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Glioblastoma multiforme (GBM) is the most common and deadliest primary brain tumor in adults. Despite the advances in GBM treatment, outcomes remain poor, with a 2-year survival rate of less than 5%. Hyperbaric oxygen (HBO) therapy is an intermittent, high-concentration, short-term oxygen therapy used to increase cellular oxygen content. In this study, we evaluated the effects of HBO therapy, alone or combined with other treatment modalities, on GBM in vitro and in vivo. In the in vitro analysis, we used a 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay to assess the effects of HBO therapy alone, a colony formation assay to analyze the effects of HBO therapy combined with radiotherapy and with temozolomide (TMZ), and a neurosphere assay to assess GBM stemness. In the in vivo analysis, we used immunohistochemical staining and in vivo bioluminescence imaging to assess GBM stemness and the therapeutic effect of HBO therapy alone or combined with TMZ or radiotherapy, respectively. HBO therapy did not affect GBM cell viability, but it did reduce the analyzed tumors’ ability to form cancer stem cells. In addition, HBO therapy increased GBM sensitivity to TMZ and radiotherapy both in vitro and in vivo. HBO therapy did not enhance tumor growth and exhibited adjuvant effects to chemotherapy and radiotherapy through inhibiting GBM stemness. In conclusion, HBO therapy shows promise as an adjuvant treatment for GBM by reducing cancer stem cell formation and enhancing sensitivity to chemotherapy and radiotherapy.

This study aimed to investigate the regulatory role of Aldo-keto reductase family 1 member B1 (AKR1B1) in glioma cell proliferation through p38 MAPK activation to control Bcl-2/BAX/caspase-3 apoptosis signaling. AKR1B1 expression was quantified in normal human astrocytes, glioblastoma multiforme (GBM) cell lines, and normal tissues by using quantitative real-time polymerase chain reaction. The effects of AKR1B1 overexpression or knockdown and those of AKR1B1-induced p38 MAPK phosphorylation and a p38 MAPK inhibitor (SB203580) on glioma cell proliferation were determined using an MTT assay and Western blot, respectively. Furthermore, the AKR1B1 effect on BAX and Bcl-2 expression was examined in real-time by Western blot. A luminescence detection reagent was also utilized to identify the effect of AKR1B1 on caspase-3/7 activity. The early and late stages of AKR1B1-induced apoptosis were assessed by performing Annexin V-FITC/PI double-staining assays. AKR1B1 expression was significantly downregulated in glioma tissues and GBM cell lines (T98G and 8401). Glioma cell proliferation was inhibited by AKR1B1 overexpression but was slightly increased by AKR1B1 knockdown. Additionally, AKR1B1-induced p38 MAPK phosphorylation and SB203580 reversed AKR1B1′s inhibitory effect on glioma cell proliferation. AKR1B1 overexpression also inhibited Bcl-2 expression but increased BAX expression, whereas treatment with SB203580 reversed this phenomenon. Furthermore, AKR1B1 induced caspase-3/7 activity. The induction of early and late apoptosis by AKR1B1 was confirmed using an Annexin V-FITC/PI double-staining assay. In conclusion, AKR1B1 regulated glioma cell proliferation through the involvement of p38 MAPK-induced BAX/Bcl-2/caspase-3 apoptosis signaling. Therefore, AKR1B1 may serve as a new therapeutic target for glioma therapy development.

Numerous studies have considered galectin-3 or Glycogen synthase kinase 3 beta (GSK3B) as a potential prognosis marker for various cancers. However, the correlation between the protein expression of galectin-3/GSK3B and the clinical parameters of astrocytoma has not been reported. This study aims to validate the correlation between the clinical outcomes and protein expression of galectin-3/GSK3B in astrocytoma. Immunohistochemistry staining was performed to detect galectin-3/GSK3B protein expression in patients with astrocytoma. The Chi-square test, Kaplan−Meier evaluation, and Cox regression analysis were used to determine the correlation between clinical parameters and galectin-3/GSK3B expression. Cell proliferation, invasion, and migration were compared between a non-siRNA group and a galectin-3/GSK3B siRNA group. Protein expression in galectin-3 or GSK3B siRNA-treated cells was evaluated using western blotting. Galectin-3 and GSK3B protein expression were significantly positively correlated with the World Health Organization (WHO) astrocytoma grade and overall survival time. Multivariate analysis revealed that WHO grade, galectin-3 expression, and GSK3B expression were independent prognostic factors for astrocytoma. Galectin-3 or GSK3B downregulation induced apoptosis and decreased cell numbers, migration, and invasion. siRNA-mediated gene silencing of galectin-3 resulted in the downregulation of Ki-67, cyclin D1, VEGF, GSK3B, p-GSK3B Ser9 (p-GSK3B S9), and β-catenin. In contrast, GSK3B knockdown only decreased Ki-67, VEGF, p-GSK3B S9, and β-catenin protein expression but did not affect cyclin D1 and galectin-3 protein expression. The siRNA results indicated that GSK3B is downstream of the galectin-3 gene. These data support that galectin-3 mediated tumor progression by upregulating GSK3B and β-catenin protein expression in glioblastoma. Therefore, galectin-3 and GSK3B are potential prognostic markers, and their genes may be considered to be anticancer targets for astrocytoma therapy.

Although the expression of p53 and epidermal growth factor receptor (EGFR) is associated with therapeutic resistance and patient outcomes in many malignancies, the relationship in astrocytomas is unclear. This study aims to correlate p53 and EGFR expression in brain astrocytomas with overall patient survival. Eighty-two patients with astrocytomas were enrolled in the study. Semi-quantitative p53 and EGFR immunohistochemical staining was measured in tumor specimens. The mean follow-up after astrocytoma surgery was 18.46 months. The overall survival rate was 83%. Survival was reduced in EGFR-positive patients compared with survival in EGFR-negative patients (p < 0.05). However, no significant differences in survival were detected between patients with high and low p53 expression. In patients with low p53 expression, positive EGFR staining was associated with significantly worse survival compared with patients with negative EGFR staining (log-rank test: p < 0.001). Survival rates in positive and negative EGFR groups with high p53 protein expression were similar (log-rank test: p = 0.919). The IC50 of an EGFR inhibitor was higher in GBM cells with high p53 protein expression compared with the IC50 in cells with low p53 expression. Combined EGFR and p53 expression may have prognostic significance in astrocytomas.