Lower dose of metformin combined with artesunate induced autophagy-dependent apoptosis of glioblastoma by activating ROS-AMPK-mTOR axis.

Glioblastoma multiform (GBM), one of the most common, aggressive primary brain tumours, demonstrates resistance to radiotherapy and chemotherapy after surgical resection and treatment failure. Metformin (MET) has been shown to suppress the proliferative capacity and invasion ability of GBM cells by activating AMPK and inhibiting mTOR, but the effective dose exceeded the maximum tolerated dose. Artesunate (ART) can exert certain anti-tumour effects by activating the AMPK-mTOR axis and inducing autophagy in tumour cells. Therefore, this study investigated the effects of MET combined with ART combination therapy on autophagy and apoptosis in GBM cells. MET combined with ART treatment effectively suppressed the viability, mono-cloning ability, migration and invasion capacities, as well as metastatic ability of GBM cells. The underlying mechanism involved modulation of the ROS-AMPK-mTOR axis, which was confirmed using 3-methyladenine and rapamycin to inhibit or promote the effects of MET combined with ART, respectively. The study findings suggest that MET used in combination with ART can induce autophagy-dependent apoptosis in GBM cells by activating the ROS-AMPK-mTOR pathway, providing a potential new treatment for GBM.

Tetrandrine alleviates inflammation and neuron apoptosis in experimental traumatic brain injury by regulating the IRE1α/JNK/CHOP signal pathway.

AIM: The aim of this study was to investigate the therapeutic roles of Tetrandrine (TET) on traumatic brain injury (TBI) and the underlying mechanism. METHOD: Traumatic injury model of hippocampal neurons and TBI mouse model were established to evaluate the therapeutic effects. The expression of neuron-specific enolase (NSE), Caspase 3, and Caspase 12 was detected by immunofluorescence. The expression of TNF-α, NF-κB, TRAF1, ERS markers (GADD34 and p-PERK), IRE1α, CHOP, JNK, and p-JNK were evaluated by western blot. Flow cytometry was used to determine the apoptosis of neurons. Brain injury was assessed by Garcia score, cerebral water content, and Evan blue extravasation test. Hematoxylin and eosin staining was used to determine the morphological changes of hippocampal tissue. Apoptosis was assessed by TUNEL staining. RESULT: In traumatic injury model of hippocampal neurons, TET downregulated NSE, TNF-α, NF-κB, TRAF1, GADD34, p-PERK, IRE1α, CHOP, and p-JNK expression. TET reduced Caspase 3 and Caspase 12 cleavage. Apoptosis rate was inhibited by the introduction of TET. TET improved the Garcia neural score, decreased the cerebral water content and Evans blue extravasation, and reduced NSE, TNF-α, NF-κB, TRAF1, IRE1α, CHOP, and p-JNK expression in mice with TBI, which was significantly reversed by Anisomycin, a JNK selective activator. CONCLUSION: TET alleviated inflammation and neuron apoptosis in experimental TBI by regulating the IRE1α/JNK/CHOP signal pathway.

Identification of Prognostic Biomarkers of Glioblastoma Based on Multidatabase Integration and Its Correlation with Immune-Infiltration Cells.

Background: Glioblastoma (GBM) is the most malignant of all known intracranial tumors; meanwhile, most patients have a poor prognosis. In order to improve the poor prognosis of GBM patients as much as possible, it is specifically significant to identify biomarkers related to the gene diagnosis and gene therapy. Methods: In this study, a total of 343 GBM specimens and 259 nontumor specimens were collected from four Gene Expression Omnibus (GEO) datasets and The Cancer Genome Atlas (TCGA) database; then, we analyzed the differentially expressed genes (DEGs) from the above data. Through Venn diagram analysis, 54 common upregulated DEGs and 22 common downregulated DEGs were triumphantly recognized. Results: On the basis of the degree of formation communication in protein-protein interaction network (PPIN), the 10 upregulated central genes were ranked, incorporating LOX, IGFBP3, CD44, TIMP1, FN1, VEGFA, POSTN, COL1A1, COL1A2, and COL3A1. By combining the expression levels and the clinical features of GBM, we found that four hub genes (TIMP1, FN1, POSTN, and LOX) were significantly upregulated and related to poor prognosis of GBM. Meanwhile, univariate and multivariate Cox regression analysis suggested that TIMP1 could be one of the independent prognostic factors for GBM patients. Furthermore, TIMP1 was particularly correlated with the immune marker of macrophage M1, macrophage M2, neutrophils, tumor-associated macrophage, and Tregs. We then analyzed the role of TIMP1 in GBM cancer cell lines by relevant experiments, which indicated that TIMP1 knockdown resulted in the decreased cell proliferation, migration, and invasion. Conclusions: Taken together, these findings demonstrated that TIMP1 might be a new biomarker to determine prognosis and immune infiltration of GBM patients.

Tetrandrine Ameliorates Traumatic Brain Injury by Regulating Autophagy to Reduce Ferroptosis.

Traumatic brain injury (TBI) is the leading cause of death and disability in trauma patients. However, the effects and mechanism of autophagy after TBI remain unclear. This study aimed to investigate whether tetrandrine could ameliorate TBI through autophagy to reduce ferroptosis. A mice model for TBI was implemented. Behavioral and histomorphological experiments were performed to evaluate outcomes of the mice. The ferroptosis levels was detected by Perls staining. Enzyme-linked immunosorbent assay (ELISA) was applied to detect malondialdehyde (MDA), glutathione (GSH), and glutathione peroxidase 4 (GPX4) levels in the brain tissue. Western blot test was performed to detect Beclin 1, light chain 3 (LC3) II/I, p62, GPX4, SCL7A11, and ferritin heavy chain 1 (FTH1) levels, and the expression of LC3B, Beclin 1, GPX4, and FTH1 in the brain tissue was detected by immunofluorescence (IF). The behavioral and histomorphological results demonstrated that tetrandrine improved the neurological function and cerebral edema on days 1, 3, and 7 after TBI. The ELISA results suggested that tetrandrine reduced the MDA concentration and increased GSH concentration on days 1, 3, and 7 after TBI. IF staining and Perls staining reflected that tetrandrine promoted autophagy and inhibited ferroptosis on days 1, 3, and 7 after TBI, respectively. Tetrandrine further improved the neurological function, cerebral edema, autophagy, and ferroptosis on days 1, 3, and 7 after TBI after adding the autophagy inducer rapamycin. The effect of TET in alleviating TBI increased with the increase of time and dose. Tetrandrine ameliorated TBI by regulating autophagy to reduce ferroptosis, providing a new therapeutic strategy for TBI.

Suppression of CLC-3 chloride channel reduces the aggressiveness of glioma through inhibiting nuclear factor-κB pathway.

CLC-3 chloride channel plays important roles on cell volume regulation, proliferation and migration in normal and cancer cells. Recent growing evidence supports a critical role of CLC-3 in glioma metastasis, however, the mechanism underlying is unclear. This study finds that CLC-3 is upregulated in glioma tissues and positively correlated with WHO histological grade. Patients with high CLC-3 expression had an overall shorter survival time, whereas patients with low expression of CLC-3 had a better survival time. Silencing endogenous CLC-3 with ShCLC-3 adenovirus significantly decreases volume-regulated chloride currents, inhibits the nuclear translocation of p65 subunit of Nuclear Factor-κB (NF-κB), decreases transcriptional activity of NF-κB, reduces MMP-3 and MMP-9 expression and decreases glioma cell migration and invasion. Taken together, these results suggest CLC-3 promotes the aggressiveness of glioma at least in part through nuclear factor-κB pathway, and might be a novel prognostic biomarker and therapeutic target for glioma.

PLD1 overexpression promotes invasion and migration and function as a risk factor for Chinese glioma patients.

Glioma is a lethal disease with few effective therapeutic options. Recently, insights into cancer biology had suggested that abnormal lipid metabolism was a risk factor for various human malignancies, including glioma. As a key enzyme implicated in lipid metabolism, PLD1 was overexpression in multiple human cancers, and it was stated to be responsible for aggressive phenotypes, such as angiogenesis and chemoresistance. However, there was still much to know about its expression and function in glioma. In the present study, we showed that PLD1 was overexpression in clinical samples of glioma. In addition, the correlation assay revealed that PLD1 overexpression was correlated with poor differentiation (p = 0.04), and it was responsible for a poor prognosis for the patients (p = 0.009). Furthermore, we showed in COX regression assay that PLD1 was a risk factor for glioma (p = 0.018, HR = 0.461, 95% CI = 0.243-0.887). Consistently, we found that PLD1 was overexpression in glioma cell lines, and it could facilitate the proliferation and migration. Taken together, our study suggested that PLD1 was pro-tumoral in glioma, and that further studies were urgently needed so as to define whether it was a novel therapeutic target for the disease.

Hippocampal Src kinase is required for novelty-induced enhancement of contextual fear extinction.

Exposure to a novel environment enhances the extinction of contextual fear through the "tagging-and-capture" process. However, the underlying molecular mechanisms of novelty-induced enhancement of fear extinction are still unclear. NMDA receptor activity was recently revealed to be required for the enhancement of fear extinction caused by exposure to novelty. Src family kinases (SFKs) act as a molecular hub for regulation of NMDA receptors. We hypothesized that SFKs might be involved in novelty-induced enhancement of fear extinction. We found that the enhancement of fear extinction induced by novelty exposure is accompanied by Src kinase phosphorylation and activation in a restricted time window. Furthermore, intrahippocampal infusion of SFKs inhibitor PP2 inhibits Src kinase phosphorylation and activation, attenuates the activation of NR2B-containing NMDA receptors, and thereby reverses the enhancement of fear extinction induced by novelty exposure. These results suggested that Src kinase may serve as a behavioral tag in the procedural enhancement of fear extinction by novelty exposure.