A CRISPR-Cas12a-based electrochemical biosensor for the detection of microphthalmia-associated transcription factor.

A novel electrochemical biosensor that combines the CRISPR-Cas12a system with a gold electrode is reported for the rapid and sensitive detection of microphthalmia-associated transcription factor (MITF). The biosensor consists of a gold electrode modified with DNA1, which contains the target sequence of MITF and is labeled with ferrocene, an electroactive molecule. The biosensor also includes hairpin DNA, which has a binding site for MITF and can hybridize with helper DNA to form a double-stranded complex that activates CRISPR-Cas12a. When MITF is present, it binds to hairpin DNA and prevents its hybridization with helper DNA, thus inhibiting CRISPR-Cas12a activity and preserving the DPV signal of ferrocene. When MITF is absent, hairpin DNA hybridizes with helper DNA and activates CRISPR-Cas12a, which cleaves DNA1 and releases ferrocene, thus reducing the DPV signal. The biosensor can detect MITF with high sensitivity (with an LOD of 8.14 fM), specificity, and accuracy in various samples, such as cell nuclear extracts and human serum. The biosensor can also diagnose and monitor melanocyte-related diseases and melanin production. This work provides a simple, fast, sensitive, and cost-effective biosensor for MITF detection and a valuable tool for applications in genetic testing, disease diagnosis, and drug screening.

Targeting MYH9 represses USP14-mediated NAP1L1 deubiquitination and cell proliferation in glioma.

Myosin heavy chain 9 (MYH9) plays an important role in a number of diseases. Nevertheless, the function of MYH9 in glioma is unclear. The present research aimed to investigate the role of MYH9 in glioma and determine whether MYH9 is involved in the temozolomide chemoresistance of glioma cells. Our results showed that MYH9 increased the proliferation and temozolomide resistance of glioma cells. The mechanistic experiments showed that the binding of MYH9 to NAP1L1, a potential promoter of tumor proliferation, inhibited the ubiquitination and degradation of NAP1L1 by recruiting USP14. Upregulation of NAP1L1 increased its binding with c-Myc and activated c-Myc, which induced the expression of CCND1/CDK4, promoting glioma cell temozolomide resistance and proliferation. Additionally, we found that MYH9 upregulation was strongly related to patient survival and is therefore a negative factor for patients with glioma. Altogether, our results show that MYH9 plays a role in glioma progression by regulating NAP1L1 deubiquitination. Thus, targeting MYH9 is a potential therapeutic strategy for the clinical treatment of glioma in the future.

Role of long noncoding RNA MEG3/miR-378/GRB2 axis in neuronal autophagy and neurological functional impairment in ischemic stroke.

Autophagy has been shown to maintain neural system homeostasis during stroke. However, the molecular mechanisms underlying neuronal autophagy in ischemic stroke remain poorly understood. This study aims to investigate the regulatory mechanisms of the pathway consisting of MEG3 (maternally expressed gene 3), microRNA-378 (miR-378), and GRB2 (growth factor receptor-bound protein 2) in neuronal autophagy and neurological functional impairment in ischemic stroke. A mouse model of the middle cerebral artery occluded-induced ischemic stroke and an in vitro model of oxygen-glucose deprivation-induced neuronal injury were developed. To understand the role of the MEG3/miR-378/GRB2 axis in the neuronal regulation, the expression of proteins associated with autophagy in neurons was measured by Western blotting analysis, and neuron death was evaluated using a lactate dehydrogenase leakage rate test. First, it was found that the GRB2 gene, up-regulated in middle cerebral artery occluded-operated mice and oxygen-glucose deprivation-exposed neurons, was a target gene of miR-378. Next, miR-378 inhibited neuronal loss and neurological functional impairment in mice, as well as neuronal autophagy and neuronal death by silencing of GRB2. Confirmatory in vitro experiments showed that MEG3 could specifically bind to miR-378 and subsequently up-regulate the expression of GRB2, which in turn suppressed the activation of Akt/mTOR pathway. Taken together, these findings suggested that miR-378 might protect against neuronal autophagy and neurological functional impairment and proposed that a MEG3/miR-378/GRB2 regulatory axis contributed to better understanding of the pathophysiology of ischemic stroke.

CCT5 interacts with cyclin D1 promoting lung adenocarcinoma cell migration and invasion.

Cyclin D1 (CCND1) has been identified as a metastatic promoter in various tumors including lung adenocarcinoma (LUAD), a subtype of non small cell lung cancer (NSCLC). The previous observation revealed that CCND1 was upregulated in NSCLC and predicted poor prognosis of LUAD patients. In this study, we examined a chaperonin containing TCP1 subunit 5 (CCT5) protein interacts with CCND1 in LUAD. Immunofluorescence demonstrated the co-localization of CCT5 and CCND1 protein in LUAD cells. CCT5 expression was detected with both immunohistochemistry (IHC) and bioinformatics analyses. Similar with the expression pattern of CCND1, CCT5 displayed a high level in LUAD tissues compared to non cancerous lung specimens. Patients with high CCT5 expression showed a significant shorter overall survival relative to those with low expression level. Furthermore, upregulated CCT5 exhibited significant positive correlation with TNM stage of LUAD patients in both IHC analyses and bioinformatics. Knocking down CCT5 remarkably inhibited LUAD cell migration and invasion in vitro by inactivating PI3K/AKT and its downstream EMT signals, which could abrogated the accelerated migration and invasion caused by CCND1 overexpression. In summary, our study discovered a highly expressed protein CCT5 in LUAD which interacted with CCND1 and promoted migration and invasion of LUAD cells by positively moderating PI3K/AKT-induced EMT pathway.

miR-5188 augments glioma growth, migration and invasion through an SP1-modulated FOXO1-PI3K/AKT-c-JUN-positive feedback circuit.

The biological effect and molecular mechanism of miR-5188 have not been thoroughly investigated. The study aims at elucidating the role of miR-5188 in glioma progression. Human glioma cell lines and tissues were used for functional and expression analysis. Cellular and molecular techniques were performed to explore the functions and mechanisms of miR-5188 in glioma. In our investigation, we demonstrated that miR-5188 promoted cell proliferation, the G1/S transition of the cell cycle, migration and invasion in glioma and reduced the lifespan of glioma-bearing mice. miR-5188 directly targeted FOXO1 and activated PI3K/AKT-c-JUN signalling, which enhanced miR-5188 expression. Moreover, the c-JUN transcription factor functionally bound to the miR-5188 promoter region, forming the positive feedback loop. The feedback loop promoted glioma progression through activating the PI3K/AKT signalling, and this loop is augmented by the interaction between SP1 and c-JUN. Moreover, it was also found that the miR-5188/FOXO1 axis is facilitated by SP1-activated PI3K/AKT/c-JUN signalling. In glioma samples, miR-5188 expression was found to be an unfavourable factor and was positively associated with the mRNA levels of SP1 and c-JUN, whereas negatively associated with the mRNA levels of FOXO1. Our investigation demonstrates that miR-5188 could function as a tumour promoter by directly targeting FOXO1 and participating in SP1-mediated promotion of cell growth and tumorigenesis in glioma.

Phytanoyl-CoA 2-Hydroxylase-Interacting Protein-Like Gene Is a Therapeutic Target Gene for Glioblastoma Multiforme.

Glioblastoma multiforme (GBM) is the most common primary CNS cancer and has a poor prognosis. This study searched for significant genes and the mechanisms involved in GBM. We used the Gene Expression Omnibus (GEO) to test the WHO normal and IV glioma database, used R tool to identify the significant gene, and finally, combined these with The Cancer Genome Atlas (TCGA) to verify the significant genes. Subsequently, we explored the biological mechanisms involved. Phytanoyl-CoA 2-hydroxylase-interacting protein-like gene (PHYHIPL) is downregulated in grade IV glioma (GBM). The downregulation of PHYHIPL in GBM is accompanied by poor overall survival in the TCGA database, which indicates that PHYHIPL is a protection gene in GBM development. Bioinformatics analysis shows that the poor prognosis with downregulated PHYHIPL may be the result of the TNF signaling pathway and the IL-17 signaling pathway, but good prognosis accompanied by upregulated PHYHIPL may be the result of retrograde endocannabinoid signaling and the cAMP signaling pathway. Protein-protein interactions (PPI) net indicated that PHYHIPL may play a vital role in cell metabolism, and we hypothesize that the downregulation mechanism may be the result of mutations of the ß-catenin gene and the endogenous siRNA, as shown in previous studies. PHYHIPL may be a target gene for the treatment and prognosis of GBM.

The Tumorgenicity of Glioblastoma Cell Line U87MG Decreased During Serial In Vitro Passage.

Established cancer cell lines are routinely used to study cancer. Several factors such as serial passage may affect the reproducibility of experiments with cancer cell lines, but few researches focused on these changes. In the present study, different morphology and decreased tumorigenicity were observed in late passage U87MG cells. In vitro experiments further revealed that late passage U87MG cells possessed lower invasion properties than early passage, whereas no significant differences of proliferation and migration were found between early and late passage U87MG cells. In particular, we confirmed that late passage U87MG cells exhibited more epithelial phenotype with decreased PI3K/Akt pathway and TGF-ß pathway expressions at protein level. In summary, our results focused on the changes of U87MG cells during serial in vitro passage, suggested that passage-induced changes may lead to notable changes of biological characteristics and several molecular transitions in cancer cell lines, indicating the necessity to shorten experiment-span and accomplish experiments with the same or similar passage cancer cell strains.

Alpha-enolase as a potential cancer prognostic marker promotes cell growth, migration, and invasion in glioma.

BACKGROUND: The success of using glycolytic inhibitors for cancer treatment relies on better understanding the roles of each frequently deregulated glycolytic genes in cancer. This report analyzed the involvement of a key glycolytic enzyme, alpha-enolase (ENO1), in tumor progression and prognosis of human glioma. METHODS: ENO1 expression levels were examined in glioma tissues and normal brain (NB) tissues. The molecular mechanisms of ENO1 expression and its effects on cell growth, migration and invasion were also explored by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay, Transwell chamber assay, Boyden chamber assay, Western blot and in vivo tumorigenesis in nude mice. RESULTS: ENO1 mRNA and protein levels were upregulated in glioma tissues compared to NB. In addition, increased ENO1 was associated disease progression in glioma samples. Knocking down ENO1 expression not only significantly decreased cell proliferation, but also markedly inhibited cell migration and invasion as well as in vivo tumorigenesis. Mechanistic analyses revealed that Cyclin D1, Cyclin E1, pRb, and NF-κB were downregulated after stable ENO1 knockdown in glioma U251 and U87 cells. Conversely, knockdown of ENO1 resulted in restoration of E-cadherin expression and suppression of mesenchymal cell markers, such as Vimentin, Snail, N-Cadherin, ß-Catenin and Slug. Furthermore, ENO1 suppression inactivated PI3K/Akt pathway regulating the cell growth and epithelial-mesenchymal transition (EMT) progression. CONCLUSION: Overexpression of ENO1 is associated with glioma progression. Knockdown of ENO1 expression led to suppressed cell growth, migration and invasion progression by inactivating the PI3K/Akt pathway in glioma cells.

Electrochemical biosensing for E.coli detection based on triple helix DNA inhibition of CRISPR/Cas12a cleavage activity.

BACKGROUND: Escherichia coli (E.coli) is both a commensal and a foodborne pathogenic bacterium in the human gastrointestinal tract, posing significant potential risks to human health and food safety. However, one of the major challenges in E.coli detection lies in the preparation and storage of antibodies. In traditional detection methods, antibodies are indispensable, but their instability often leads to experimental complexity and increased false positives. This underscores the need for new technologies and novel sensors. Therefore, the development of a simple and sensitive method for analyzing E.coli would make significant contributions to human health and food safety. RESULTS: We constructed an electrochemical biosensor based on triple-helical DNA and entropy-driven amplification reaction (EDC) to inhibit the cleavage activity of Cas12a, enabling high-specificity detection of E.coli. Replacing antibodies with nucleic acid aptamers (Apt) as recognition elements, we utilized the triple-helical DNA generated by the binding of DNA2 and DNA5/DNA6 double-helical DNA through the entropy-driven amplification reaction to inhibit the collateral cleavage activity of clustered regularly interspaced short palindromic repeats gene editing system (CRISPR) and its associated proteins (Cas). By converting E.coli into electrical signals and recording signal changes in the form of square wave voltammetry (SWV), rapid detection of E.coli was achieved. Optimization of experimental conditions and data detection under the optimal conditions provided high sensitivity, low detection limits, and high specificity. SIGNIFICANCE: With a minimal detection limit of 5.02 CFU/mL and a linear range of 1 × 102 - 1 × 107 CFU/mL, the suggested approach was successfully verified to analyze E.coli at various concentrations. Additionally, after examining E.coli samples from pure water and pure milk, the recoveries ranged between 95.76 and 101.20%, demonstrating the method's applicability. Additionally, it provides a feasible research direction for the detection of pathogenic bacteria causing other diseases using the CRISPR/Cas gene editing system.

A novel electrochemical biosensor for B-type natriuretic peptide detection based on CRISPR/Cas13a and chain substitution reaction.

B-type natriuretic peptide (BNP) is a biomarker for heart failure, a serious and prevalent disease that requires rapid and accurate diagnosis. In this study, we developed a novel electrochemical biosensor for BNP detection based on CRISPR/Cas13a and chain substitution reaction. The biosensor consists of a DNA aptamer that specifically binds to BNP, a T7 RNA polymerase that amplifies the signal, a CRISPR/Cas13a system that cleaves the target RNA, and a two-dimensional DNA nanoprobe that generates an electrochemical signal. The biosensor exhibits high sensitivity, specificity, and stability, with a detection limit of 0.74 aM. The biosensor can also detect BNP in human serum samples with negligible interference, demonstrating its potential for clinical and point-of-care applications. This study presents a novel strategy for integrating CRISPR/Cas13a and chain substitution reaction into biosensor design, offering a versatile and effective platform for biomolecule detection.

Proximity hybridization induced molecular machine for signal-on electrochemical detection of α-synuclein oligomers.

α-synuclein oligomer is a marker of Parkinson's disease. The traditional enzyme-linked immunosorbent assay for α-synuclein oligomer detection is not conducive to large-scale application due to its time-consuming, high cost and poor stability. Recently, DNA-based biosensors have been increasingly used in the detection of disease markers due to their high sensitivity, simplicity and low cost. In this study, based on the DNAzyme-driven DNA bipedal walking method, we developed a signal-on electrochemical sensor for the detection of α-syn oligomers. Bipedal DNA walkers have a larger walking area and faster walking kinetics, providing higher amplification efficiency compared to conventional DNA walkers. The DNA walker is driven via an Mg2+-dependent DNAzyme, and the binding-induced DNA walker will continuously clamp the MB, resulting in the proliferation of Fc confined near the GE surface. The linear range and limit of detection were 1 fg/mL to 10 pg/mL and 0.57 fg/mL, respectively. The proposed signal-on electrochemical sensing strategy is more selective. It will play a significant role in the sensitive and precise electrochemical analysis of other proteins.

Comprehensive analysis of prognosis of patients with GBM based on 4 m6A-related lncRNAs and immune cell infiltration.

Objective: To investigate the immune cell infiltration status in glioblastoma multiforme (GBM) and construct a novel prognostic risk model that can predict patients' prognosis. Methods: The Cancer Genome Atlas (TCGA) database was used to obtain RNA-sequence information and relevant clinical data. We performed Pearson correlation, univariate Cox regression to screen m6A-related prognostic lncRNA. GMB patients' samples were separated into different clusters through the ConsensusClusterPlus package. The risk score model was established through LASSO regression analysis. Besides, KEGG pathway enrichment analysis was implemented. CIBERSORT algorithm was used to analyze the difference of 22 types of immune cell infiltration in different cluster of GBM patient. Cox regression analyses were used to verify the independence of the model and correlation analysis was performed to demonstrate the link between our model and clinical characteristics of GBM patients. Experiments were used to validate the differential expression of the model lncRNA in patients with different prognosis. Results: 17 lncRNA related to prognosis were screened from 1021 m6A-related lncRNAs. Further, four m6A-related lncRNAs that were significantly correlated with GBM prognosis were selected to establish our prognostic risk model, which had excellent accuracy and can independently predict the prognosis of GBM patients. The infiltration fractions of T regulatory cells, T cells CD4 memory activated and neutrophils were positively associated with risk score, which suggested a significant relationship between the model and tumor immune microenvironment. Conclusion: The m6A-related RNA risk model offered potential for identifying biomarkers of therapy and predicting prognosis of GBM patients.

Proximity hybridization induced bipedal DNA walker for label-free electrochemical detection of apolipoprotein A4 based on DNA meditated Ag nanoparticles growth.

Apolipoprotein A4 (Apo-A4) is considered as a prospective molecular biomarker for diagnosis of depression due to its neurosynaptic toxicity. Here, we propose a neighboring hybridization induced catalyzed hairpin assembly (CHA) driven bipedal DNA walker that mediates hybridization of Ag nanoparticles (Ag NPs) with DNA probes for highly sensitive electrochemical quantitative detection of Apo-A4. Driven by CHA, this bipedal DNA walker can spread all over the surface of the sensor, induce the HP1-HP2 double chain structure, make the surface of the sensor negatively charged, and adsorb a large number of Ag ions. After chemical reduction with hydroquinone, the Ag NPs formed provide signal tracers for electrochemical dissolution analysis of the target. The Ag NPs formed by chemical reduction of hydroquinone can provide signal traces for electrochemical stripping analysis of target thrombin. The linear range of this method is from 10 pg mL-1 to 1000 ng mL-1, and the detection limit is 5.1 pg mL-1. This enzyme-free and labeling detection method provides a new strategy for rapid clinical detection of Apo-A4 and accurate identification of depression.

Integrating machine learning and bioinformatics analysis to m6A regulator-mediated methylation modification models for predicting glioblastoma patients' prognosis and immunotherapy response.

BACKGROUND: Epigenetic regulations of immune responses are essential for cancer development and growth. As a critical step, comprehensive and rigorous explorations of m6A methylation are important to determine its prognostic significance, tumor microenvironment (TME) infiltration characteristics and underlying relationship with glioblastoma (GBM). METHODS: To evaluate m6A modification patterns in GBM, we conducted unsupervised clustering to determine the expression levels of GBM-related m6A regulatory factors and performed differential analysis to obtain m6A-related genes. Consistent clustering was used to generate m6A regulators cluster A and B. Machine learning algorithms were implemented for identifying TME features and predicting the response of GBM patients receiving immunotherapy. RESULTS: It is found that the m6A regulatory factor significantly regulates the mutation of GBM and TME. Based on Europe, America, and China data, we established m6Ascore through the m6A model. The model accurately predicted the results of 1206 GBM patients from the discovery cohort. Additionally, a high m6A score was associated with poor prognoses. Significant TME features were found among the different m6A score groups, which demonstrated positive correlations with biological functions (i.e., EMT2) and immune checkpoints. CONCLUSIONS: m6A modification was important to characterize the tumorigenesis and TME infiltration in GBM. The m6Ascore provided GBM patients with valuable and accurate prognosis and prediction of clinical response to various treatment modalities, which could be useful to guide patient treatments.

IGFBP2 from a novel copper metabolism-associated biomarker promoted glioma progression and response to immunotherapy.

Introduction: Copper metabolism encompasses all cellular metabolic processes involving copper ions and plays a significant role in the pathogenesis of diseases, including cancer. Furthermore, copper is intricately involved in various processes related to nucleotide metabolism. However, a comprehensive analysis of copper metabolism in gliomas remains lacking despite its importance. Methods: To address this gap, glioma patients were stratified based on the expression levels of copper metabolism-related genes. By utilizing machine learning techniques, a novel copper metabolism-associated biomarker was developed. The potential of this biomarker in prognosis, mutation analysis, and predicting immunotherapy response efficiency in gliomas was systematically investigated. Results: Notably, IGFBP2, identified as a glioma tumor promoter, was found to promote disease progression and influence immunotherapy response. Additionally, glioma-derived IGFBP2 was observed to enhance microglial migration. High IGFBP2 expression in GBM cells facilitated macrophage interactions through the EGFR, CD63, ITGB1, and CD44 signaling pathways. Discussion: Overall, the copper metabolism-associated biomarker shows promising potential to enhance the clinical management of gliomas, offering valuable insights into disease prognosis and treatment strategies.

GPX8 regulates pan-apoptosis in gliomas to promote microglial migration and mediate immunotherapy responses.

Introduction: Gliomas have emerged as the predominant brain tumor type in recent decades, yet the exploration of non-apoptotic cell death regulated by the pan-optosome complex, known as pan-apoptosis, remains largely unexplored in this context. This study aims to illuminate the molecular properties of pan-apoptosis-related genes in glioma patients, classifying them and developing a signature using machine learning techniques. Methods: The prognostic significance, mutation features, immunological characteristics, and pharmaceutical prediction performance of this signature were comprehensively investigated. Furthermore, GPX8, a gene of interest, was extensively examined for its prognostic value, immunological characteristics, medication prediction performance, and immunotherapy prediction potential. Results: Experimental techniques such as CCK-8, Transwell, and EdU investigations revealed that GPX8 acts as a tumor accelerator in gliomas. At the single-cell RNA sequencing level, GPX8 appeared to facilitate cell contact between tumor cells and macrophages, potentially enhancing microglial migration. Conclusions: The incorporation of pan-apoptosis-related features shows promising potential for clinical applications in predicting tumor progression and advancing immunotherapeutic strategies. However, further in vitro and in vivo investigations are necessary to validate the tumorigenic and immunogenic processes associated with GPX8 in gliomas.

miR-150-3p enhances neuroprotective effects of neural stem cell exosomes after hypoxic-ischemic brain injury by targeting CASP2.

Brains are vulnerable to ischemic/hypoxic damage, which are directly caused by stroke, hypoxic-ischemic encephalopathy and other cerebral diseases. Currently, therapeutic strategies against cerebral ischemia and hypoxia are extremely limited. Recent studies have indicated that stem cell-derived exosomes play a neuroprotective role in hypoxic-ischemic brain injury. However, the treatment mechanism remains unclear. In this study, we cultured neural stem cells (NSCs) in vitro successfully. Exosomes isolated from NSCs (NSCs-Ex) inhibited the apoptosis while promoting the proliferation of SH-SY5Y cells both in normal and oxygen-glucose deprivation (OGD) culture conditions. Moreover, in vivo studies demonstrated that NSCs-Ex significantly reduced the infarction area in the middle cerebral artery occlusion (MCAO) model and suppressed the apoptosis of neurons. Furthermore, miR-150-3p was identified as the most abundantly expressed miRNA in exosomes compared to their parent NSCs. The miR-150-3p mimic displayed neuroprotective effects while miR-150-3p inhibitor exacerbated nerve injury both in vivo and in vitro. We further identified CASP2 as a miR-150-3p target. Thus, our data indicate that NSC-Ex facilitate the neuroprotective effects via transfer of miR-150-3p which targets CASP2, thus suppressing neuronal apoptosis after brain injury. Our results suggest that NSCs-Ex prevent cerebral injury by transferring miR-150-3p which promotes neurons proliferation by inhibiting CASP2 signaling pathway.

Beclin­1 exerts protective effects against cerebral ischemia­reperfusion injury by promoting DNA damage repair through a non­autophagy­dependent regulatory mechanism.

Cerebral ischemia­reperfusion (I/R) can result in severe brain injury, for which there are no optimal treatment options. I/R is often accompanied by increased autophagy. Beclin­1, a central player in autophagy, has been extensively studied in I/R; however, to date, at least to the best of our knowledge, there are no definitive descriptions of its specific role. Thus, the aim of the present study was to explore the regulatory role played by Beclin­1 in I/R. In vivo experiments were performed using an animal model of brain I/R with male Sprague­Dawley rats. Brain tissue damage was observed using 2,3,5­triphenyltetrazolium chloride, and hematoxylin and eosin staining. Tissue apoptosis levels were evaluated using a TUNEL assay, as well as western blot analysis. Immunofluorescence together with western blot analysis was used to detect autophagy in the tissues. Immunohistochemistry and western blot analysis were used to analyze DNA double­stranded breaks (DSBs). Moreover, HT22 cells overexpressing Beclin­1 were subjected to oxygen glucose deprivation/reoxygenation injury to simulate I/R pathological damage in vitro. Apoptosis was assessed using TUNEL and flow cytometric assays in this in vitro model, and autophagy was assessed using immunofluorescence and western blot analysis. The DSBs of the cells were analyzed using western blot analysis. I/R activated autophagy and induced DSBs. Autophagy inhibitors decreased brain tissue damage and reduced cell apoptosis; however, the degree of decrease in damage and apoptosis was not highly associated with the change in autophagy, and the frequency of DSBs slightly increased. The overexpression of Beclin­1 in neurons significantly attenuated I/R­induced damage and promoted DSB repair. On the whole, the present study demonstrates that Beclin­1 protects neurons from ischemic damage through the non­autophagy­dependent regulation of DNA repair processes.

NAP1L1 promotes proliferation and chemoresistance in glioma by inducing CCND1/CDK4/CDK6 expression through its interaction with HDGF and activation of c-Jun.

The prognosis of glioma is poor as its pathogenesis and mechanisms underlying cisplatin chemoresistance remain unclear. Nucleosome assembly protein 1 like 1 (NAP1L1) is regarded as a hallmark of malignant tumors. However, the role of NAP1L1 in glioma remains unknown. In this study, we aimed to investigate the molecular functions of NAP1L1 in glioma and its involvement in cisplatin chemoresistance, if any. NAP1L1 was found to be upregulated in samples from The Cancer Genome Atlas (TCGA) database. Immunohistochemistry indicated that NAP1L1 and hepatoma-derived growth factor (HDGF) were enhanced in glioma as compared to the para-tumor tissues. High expressions of NAP1L1 and HDGF were positively correlated with the WHO grade, KPS, Ki-67 index, and recurrence. Moreover, NAP1L1 expression was also positively correlated with the HDGF expression in glioma tissues. Functional studies suggested that knocking down NAP1L1 could significantly inhibit glioma cell proliferation both in vitro and in vivo, as well as enhance the sensitivity of glioma cells to cisplatin (cDDP) in vitro. Mechanistically, NAP1L1 could interact with HDGF at the protein level and they co-localize in the cytoplasm. HDGF knockdown in NAP1L1-overexpressing glioma cells significantly inhibited cell proliferation. Furthermore, HDGF could interact with c-Jun, an oncogenic transcription factor, which eventually induced the expressions of cell cycle promoters, CCND1/CDK4/CDK6. This finding suggested that NAP1L1 could interact with HDGF, and the latter recruited c-Jun, a key oncogenic transcription factor, that further induced CCND1/CDK4/CDK6 expression, thereby promoting proliferation and chemoresistance in glioma cells. High expression of NAP1L1 in glioma tissues indicated shorter overall survival in glioma patients.