GRIK1 promotes glioblastoma malignancy and is a novel prognostic factor of poor prognosis.

Primary tumors of the central nervous system (CNS) are classified into over 100 different histological types. The most common type of glioma is derived from astrocytes, and the most invasive glioblastoma (WHO IV) accounts for over 57% of these tumors. Glioblastoma (GBM) is the most common and fatal tumor of the CNS, with strong growth and invasion capabilities, which makes complete surgical resection almost impossible. Despite various treatment methods such as surgery, radiotherapy, and chemotherapy, glioma is still an incurable disease, and the median survival time of patients with GBM is shorter than 15 months. Thus, molecular mechanisms of GBM characteristic invasive growth need to be clarified to improve the poor prognosis. Glutamate ionotropic receptor kainate type subunit 1 (GRIK1) is essential for brain function and is involved in many mental and neurological diseases. However, GRIK1's pathogenic roles and mechanisms in GBM are still unknown. Single-nuclear RNA sequencing of primary and recurrent GBM samples revealed that GRIK1 expression was noticeably higher in the recurrent samples. Moreover, immunohistochemical staining of an array of GBM samples showed that high levels of GRIK1 correlated with poor prognosis of GBM, consistent with The Cancer Genome Atlas database. Knockdown of GRIK1 retarded GBM cells growth, migration, and invasion. Taken together, these findings show that GRIK1 is a unique and important component in the development of GBM and may be considered as a biomarker for the diagnosis and therapy in individuals with GBM.

Early outcomes of endoscopic endonasal approach pituitary adenomas resection with minimal nasal injury.

ABSTRACT: To report the results of a consecutive series of pituitary adenomas resected through endoscopic endonasal approach (EEA) with minimal nasal injury.Retrospectively review tumor characteristics and surgical outcomes of a consecutive series of EEA pituitary adenomas resection performed mainly by a single author between March 2018 and June 2019.A total of 75 endoscopic endonasal approach pituitary adenoma resections were performed by the authors' team. Of the 75 patients, 28 through mononostril EEA, 47 through Binonostril EEA. Hadad-Bassagasteguy vascularized nasoseptal flap was harvested in only 4 (5.3%) patients with a high risk of postoperative cerebrospinal fluid leak, and one side middle turbinate only been resected in 2 (2.7%) patients, other patients preserved bilateral middle turbinate. Of the 75 patients, gross total resection is 74.7%, near-total resection is 16.0%. Endocrinological remission was achieved in 76.9% of GH-secreting adenomas, 61.5% of prolactin-secreting adenomas. The postoperative cerebrospinal fluid leak rate was 2.7%. Two patients had suprasellar hemorrhage, 1 patient had perioperative stroke, 2 patients had permanent diabetes insipidus, no cranial nerve deficits, internal carotid artery injury, anosmia, and death. The sino-nasal function was measured with the Sino-Nasal Outcome Test-22 and visual analog scale for olfaction preoperatively and postoperatively, and there was no statistically significant difference.The EEA is an effective approach to resect pituitary adenomas, the gross total resection and near-total resection rate and endocrinological remission rate are satisfactory. The EEA is a safe approach, as the complication rate is acceptable compared with those reported in the previous series of microscopic and endoscopic approaches. These results can be achieved with minimal nasal injury.

Texture Analysis of High b-Value Diffusion-Weighted Imaging for Evaluating Consistency of Pituitary Macroadenomas.

BACKGROUND: Preoperative evaluation of the consistency of pituitary macroadenomas is important for neurosurgeons to prepare the surgical plan. PURPOSE: To evaluate the diagnostic performance of texture analysis (TA) of diffusion-weighted imaging (DWI) at a standard b-value (b = 1000 s/mm2 ) and a high b-value (b = 2000 s/mm2 ) for their ability to assess the tumor consistency of pituitary macroadenomas. STUDY TYPE: Retrospective. POPULATION/SUBJECTS: Fifty patients with histologically confirmed pituitary macroadenomas were classified as soft (n = 37) or hard (n = 13) types. FIELD STRENGTH/SEQUENCE: Coronal T2 -weighted imaging (T2 WI), Readout Segmentation of Long Variable Echo-trains (RESOLVE) DWI at b = 1000 s/mm2 and b = 2000 s/mm2 were acquired with 3.0T MRI. ASSESSMENT: The corresponding apparent diffusion coefficient (ADC) maps (ADC1000 and ADC2000 ) were registered to T2 WI. Regions of interest (ROIs) were manually drawn along the solid part of the tumor from the coregistered T2 WI-ADC images. The texture parameters from T2 WI, ADC1000 , and ADC2000 were acquired. STATISTICAL TESTS: The texture parameters were compared between the two types by using unpaired Student's t-test. Receiver operating characteristic (ROC) curves and logistic regression analyses were used to assess their diagnostic performance. RESULTS: Significant differences in TA parameters of ADC1000 and ADC2000 were observed between soft and hard types (P < 0.05 for all), whereas the TA of T2 WI resulted in no significant difference (P > 0.05 for all). TA of ADC2000 provided a superior diagnostic performance compared with that of ADC1000 (P = 0.038). A combination of mean value and entropy of ADC2000 yielded an AUC, a sensitivity, and a specificity of 0.911, 78.4% and 92.3%, respectively. DATA CONCLUSION: TA of ADC values were useful for assessing the tumor consistency of pituitary macroadenomas. ADC2000 may facilitate better type discrimination. LEVEL OF EVIDENCE: 3 Technical Efficacy Stage: 2 J. Magn. Reson. Imaging 2020;51:1507-1513.

MiR-184 Retarded the Proliferation, Invasiveness and Migration of Glioblastoma Cells by Repressing Stanniocalcin-2.

To investigate the repression of miR-184 on Stanniocalcin-2 (STC2) and how this axis affects the propagation, invasiveness and migration ability of glioblastoma cells. RT-PCR was employed to determine the miR-184 and STC2 mRNA expression both in tissues and cells. Western blot was employed to determine the protein expression levels. The cells were transfected via lipofection. MTT, colony formation, invasion and scratch healing assays were conducted to study the propagation, invasiveness and migratory ability of glioblastoma cells, respectively. The dual luciferase reporter gene assay was conducted to determine whether miR-184 could directly bind to STC2 mRNA 3'UTR. MiR-184 was under-expressed whereas STC2 was over-expressed in glioblastoma tissues and cell line. The up-regulation of miR-184 significantly suppressed the propagation, migratory ability and invasion of glioblastoma cells, whereas the over-expression of STC2 restored this effect. MiR-184 was confirmed to directly target STC2. MiR-184 could retard the propagation, invasiveness and migratory ability of glioblastoma cells by suppressing STC2.

MicroRNA-622 suppresses the proliferation of glioma cells by targeting YAP1.

It has recently been shown that miR-622 plays a tumor suppressive role in many human cancers. However, the exact function and underlying mechanism are still unknown. Here, we reported that the level of miR-622 is clearly reduced in human glioma tissues in comparison with normal brain tissues and is negatively correlated with the histological grades. Additionally, ectopically expressed miR-622 significantly inhibited cell proliferation and induced cell cycle arrest at the G0/G1 phase in glioma cells. Furthermore, the bioinformatics analysis revealed that YAP1 possesses putative miR-622-binding sites within its 3'UTR. Consequently, an elevated miR-622 level was found to suppress the luciferase reporter activity of YAP1 3'UTR, and the effect was diminished by the deletion of the miR-622 seed binding site. In addition, the level of YAP1 protein expression was significantly decreased after the overexpression of miR-622. These results indicate a negative link between miR-622 and YAP1 and further confirm that YAP1 is a direct target of miR-622, suggesting that miR-622 could be a new important therapeutic strategy for gliomas treatment.

miR-330-5p suppresses glioblastoma cell proliferation and invasiveness through targeting ITGA5.

The present study intended to investigate the biological effects of miR-330-5p on glioblastoma (GBM) cell proliferation and invasiveness by targeting integrin α5 (ITGA5). The expressions of miR-330-5p and ITGA5 mRNA in GBM cell lines (U87, U251, and U373) and normal brain glial cell line (HEB) were detected using RT-qPCR. Protein expression of ITGA5 was examined using Western blot. The present study used MTT assay, colony formation assay, Transwell assay, wound healing assay, and flow cytometry analysis in order to determine the biological functions of GBM cells (including cell proliferation, invasion, migration, apoptosis, and cell cycle). The present study applied dual-luciferase reporter gene assay to identify the target relationship between miR-330-5p and ITGA5. miR-330-5p was low-expressed in GBM cell lines while ITGA5 was high-expressed compared with HEB. miR-330-5p could directly target ITGA5 as well as suppress its expression in GBM cells. Up-regulation of miR-330-5p and down-regulation of ITGA5 both have an inhibitory effect on cell proliferation, invasion, and migration. Meanwhile, they could also promote GBM cell apoptosis. miR-330-5p could suppress proliferation and invasion of GBM cells through targeting ITGA5.

Human herpesvirus 6 latent infection in patients with glioma.

The etiology of glioma remains unclear so far. Human herpesvirus 6 (HHV-6) might be associated with glioma, but there is no direct evidence to support this. High percentages of HHV-6 DNA and protein were detected in tissue from gliomas, compared with normal brain tissue. In addition, a strain of HHV-6A was isolated from the fluid specimens from glioma cysts. High levels of interleukin 6 (IL-6), interleukin 8 (IL-8), tumor necrosis factor α, and transforming growth factor ß (TGF-ß) were detected in the cyst fluid specimens from HHV-6-positive patients with glioma. Furthermore, HHV-6A infection promoted IL-6, IL-8, and TGF-ß production in astrocyte cultures. Our studies strongly suggest the involvement of HHV-6 infection in the pathogenesis of glioma.

The apoptosis-resistance in t-AUCB-treated glioblastoma cells depends on activation of Hsp27.

We previously reported that sEH inhibitor t-AUCB suppresses the growth of human glioblastoma U251 and U87 cell lines and induces cell-cycle G0/G1 phase arrest. In present study, we found even 96 h-treatment of 200 µM t-AUCB can not induce apoptosis in U251 and U87 cells. We also revealed that 200 µM t-AUCB significantly elevates the activation of p38 MAPK, MAPKAPK2 and Hsp27. The p38 MAPK inhibitor SB203580 and the inhibitor of Hsp27 phosphorylation, KRIBB3, were used to investigate the mechanism of the apoptosis-resistance. The results showed that, after blocking the activation of Hsp27 by SB203580 or KRIBB3, 200 µM t-AUCB significantly induces apoptosis and increases caspase-3 activities in U251 and U87 cells. Our data demonstrated that t-AUCB induces cell apoptosis after blocking itself-induced activation of Hsp27, and that the activation of Hsp27 may confer chemoresistance in GBM cells. The combination of t-AUCB and the inhibitor of Hsp27 phosphorylation may be a potential strategy for treatment of glioblastoma.

Anisomycin induces glioma cell death via down-regulation of PP2A catalytic subunit in vitro.

AIM: To examine the effects of anisomycin on glioma cells and the related mechanisms in vitro. METHODS: The U251 and U87 human glioblastoma cell lines were tested. The growth of the cells was analyzed using a CCK-8 cell viability assay. Apoptosis was detected using a flow cytometry assay. The expression of proteins and phosphorylated kinases was detected using Western blotting. RESULTS: Treatment of U251 and U87 cells with anisomycin (0.01-8 µmol/L) inhibited the cell growth in time- and concentration-dependent manners (the IC(50) values at 48 h were 0.233±0.021 and 0.192±0.018 µmol/L, respectively). Anisomycin (4 µmol/L) caused 21.5%±2.2% and 25.3%±3.1% of apoptosis proportion, respectively, in U251 and U87 cells. In the two cell lines, anisomycin (4 µmol/L) activated p38 MAPK and JNK, and inactivated ERK1/2. However, neither the p38 MAPK inhibitor SB203580 (10 µmol/L) nor the JNK inhibitor SP600125 (10 µmol/L) prevented anisomycin-induced cell death. On the other hand, anisomycin (4 µmol/L) reduced the level of PP2A/C subunit (catalytic subunit) in a time-dependent manner in the two cell lines. Treatment of the two cell lines with the PP2A inhibitor okadaic acid (100 nmol/L) caused marked cell death. CONCLUSION: Anisomycin induces glioma cell death via down-regulation of PP2A catalytic subunit. The regulation of PP2A/C exression by anisomycin provides a clue to further study on its role in glioma therapy.

t-AUCB, an improved sEH inhibitor, suppresses human glioblastoma cell growth by activating NF-κB-p65.

Although sEH inhibitors are well studied in inflammatory and cardiovascular diseases, their effects on gliomas are unclear. In this study, we investigated the effects of t-AUCB, a more potent and selective sEH inhibitor, on U251 and U87 human glioblastoma cell lines and the HepG2 human hepatocellular carcinoma cell line. Our results showed that t-AUCB efficiently inhibited sEH activities in all three cell lines (the inhibition rate was more than 80% in each) and suppressed U251 and U87 cell growth in a dose-dependent manner, but exhibited no cell growth inhibition on HepG2. We detected high levels of phosphorylated NF-κB-p65 (Ser536) in t-AUCB-treated U251 and U87 cells, and then found that the NF-κB inhibitor PDTC can completely abolish t-AUCB-induced growth inhibition. This indicated that t-AUCB suppresses U251 and U87 cell growth by activating NF-κB-p65. Moreover, we found that t-AUCB induces cell-cycle G0/G1 phase arrest by regulating Cyclin D1 mRNA and protein levels and CDC2 (Thr161) phosphorylation level. We propose to further test this promising reagent for its anti-glioma activity in clinical relevant orthotopic brain glioma models.

In vitro and in vivo evaluation of ribavirin and pleconaril antiviral activity against enterovirus 71 infection.

Enterovirus 71(EV71) causes recurring outbreaks of hand, foot and mouth disease and encephalitis leading to complications or death in young children. More effective antiviral drugs are needed to prevent or reduce EV71-related disease and complications. However, there are no standard models currently in use to evaluate activity against EV71 infection both in vitro and in vivo. In this study, the activity of ribavirin and pleconaril against EV71 infection was evaluated in two models. An in vitro EV71 infection model was developed in RD cells, and an in vivo EV71 infection model was applied. Ribavirin and pleconaril effectively increased the viability of infected cells. Pleconaril reduced the morbidity and mortality of one-day-old infected mice, but ribavirin did not protect the infected mice. In all, the results demonstrated that infected cells and infected mice can be used to evaluate antiviral activity of ribavirin and pleconaril against EV71 infection in vitro and in vivo.

The effect of lentivirus-mediated TH and GDNF genetic engineering mesenchymal stem cells on Parkinson's disease rat model.

This study was designed to assess the potential therapeutic efficacy of gene-modified mesenchymal stem cells (MSCs), MSCs-TH and MSCs-GDNF, in PD rats. Fifty-nine PD rat models were divided into five groups and then the gene-modified MSCs were transplanted into the striatum of rats according to the design. Apomorphine-induced rotational behavior in rats was observed weekly; rats which received both MSCs-TH and MSCs-GDNF showed the most significant improvement compared with those in other groups (P < 0.01). Three weeks later, immunohistochemistry analysis found TH-positive cells and GDNF-positive cells in striatal. Eight weeks later, PD rats were killed. HPLC and ELISA results showed DA and GDNF content in striatum of rats which received both MSCs-TH, and MSCs-GDNF was considerably higher compared with those of other groups (P < 0.01),respectively. In conclusion, our results suggest that combined transplantation of MSCs expressing TH and GDNF can lead to remarkable therapeutic effects in a rat model of PD.

MicroRNA-181a sensitizes human malignant glioma U87MG cells to radiation by targeting Bcl-2.

Radiotherapy is widely used in cancer treatment and biological studies. Multiple mechanisms induced by radiation, especially changes of the expression profile of genes, lead to the disruption of cellular homeostasis. MicroRNAs (miRNAs) are important post-transcriptional gene regulators and play an important role in response to cellular stress. Here we investigated the profiles of miRNA expression following exposure to radiation and the possible role of miRNAs in the modulation of radiosensitivity in the glioblastoma multiform U87MG cell line. MiRNA expression profiles revealed a limited set of miRNAs with altered expression in U87MG cells in response to radiation treatment. MiR-181a, a member of miR-181 family, was one of the down-regulated miRNAs, whose expression was further validated by qRT-PCR. The target mRNAs of radiation-responsive miRNAs were predicted with a target prediction tool. Transiently overexpressed miR-181a significantly sensitized malignant glioma cells to radiation treatment concurrent with the down-regulation of the protein Bcl-2 (B cell lymphoma/lewkmia-2). It indicates that miR-181a may modulate radiosensitivity by targeting Bcl-2 in human malignant glioma cells. These data suggest that radiation can affect miRNA expression, which regulates the cellular response, and miR-181a could be a target for enhancing the effect of radiation treatment on malignant glioma cells.

Pharmacokinetics and the bystander effect in CD::UPRT/5-FC bi-gene therapy of glioma.

BACKGROUND: Cytosine deaminase (CD) converts 5-fluorocytosine (5-FC) to 5-fluorouracil (5-FU) in CD/5-FC gene therapy, 5-FU will be mostly converted into nontoxic beta-alanine without uracil phosphoribosyltransferase (UPRT). UPRT catalyzes the conversion of 5-FU to 5-fluorouridine monophosphate, which directly kills CD::UPRT-expressing cells and surrounding cells via the bystander effect. But the pharmacokinetics and the bystander effect of CD::UPRT/5-FC has not been verified in vivo and in vitro. Before the CD::UPRT/5-FC bi-gene therapy system is used in clinical trial, it is essential to monitor the transgene expression and function in vivo. Thus, we developed a preclinical tumor model to investigate the feasibility of using (19)F-magnetic resonance spectroscopy ((19)F-MRS) and optical imaging to measure non-invasive CD and UPRT expression and its bystander effect. METHODS: C6 and C6-CD::UPRT cells were cultured with 5-FC. The medium, cells and their mixture were analyzed by (19)F-MRS. Rats with intracranial xenografted encephalic C6-CD::UPRT glioma were injected intraperitoneally with 5-FC and their (19)F-MRS spectra recorded. Then the pharmacokinetics of 5-FC was proved. Mixtures of C6 and C6-CD::UPRT cells at different ratios were cultured with 5-FC and the cytotoxic efficacy and survival rate of cells recorded. To determine the mechanism of the bystander effect, the culture media from cell comprising 25% and 75% C6-CD::UPRT cells were examined by (19)F-MRS. A comparative study of mean was performed using analysis of variance (ANOVA). RESULTS: (19)F-MRS on samples from C6-CD::UPRT cells cultured with 5-FC showed three broad resonance signals corresponding to 5-FC, 5-FU and fluorinated nucleotides (F-Nuctd). For the C6 mixture, only the 5-FC peak was detected. In vivo serial (19)F-MRS spectra showed a strong 5-FC peak and a weak 5-FU peak at 20 minutes after 5-FC injection. The 5-FU concentration reached a maximum at about 50 minutes. The F-Nuctd signal appeared after about 1 hour, reached a maximum at around 160 minutes, and was detectable for several hours. At a 10% ratio of C6-CD::UPRT cells, the survival rate was (79.55 +/- 0.88)% (P < 0.01). As the C6-CD::UPRT ratio increased, the survival rate of the cells decreased. (19)F-MRS showed that the signals for 5-FU and F-Nuctd in the culture medium increased as the ratio of C6-CD::UPRT in the mixture increased. CONCLUSIONS: (19)F-MRS studies indicated that C6-CD::UPRT cells could effectively express CD and UPRT enzymes. The CD::UPRT/5-FC system showed an obvious bystander effect. This study demonstrated that CD::UPRT/5-FC gene therapy is suitable for 5-FC to F-Nuctd metabolism; and (19)F-MRS can monitor transferred CD::UPRT gene expression and catalysis of substrates noninvasively, dynamically and quantitatively.

Radiofrequency thermocoagulation-assisted surgery for intracranial giant vasogenic tumors.

BACKGROUND: We report the novel use of radiofrequency thermocoagulation to facilitate surgical excision of intracranial giant vasogenic tumors and detail the operative procedures and patient outcomes. MATERIALS AND METHODS: There were 2 patients with intracranial giant vasogenic tumors. The first was an extracerebral deep-seated cavernous angioma in the cavernous sinus (largest diameter: 8 cm), and the other was a hemangiopericytoma accreting the left confluence sinus (largest diameter: 9.2 cm). The tumors were well exposed during surgery and separated from the surrounding brain tissue by blunt dissection. The external surface of each tumor was devascularized. Radiofrequency thermocoagulation was applied in multiple cycles with each cycle encompassing a 3-cm-diameter volume to coagulate the inner tissue of the tumors prior to resection. The tumors were then resected in a piecemeal fashion starting from the thermocoagulated regions until complete removal was achieved. RESULTS: With radiofrequency thermocoagulation assistance, the 2 intracranial giant vasogenic tumors were removed completely with no bleeding. The surrounding brain tissue, cranial nerves, and vessels were kept intact. Patient recovery was uneventful. No complications and no tumor recurrences have occurred over a 2-year follow-up period. CONCLUSIONS: Radiofrequency thermocoagulation is extremely effective in controlling bleeding during surgical excision of intracranial giant vasogenic tumors. This improves the ease and safety of such procedures and allows for complete removal of tumors.

[Endoscopic transseptal transsphenoidal pituitary adenoma microsurgery].

OBJECTIVE: To introduce the approach and technique of endoscopic transseptal transsphenoidal pituitary adenoma microsurgery. METHOD: Endoscopic transseptal transsphenoidal pituitary adenoma microsurgery were performed in 23 patients. RESULT: Tumors were total resected in all of the patients. The mean volume of bleeding was 50 ml and there were not any severe complications. CONCLUSION: Compared with transsphenoidal pituitary adenoma microsurgery and endoscopic transsphenoidal pituitary adenoma surgery, endoscopic transseptal transsphenoidal pituitary adenoma microsurgery could combine the benefits of the two approaches. The approach has the benefit of microtrauma, convenience and could resect the tumor thoroughly. We could do best in treating suitable patients with good endoscopic technique.