. A) Transcriptional activity of GFAP, as detected by qRT-PCR assay and grouped by tumor grade. B) Kaplan‑Meier curve for grade IV astrocytoma patient overall survival in accordance to GFAP mRNA expression. Increased GFAP expression - patient group, exhibiting higher GFAP mRNA expression than the reference human brain control (Log2(2‑ΔΔCt) > 0). Decreased GFAP expression - patient group, featuring lower GFAP mRNA expression than the reference human brain control (Log2(2‑ΔΔCt) < 0) . C) Translational activity of GFAP, as observed by western blot assay and grouped by tumor grade. D) GFAP and Beta-actin protein bands, as captured by western blot assay. The molecular weight of GFAP isoforms ranges from 50 to 38 kDa, whereas Beta-actin protein is at approximately 42 kDa. In all graphs GFAP expression values at mRNA and protein levels were normalized to that of the reference gene (Beta-actin). GFAP transcriptional activity for tumor samples is in relation to the reference human brain control.
Figure 2
Figure 2
Rs2070935 polymorphism is linked with varying GFAP expression and different grade IV astrocytoma patient survival. A) Kaplan‑Meier curve for grade IV astrocytoma patient overall survival in accordance to rs2070935 genotypes. B) GFAP transcriptional activity, as detected by qRT-PCR assay and subdivided by genotypes of rs2070935 as well as tumor grade. C) GFAP translational activity, as observed by western blot assay and grouped by rs2070935 polymorphism as well as astrocytoma grade. D) Correlation between GFAP transcriptional and translational activities in the group of rs2070935 CA heterozygotes. In all graphs GFAP expression values at mRNA and protein levels were normalized to that of the reference gene (Beta-actin). GFAP transcriptional activity for tumor samples is in relation to the reference human brain control.
Figure 3
Figure 3
Rs2070935 polymorphism is associated with different GFAP isoform quantities in human astrocytoma. 38 kDa proportion (%) - a percentile value, which represents the signal intensity of the 38 kDa GFAP isoform, compared to the signal intensity of the whole sample.

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