Knockdown of STIM1 improves neuronal survival after traumatic neuronal injury through regulating mGluR1-dependent Ca(2+) signaling in mouse cortical neurons.

Activation of glutamate receptors and followed increase of intracellular calcium concentration is a key pathological mechanism involved in secondary neuronal injury after traumatic brain injury (TBI). Stromal interaction molecule (STIM) proteins are considered to be important players in regulating neuronal Ca(2+) homeostasis under normal aging and pathological conditions. Here, we investigated the role of STIM1 in regulating metabotropic glutamate receptor 1 (mGluR1)-related Ca(2+) signaling and neuronal survival by using an in vitro traumatic neuronal injury (TNI) model. The expression of STIM1 was significantly increased at both mRNA and protein levels after TNI. Down-regulation of STIM1 by specific small interfere RNA significantly preserved neuronal viability, decreased lactate dehydrogenase release, and inhibited apoptotic cell death after traumatic injury. Moreover, knockdown of STIM1 significantly alleviated the mGluR1-related increase of cytoplasmic Ca(2+) levels after TNI. By analyzing Ca(2+) imaging in Ca(2+)-free conditions, we demonstrated that the mGluR1-dependent inositol trisphosphate receptor and/or ryanodine receptor-mediated Ca(2+) release from the endoplasmic reticulum after TNI is strongly attenuated in the absence of STIM1. Together, our results demonstrate that in the mammalian nervous system, STIM1 is a key regulator of mGluR1-dependent Ca(2+) signaling and knockdown of STIM1 might be an effective intervention target in TBI.

Modified surgical method of supra- and infratentorial epidural hematoma and the related anatomical study of the squamous part of the occipital bone.

BACKGROUND: Supra- and infratentorial acute epidural hematoma (SIEDH) is a common posterior cranial fossa epidural hematoma located at the inner surface of the squamous part of the occipital bone (SOB). Traditionally, surgical treatment of the SIEDH requires a combined supra-infratentorial craniotomy. AIM: To analyze the morphological characteristics of the SOB and introduce a single supratentorial craniotomy for SIEDH. METHODS: Skull computed tomography (CT) scan data from 32 adult patients were collected from January 1, 2019 to January 31, 2020. On the median sagittal plane of the CT scan, the angle of the SOB (ASOB) was defined by two lines: Line A was defined from the lambdoid suture (LambS) to the external occipital protuberance (EOP), while line B was defined from the EOP to the posterior edge of the foramen magnum (poFM). The operative angle for the SIEDH (OAS) from the supra- to infratentorial epidural space was determined by two lines: The first line passes from the midpoint between the EOP and the LambS to the poFM, while the second line passes from the EOP to the poFM. The ASOB and OAS were measured and analyzed. RESULTS: Based on the anatomical study, a single supratentorial craniotomy was performed in 8 patients with SIEDH. The procedure and the results of the modified surgical method were demonstrated in detail. For males, the ASOB was 118.4 ± 4.7 and the OAS was 15.1 ± 1.8; for females, the ASOB was 130.4 ± 5.1 and the OAS was 12.8 ± 2.0. There were significant differences between males and females both in ASOB and OAS. The smaller the ASOB was, the larger the OAS was. The bone flaps in 8 patients were designed above the transverse sinus intraoperatively, and the SIEDH was completely removed without suboccipital craniotomy. The SOB does not present as a single straight plane but bends at an angle around the EOP and the superior nuchal lines. The OAS was negatively correlated with the ASOB. CONCLUSION: The single supratentorial craniotomy for SIEDH is reliable and effective.

Downregulation of Astrocyte Elevated Gene-1 Expression Combined with All-Trans Retinoic Acid Inhibits Development of Vasculogenic Mimicry and Angiogenesis in Glioma.

OBJECTIVE: This study aimed to investigate the effects of downregulating astrocyte elevated gene-1 (AEG-1) expression combined with all-trans retinoic acid (ATRA) on vasculogenic mimicry (VM) formation and angiogenesis in glioma. METHODS: U87 glioma cells were transfected with AEG-1 shRNA lentiviral vectors (U87-siAEG-1) and incubated in a medium containing 20 µmol/L ATRA. Matrigel-based tube formation assay was performed to evaluate VM formation, and the cell counting kit-8 (CCK-8) assay was used to analyze the proliferation of glioma cells in vitro. Reverse transcription-quantitative polymerase chain reaction and Western blot analysis were used to investigate the mRNA and protein expression of related genes, respectively. Glioma xenograft models were generated via subcutaneous implantation of glioma cells in nude mice. Tumor-bearing mice received an intraperitoneal injection of ATRA (10 mg/kg per day). Immunohistochemistry was used to evaluate the expression of related genes and the microvessel density (MVD) in glioma xenograft models. CD34/periodic acid-Schiff double staining was performed to detect VM channels in vivo. The volume and weight of tumors were measured, and a tumor growth curve was drawn to evaluate tumor growth. RESULTS: A combination of ATRA intervention and downregulation of AEG-1 expression significantly inhibited the proliferation of glioma cells in vitro and glioma VM formation in vitro and in vivo. It also significantly decreased MVD and inhibited tumor growth. Further, the expression levels of matrix metalloproteinase (MMP)-2, MMP-9, vascular endothelial-cadherin (VE-cadherin), and vascular endothelial growth factor (VEGF) in glioma significantly decreased in vivo and in vivo. CONCLUSION: Hence, a combinatorial approach might be effective in treating glioma through regulating MMP-2, MMP-9, VEGF, and VE-cadherin expression.

Three-dimensional time-of-flight magnetic resonance angiography combined with high resolution T2-weighted imaging in preoperative evaluation of microvascular decompression.

BACKGROUND: Neurovascular compression (NVC) is the main cause of primary trigeminal neuralgia (TN) and hemifacial spasm (HFS). Microvascular decompression (MVD) is an effective surgical method for the treatment of TN and HFS caused by NVC. The judgement of NVC is a critical step in the preoperative evaluation of MVD, which is related to the effect of MVD treatment. Magnetic resonance imaging (MRI) technology has been used to detect NVC prior to MVD for several years. Among many MRI sequences, three-dimensional time-of-flight magnetic resonance angiography (3D TOF MRA) is the most widely used. However, 3D TOF MRA has some shortcomings in detecting NVC. Therefore, 3D TOF MRA combined with high resolution T2-weighted imaging (HR T2WI) is considered to be a more effective method to detect NVC. AIM: To determine the value of 3D TOF MRA combined with HR T2WI in the judgment of NVC, and thus to assess its value in the preoperative evaluation of MVD. METHODS: Related studies published from inception to September 2022 based on PubMed, Embase, Web of Science, and the Cochrane Library were retrieved. Studies that investigated 3D TOF MRA combined with HR T2WI to judge NVC in patients with TN or HFS were included according to the inclusion criteria. Studies without complete data or not relevant to the research topics were excluded. The Quality Assessment of Diagnostic Accuracy Studies checklist was used to assess the quality of included studies. The publication bias of the included literature was examined by Deeks' test. An exact binomial rendition of the bivariate mixed-effects regression model was used to synthesize data. Data analysis was performed using the MIDAS module of statistical software Stata 16.0. Two independent investigators extracted patient and study characteristics, and discrepancies were resolved by consensus. Individual and pooled sensitivities and specificities were calculated. The I² statistic and Q test were used to test heterogeneity. The study was registered on the website of PROSERO (registration No. CRD42022357158). RESULTS: Our search identified 595 articles, of which 12 (including 855 patients) fulfilled the inclusion criteria. Bivariate analysis showed that the pooled sensitivity and specificity of 3D TOF MRA combined with HR T2WI for detecting NVC were 0.96 [95% confidence interval (CI): 0.92-0.98] and 0.92 (95%CI: 0.74-0.98), respectively. The pooled positive likelihood ratio was 12.4 (95%CI: 3.2-47.8), pooled negative likelihood ratio was 0.04 (95%CI: 0.02-0.09), and pooled diagnostic odds ratio was 283 (95%CI: 50-1620). The area under the receiver operating characteristic curve was 0.98 (95%CI: 0.97-0.99). The studies showed no substantial heterogeneity (I2 = 0, Q = 0.001 P = 0.50). CONCLUSION: Our results suggest that 3D TOF MRA combined with HR T2WI has excellent sensitivity and specificity for judging NVC in patients with TN or HFS. This method can be used as an effective tool for preoperative evaluation of MVD.

Anti-tumor effect of lentivirus-mediated gene transfer of alphastatin on human glioma.

Alphastatin, an endogenous angiogenesis inhibitor, has recently been used as an anticancer agent in several tumor models. This study was to investigate whether local sustained long-term expression of alphastatin could serve to diminish tumor growth of a human xenograft glioma model. We found that the recombinant alphastatin lentiviruses were able to stably infect HUVECs, and infected HUVECs could sustainably secrete alphastatin, which exhibited potent inhibitory effects on HUVECs migration, differentiation but not proliferation induced by vascular endothelial growth factor (VEGF) or basic fibroblast growth factor(bFGF). And the expression of secreted protein alphastatin markedly decreased tumor vascularization and inhibited tumor growth. Additionally, alphastatin inhibited VEGF- or bFGF-induced initial stage of angiogenesis by reducing JNk and ERK phosphorylation in vitro. Taken together, these data demonstrate that secreted protein alphastatin inhibits VEGF- or bFGF-induced angiogenesis by suppressing JNK and ERK kinases activation pathways in HUVECs, and markedly inhibits tumor angiogenesis in vivo. Consequently lentivirus-mediated gene transfer might represent an effective strategy for expression of alphastatin to achieve inhibition of human malignant glioma proliferation and tumor progression.

Homer1/mGluR1-mediated ER stress contributes to lysophosphatidic acid-induced neurotoxicity in cortical neurons.

Lysophosphatidic acid (LPA) is a glycerophospholipid that can be detected in serum, saliva and cerebrospinal fluid. However, the effect of LPA on neuronal death and survival has not been fully determined. In the present study, we investigated the potential neurotoxic effect of LPA in primary cultured cortical neurons. Treatment with LPA (0.5, 1 and 5 µM) markedly decreased neuronal viability, increased lactate dehydrogenase (LDH) release and promoted apoptosis in cortical neurons. The results of western blot showed that LPA increased the expression of endoplasmic reticulum (ER) stress associated factors, and the protein misfolding inhibitor 4-phenylbutyric acid (4-PBA) attenuated LPA-induced toxicity. In addition, treatment with LPA did not alter the expression and distribution of Homer1 in cortical neurons. The protein levels of metabotropic glutamate receptor 1 (mGluR1), but not metabotropic glutamate receptor 5 (mGluR5), were significantly increased by LPA at 12 and 24 h after treatment. Knockdown of Homer1 using specific siRNA partially prevented the LPA-induced neurotoxicity and ER stress. Furthermore, the results of Ca2+ imaging showed that treatment with LPA induced intracellular Ca2+ release, which could be partially prevented by 4-PBA and downregulation of Homer1. The LPA-induced intracellular Ca2+ release was associated with ER Ca2+ release through the Homer1-mGluR1 pathway. In summary, our results showed that LPA treatment induced ER stress and apoptosis in cortical neurons, and its neurotoxicity was partially mediated by Ca2+ release from the ER via the Homer1/mGluR1 pathway.

Ten-eleven translocation 1 regulates methylation of autophagy-related genes in human glioma.

Ten-eleven translocation 1 catalyzes the conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), which plays an important role in epigenetics and is related to the malignant biological behavior of tumors. However, its regulatory role in glioma remains unclear. In this study, the levels of 5mC and 5hmC were detected using immunohistochemistry, dot-blot, hMeDIP-chip, and western blot in glioma tissues and normal brain tissues, whereas 5hmC differentially enriched genes were determined and further validated. The level of 5hmC in gliomas was decreased, whereas 5mC was increased. 5hmC highly enriched 10 functional protein-coding genes and 10 signaling pathways were identified using hMeGIP-chip in glioma tissues. Two autophagy-related genes, ATG13 and DNA damage-regulated autophagy modulator protein 1, with low enrichment of 5hmC in glioma tissues were verified in the promoter region, and hMeGIP-PCR further confirmed this result in U251 cells. Immunohistochemistry further confirmed that autophagy level in glioma tissues was lower than that of normal controls, and negatively correlated with WHO grade. This study indicates that ten-eleven translocation 1 may be involved in the development and progression of glioma through demethylation regulating a variety of cellular functions and signaling pathways, and autophagy is one of the regulatory mechanisms.

Knockdown of RTN1-C attenuates traumatic neuronal injury through regulating intracellular Ca2+ homeostasis.

Reticulons (RTNs) are a family of membrane-bound proteins that are dominantly localized to the endoplasmic reticulum (ER) membrane. RTN1-C is one member of RTNs abundantly expressed in the brain and has been shown to mediate neuronal injury in cerebral ischemia models. In the present study, we investigated the role of RTN1-C in an in vitro brain trauma model mimicked by traumatic neuronal injury (TNI) in primary cultured cortical neurons. TNI increased the expression of RTN1-C in cortical neurons but had no effect on RTN1-A and RTN1-B. Knockdown of RTN1-C with specific siRNA (Si-RTN1-C) significantly decreased cytotoxicity and apoptosis after TNI. The results of Ca2+ imaging showed that intracellular Ca2+ overload induced by TNI was attenuated by RTN1-C knockdown. Furthermore, the activation of metabotropic glutamate receptor 1 (mGluR1)-induced Ca2+ response was partially prevented by Si-RTN1-C transfection. We also evaluated the role of RTN1-C in store-operated Ca2+ entry (SOCE) in cortical neurons using the ER Ca2+ inducer thapsigargin (Tg). The results showed that knockdown of RTN1-C alleviated the SOCE-mediated Ca2+ influx and decreased the expression of stromal interactive molecule 1 (STIM1). In summary, the present study found that knockdown of RTN1-C protected neurons against TNI via preservation of intracellular Ca2+ homeostasis, which was associated with the inhibition of mGluR1-mediated ER Ca2+ release and suppression of STIM1-related SOCE. Thus, RTN1-C might represent a therapeutic target for traumatic brain injury (TBI) research.

miR-139 Functions as An Antioncomir to Repress Glioma Progression Through Targeting IGF-1 R, AMY-1, and PGC-1ß.

Gliomas are the most common primary malignant brain tumor with poor prognosis, characterized by a highly heterogeneous cell population, extensive proliferation, and migration. A lot of molecular mechanisms regulate gliomas development and invasion, including abnormal expression of oncogenes and variation of epigenetic modification. MicroRNAs could affect cell growth and functions. Several reports have demonstrated that miR-139 plays multifunctions in kinds of solid tumors through different pathways. However, the antitumor mechanisms of this miR-139 are not unveiled in detail. In this study, we not only validated the low expression level of miR-139 in glioma tissues and cell lines but also detected the effect of miR-139 on modulating gliomas proliferation and invasion both in vitro and in vivo. We identified insulin-like growth factor 1 receptor, associate of Myc 1, and peroxisome proliferator-activated receptor γ coactivator 1ß as direct targets of miR-139 and the levels of them were all inversely correlated with miR-139 in gliomas. Insulin like growth factor 1 receptor promoted gliomas invasion through Akt signaling and increased proliferation in the peroxisome proliferator-activated receptor γ coactivator 1ß-dependent way. Associate of Myc 1 also facilitated gliomas progression by activating c-Myc pathway. Overexpression of the target genes could retrieve the antitumor function of miR-139, respectively, in different degrees. The nude mice transplantation tumor experiment displayed that glioma cells stably expressed miR-139 growth much slower in vivo than the negative control cells. Taken together, these findings suggested miR-139 acted as a favorable factor against gliomas progression and uncovered a novel regulatory mechanism, which may provide a new evidenced prognostic marker and therapeutic target for gliomas.

Associations between EGFR gene polymorphisms and susceptibility to glioma: a systematic review and meta-analysis from GWAS and case-control studies.

The results of genome-wide association studies (GWAS) and case-control studies performed to investigate the associations between epidermal growth factor receptor (EGFR) gene polymorphisms and glioma risk are controversial. The aim of this systematic review and meta-analysis is to determine whether EGFR gene polymorphisms are associated with glioma risk by searching 'PubMed', 'EMBASE', 'Web of Science', 'Cochrane Library' and 'China WeiPu Library' to retrieve studies that investigated associations between EGFR gene polymorphisms and glioma risk. Four GWAS containing 35 studies and 7 case-control studies meeting the inclusion criteria were finally recruited, and 11 single-nucleotide polymorphisms (SNPs) were analyzed. The results showed a significant positive association between rs730437/rs845552 and glioma risk in Asians, and a significant negative association between them in Caucasians. In addition, rs11506105 was significantly associated with an increased risk of glioma in both Asians and Caucasians, and rs11979158 decreased the risk of glioma in Caucasians. However, no significant association was observed between rs12718945/rs17172432/rs4947492 and glioma risk in Asians, between rs2252586 and glioma risk in Caucasians, and between rs3752651 and glioma risk in either Asians or Caucasians. In conclusion, different SNPs in EGFR gene might have different impacts on the risk of glioma in various ethnicities, which offers new insights into the treatment with a target-oriented approach.

TET1 exerts its tumour suppressor function by regulating autophagy in glioma cells.

DNA methylation and demethylation play a critical role in the regulation of the molecular pathogenesis of gliomas. Tet methylcytosine dioxygenase 1 (TET1) catalyses the sequential oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine, (5hmC) leading to eventual DNA demethylation. It has been reported that TET1 is a tumour suppressor in several cancers. However, whether TET1 plays a role in glioma development is largely unclear. Different glioma specimens and corresponding normal controls were collected to analyse the expression of TET1. At the same time, TET1 of glioma U251 cells was knocked down or overexpressed to observe its effect on glioma cell proliferation and invasion as well as autophagy level. Here, we reported that the expression of TET1 in glioma tissue was significantly lower than the corresponding non-tumour normal tissues, and the concentration of TET1 is negatively correlated with the glioma WHO classification. When TET1 gene in glioma U251 cells was knocked down by CRISPR/Caspase-9 system, the proliferation and invasive ability of U251 increased remarkably. But when TET1 was overexpressed in U251 cells, the proliferation and invasion were impaired. Following the down-expression of TET1, the level of autophagy in U251 cells decreased accordingly.However, when TET1 was overexpressed in U251 cells, the level of autophagy incraesed. Furthermore, bafilomycin A1 (Baf-A1) but not 3-methyladenine (3-MA) could decrease the autophagy level of TET1-/- U251 cells as the wild-type controls. It suggests that the tumour suppressor effect of TET1 seems to be mediated by regulating the level of autophagy, and the regulation of TET1 on autophagy is at an early stage.