Shuxu Yang

miR-92b controls glioma proliferation and invasion through regulating Wnt/beta-catenin signaling via Nemo-like kinase

BACKGROUND Nemo-like kinase (NLK) is an evolutionarily conserved protein kinase involved in Wnt/beta-catenin signaling, which has been reported to be associated with gliomagenesis. In the present study, we aimed to identify a concrete mechanism of Wnt/beta-catenin pathway regulation by microRNAs (miRNAs) in glioma. METHODS Quantitative reverse-transcription polymerase chain reaction and in situ hybridization were conducted to detect the expression of miR-92b. The cell proliferation rate and cell cycle kinetics were detected using 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assay and flow cytometry, cell invasion and migration were evaluated using Transwell assay and wound healing assay, and cell apoptosis was detected using annexin V staining. Furthermore, the relevant molecules regulating proliferation and invasion were examined using Western blot analysis, immunohistochemistry, and immunofluorescence staining. Luciferase reporter assay was used to identify the direct regulation of NLK by miR-92b and beta-catenin/TCF4 activity. RESULTS We first showed that the expression of miR-92b was elevated in both glioma samples and glioma cells. Furthermore, down-regulation of miR-92b triggered growth inhibition, induced apoptosis, and suppressed invasion of glioma in vitro and in vivo. Luciferase assay and Western blot analysis revealed that NLK is a direct target of miR-92b. Restoring expression of NLK inhibited glioma proliferation and invasion. Mechanistic investigation revealed that miR-92b deletion suppressed beta-catenin/TCF-4 transcription activity by targeting NLK. Moreover, expression of NLK was inversely correlated with miR-92b in glioma samples and was predictive of patient survival in a retrospective analysis. CONCLUSIONS Our findings identify a role for miR-92b in glioma proliferation and invasion after activation of Wnt/beta-catenin signaling via NLK.

LRIG1 dictates the chemo-sensitivity of temozolomide (TMZ) in U251 glioblastoma cells via down-regulation of EGFR/topoisomerase-2/Bcl-2

In the current study, we aimed to understand the potential role of leucine-rich repeats and immunoglobulin-like domains 1 (LRIG1) in TMZ-resistance of U251 glioma cells. We established TMZ-resistant U251 clones (U251/TMZ cells), which expressed low level of LRIG1, but high levels of epidermal growth factor receptor (EGFR), topoisomerase-2 (Topo-2) and Bcl-2. Depletion of LRIG1 by the targeted RNA interference (RNAi) upregulated EGFR/Topo-2/Bcl-2 in U251 cells, and the cells were resistant to TMZ. Reversely, over-expression of LRIG1 in U251 cells downregulated EGFR/Topo-2/Bcl-2 expressions, and cells were hyper-sensitive to TMZ. Our data suggested EGFR-dependent mammalian target of rapamycin (mTOR) activation was important for Topo-2 and Bcl-2 expressions in U251/TMZ cells. The EGFR inhibitor and the mTOR inhibitor downregulated Topo-2/Bcl-2 expressions, both inhibitors also restored TMZ sensitivity in U251/TMZ cells. Finally, inhibition of Topo-2 or Bcl-2 by targeted RNAi(s) knockdown or by the corresponding inhibitor re-sensitized U251/TMZ cells to TMZ, indicating that both Topo-2 and Bcl-2 were important for TMZ resistance in the resistant U251 cells. Based on these results, we concluded that LRIG1 inhibits EGFR expression and the downstream signaling activation, interferes with Bcl-2/Topo-2 expressions and eventually sensitizes glioma cells to TMZ.

DNMT1 mediates chemosensitivity by reducing methylation of miRNA-20a promoter in glioma cells

Although methyltransferase has been recognized as a major element that governs the epigenetic regulation of the genome during temozolomide (TMZ) chemotherapy in glioblastoma multiforme (GBM) patients, its regulatory effect on glioblastoma chemoresistance has not been well defined. This study investigated whether DNA methyltransferase (DNMT) expression was associated with TMZ sensitivity in glioma cells and elucidated the underlying mechanism. DNMT expression was analyzed by western blotting. miR-20a promoter methylation was evaluated by methylation-specific PCR. Cell viability and apoptosis were assessed using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and TdT-mediated dUTP-biotin nick end labeling assays, respectively. The results showed that compared with parental U251 cells, DNMT1 expression was downregulated, miR-20a promoter methylation was attenuated and miR-20a levels were elevated in TMZ-resistant U251 cells. Methyltransferase inhibition by 5-aza-2′-deoxycytidine treatment reduced TMZ sensitivity in U251 cells. In U251/TM cells, DNMT1 expression was negatively correlated with miR-20a expression and positively correlated with TMZ sensitivity and leucine-rich repeats and immunoglobulin-like domains 1 expression; these effects were reversed by changes in miR-20a expression. DNMT1 overexpression induced an increase in U251/TM cell apoptosis that was inhibited by the miR-20a mimic, whereas DNMT1 silencing attenuated U251/TM cell apoptosis in a manner that was abrogated by miR-20a inhibitor treatment. Tumor growth of the U251/TM xenograft was inhibited by pcDNA-DNMT1 pretreatment and boosted by DNMT1-small hairpin RNA pretreatment. In summary, DNMT1 mediated chemosensitivity by reducing methylation of the microRNA-20a promoter in glioma cells.

MiR-27a regulates Wnt/beta-catenin signaling through targeting SFRP1 in glioma.

Glioma is one of the most common intracranial tumors, and the prognosis is poor, although more and more treatments are employed. Wnt/beta-catenin signaling has been reported to be associated with glioma. SFRP1 acts as an antagonist and inhibits Wnt signaling by binding to Wnt molecules. In the present study, we aimed to investigate miRNA-27a as an antineoplastic factor that inhibits the Wnt/beta-catenin pathway by binding to the SFRP1 3′-UTR in glioma in vitro. We first showed that the expression of miR-27a was elevated in both glioma samples and cell lines. Furthermore, downregulation of miR-27a induced growth inhibition, cycle arrest, and apoptosis, and suppressed invasion/migration in glioma cell lines. Quantitative real-time PCR, western blot, and luciferase assay analysis showed that SFRP1 is a direct target of miR-27a. Overexpression of SFRP1 inhibited the malignancy of glioma cell lines. Our investigation showed that downregulation of miR-27a suppressed beta-catenin/TCF-4 transcription activity by targeting SFRP1. Our findings identify a role for miR-27a in glioma cell viability, cycle, apoptosis, and invasion/migration after activation of Wnt/beta-catenin signaling through SFRP1.

The autophagy–lysosomal system in subarachnoid haemorrhage

The autophagy–lysosomal pathway is a self‐catabolic process by which dysfunctional or unnecessary intracellular components are degraded by lysosomal enzymes. Proper function of this pathway is critical for maintaining cell homeostasis and survival. Subarachnoid haemorrhage (SAH) is one of the most devastating forms of stroke. Multiple pathogenic mechanisms, such as inflammation, apoptosis, and oxidative stress, are all responsible for brain injury and poor outcome after SAH. Most recently, accumulating evidence has demonstrated that the autophagy–lysosomal pathway plays a crucial role in the pathophysiological process after SAH. Appropriate activity of autophagy–lysosomal pathway acts as a pro‐survival mechanism in SAH, while excessive self‐digestion results in cell death after SAH. Consequently, in this review article, we will give an overview of the pathophysiological roles of autophagy–lysosomal pathway in the pathogenesis of SAH. And approaching the molecular mechanisms underlying this pathway in SAH pathology is anticipated, which may ultimately allow development of effective therapeutic strategies for SAH patients through regulating the autophagy–lysosomal machinery.

Delayed Intracerebral Hemorrhage Secondary to Ventriculoperitoneal Shunt: A Case Report and Literature Review.

AbstractThe ventriculoperitoneal (VP) shunt is a routine procedure for cerebrospinal fluid (CSF) diversion, and is associated with many complications. A delayed hemorrhage after the VP shunt surgery, however, is quite rare. In this study, we report a case involving late-onset hemorrhage. The 67-year-old male patient with a history of head trauma and brain surgery underwent a VP shunt placement for hydrocephalus. The surgery course was uneventful and no bleeding was revealed in the first computed tomographic (CT) scan after the procedure. However, a massive intraparenchymal and intraventricular hemorrhage occurred 8 h following adjustment of the valve system on the 8th day after surgery.Erosion of the vasculature by catheter cannulation and a sudden reduction of CSF pressure after downregulation of the valve could be one of the possible causes of the intracerebral hemorrhage (ICH).

Neuroendocrine dysfunction and insomniain mild traumatic brain injury patients

Mild traumatic brain injury (mTBI) has been a growing public health concern in the worldwide. To investigate the subjective and objective characteristics of insomnia following mTBI and the association between insomnia and hypothalamic-pituitary-adrenal (HPA) function of mTBI patients, 59 patients with mTBI (mTBI group) were compared with 50 healthy participants (control group) in the present study. The subjective and objective measures of insomnia were respectively obtained from Pittsburgh Sleep Quality (PSQI) and polysomnography (PSG). HPA function was measured with low-dose short synacthen test (LDSST). According to the comparative and correlation analysis of the two groups, for PSQI, the scores of sleep syndrome, sleep latency, sleep efficiency, overall sleep quality and daytime dysfunction of mTBI patients were all higher, however only sleep efficiency and daytime dysfunction of mTBI patients were related with peak cortisol on lDSST; while for PSG, sleep efficiency (SE) was lower and wake after sleep onset (WASO) was longer in mTBI patients, moreover SE and WASO of mTBI patients were correlated with peak cortisol on LDSTT; for HPA function indexes, only peak cortisol on LDSST was lower in mTBI patients. These findings suggested that mTBI patients experienced more serious subjective insomnia symptoms than objective measurement, which were associated with HPA dysfunction. This study may contribute to identifying better treatment for mTBI patients with insomnia.

Norcantharidin blocks Wnt/β-catenin signaling via promoter demethylation of WIF-1 in glioma

Glioma is one of the most common primary intracranial tumors, and the prognosis is poor even though much treatment management is employed. Wnt/β-catenin signaling has been reported to be associated with glioma. Norcantharidin (NCTD) is the demethylated analog of cantharidin isolated from blister beetles, and it is reported to possess anticancer activity but less nephrotoxicity than cantharidin. Accordingly, we aimed to investigate NCTD as an anti-neoplastic drug that inhibits the Wnt/β‑catenin pathway via promoter demethylation of Wnt inhibitory factor-1 (WIF-1) in glioma growth in vitro. In the present study, we report that NCTD inhibited cell proliferation, induced apoptosis and cell cycle arrest, and suppressed cell migration and invasion in vitro. Moreover, we observed that the expression levels of WIF-1 mRNA and protein in the NCTD-treated cells were increased significantly compared with these levels in the negative control (NC) cells. Promoter demethylation was observed in the NCTD‑treated cells. In contrast, aberrant methylation was observed in the NC cells. Additionally, more investigation revealed that NCTD suppressed activity of Wnt/β-catenin signaling and transcription of β-catenin/TCF-4. Furthermore, the expression of apoptosis-related proteins Bcl-2 and cleaved caspase-3 indicated significant cell apoptosis. We provide initial evidence that NCTD reactivates WIF-1 from a methylation state, and downregulates canonical Wnt/β-catenin signaling. Our findings revealed that NCTD is effective for glioma in vitro and may be a new therapeutic option in vivo.

Glioblastoma mimicking a cerebral contusion: A case report

A 61-year-old male presented with a rare case of glioblastoma mimicking a cerebral contusion subsequent to collapsing. The patient had been medicated for hypertension for seven years and diabetes for eight years prior to hospitalization. Brain computed tomography (CT) revealed a cerebral contusion and intracerebral hemorrhage (ICH) in the left temporal region. The patient was initially administered intravenous drugs to reduce the intracranial pressure following the diagnosis of a cerebral contusion. Serial CT revealed ICH resorption. However, the patient was again admitted due to a headache and vomiting two months later. Magnetic resonance imaging (MRI) demonstrated an enhanced ring-shaped mass around the cyst cavity within the left temporal region, with surrounding edema. The patient underwent cyst puncture drainage in the temporal region. No tumor cells were identified in the cyst fluid and the culture was also negative. The patient was admitted for a headache and vomiting for the third time one month after being discharged. A cyst, tumor and meningoencephalitis were suspected following an MRI scan. The patient was treated with a left temporal craniotomy for a mass resection and biopsy. The histological diagnosis of the biopsy specimen was that of a glioblastoma. Two months later, MRI revealed a recurrence of the glioblastoma. In the present case, a brain tumor should have initially been suspected as the cause of the ICH, despite the history of craniocerebral trauma and hypertension. Early awareness of this potential cause of ICH may facilitate a more prompt diagnosis and treatment.

The comparison between dissociate bone flap cranioplasty and traditional cranioplasty in the treatment of depressed skull fractures.

ObjectiveDepressed skull fractures (DSF) with operation indications should be paid with enough attention because they have several complications and can influence esthetics. The optimal surgical method for DSF remains unclear. We explored the merits of dissociate bone flap cranioplasty. Patients and MethodsFrom July 2006 to August 2012, we performed 30 craniotomies on patients with DSF, which were divided into 2 groups: 1 group, which consisted of 18 patients, underwent dissociate bone cranioplasty; the other 12 patients underwent lever-up cranioplasty. A helical computed tomographic scan was routinely obtained after the operation and a 3-dimensional technique was performed on some patients to evaluate the postoperative condition of the flap. ResultsDissociate bone flap cranioplasty was performed on the 18 patients [11 men, 7 women: age, 26–70 (41) y]. No complications were observed in these patients. Lever-up cranioplasty was applied in the 12 patients [8 men, 4 women: age, 19–60 (41.8) y]; 2 patients had wound infection and 2 emerged with epidural hematoma. Obvious statistical significance of stability (P = 0.013) and position (P = 0.015) was found between the 2 methods. ConclusionsDissociate bone flap cranioplasty is safer, more flexible, has less complications, and has better plasticity. We advocate the use of bone flap cranioplasty in dealing with DSF.