Small-molecule Molephantin induces apoptosis and mitophagy flux blockage through ROS production in glioblastoma.

Glioblastoma (GBM), one of the most malignant brain tumors in the world, has limited treatment options and a dismal survival rate. Effective and safe disease-modifying drugs for glioblastoma are urgently needed. Here, we identified a small molecule, Molephantin (EM-5), effectively penetrated the blood-brain barrier (BBB) and demonstrated notable antitumor effects against GBM with good safety profiles both in vitro and in vivo. Mechanistically, EM-5 not only inhibits the proliferation and invasion of GBM cells but also induces cell apoptosis through the reactive oxygen species (ROS)-mediated PI3K/Akt/mTOR pathway. Furthermore, EM-5 causes mitochondrial dysfunction and blocks mitophagy flux by impeding the fusion of mitophagosomes with lysosomes. It is noteworthy that EM-5 does not interfere with the initiation of autophagosome formation or lysosomal function. Additionally, the mitophagy flux blockage caused by EM-5 was driven by the accumulation of intracellular ROS. In vivo, EM-5 exhibited significant efficacy in suppressing tumor growth in a xenograft model. Collectively, our findings not only identified EM-5 as a promising, effective, and safe lead compound for treating GBM but also uncovered its underlying mechanisms from the perspective of apoptosis and mitophagy.

Transcriptome analysis in hepatopancreases reveals the response of domesticated common carp to a high-temperature environment in the agricultural heritage rice-fish system.

Qingtian paddy field carp (PF-carp) is a local carp cultivated in the paddy field of Qingtian, Zhejiang. This rice-fish co-culture system has been recognized as one of the Globally Important Agriculture Heritage Systems (GIAHS). PF-carp has been acclimatized to the high-temperature environment of shallow paddy fields after several centuries of domestication. To reveal the physiological and molecular regulatory mechanisms of PF-carp, we chose to use 28°C as the control group and 34°C as the treatment group. We measured biochemical parameters in their serum and hepatopancreases and also performed transcriptome sequencing analysis. Compared with the control group, the serum levels of malondialdehyde (MDA), glucose (GLU), glutathione peroxidase (GSH-Px), catalase (CAT), alanine aminotransferase (ALT), and aspartate aminotransferase (AST) show no significant change. In addition, superoxide dismutase (SOD), GSH-Px, and CAT also show no significant change in hepatopancreases. We identified 1,253 differentially expressed genes (DEGs), and their pathway analysis revealed that heat stress affected AMPK signaling pathway, protein export, and other biological processes. It is worth noting that protein processing in the endoplasmic reticulum (ER) was the most significantly enriched pathway identified by the Kyoto Encyclopedia of Genes and Genomes (KEGG) and gene set enrichment analysis (GSEA). Significantly higher levels of HSP40, HSP70, HSP90, and other ubiquitin ligase-related genes were upregulated. In summary, heat stress did not lead to tissue damage, inflammation, oxidative stress, and ER stress in the hepatopancreases of PF-carp. This study provides valuable insights into the adaptation mechanism of this species to the high-temperature environment of paddy fields.

SOX2 downregulation of PML increases HCMV gene expression and growth of glioma cells.

The presence of human cytomegalovirus (HCMV) in glioblastoma (GBM) and improved outcomes of GBM patients receiving therapies targeting the virus have implicated HCMV in GBM progression. However, a unifying mechanism that accounts for the contribution of HCMV to the malignant phenotype of GBM remains incompletely defined. Here we have identified SOX2, a marker of glioma stem cells (GSCs), as a key determinant of HCMV gene expression in gliomas. Our studies demonstrated that SOX2 downregulated promyelocytic leukemia (PML) and Sp100 and consequently facilitated viral gene expression by decreasing the amount of PML nuclear bodies in HCMV-infected glioma cells. Conversely, the expression of PML antagonized the effects of SOX2 on HCMV gene expression. Furthermore, this regulation of SOX2 on HCMV infection was demonstrated in a neurosphere assay of GSCs and in a murine xenograft model utilizing xenografts from patient-derived glioma tissue. In both cases, SOX2 overexpression facilitated the growth of neurospheres and xenografts implanted in immunodeficient mice. Lastly, the expression of SOX2 and HCMV immediate early 1 (IE1) protein could be correlated in tissues from glioma patients, and interestingly, elevated levels of SOX2 and IE1 were predictive of a worse clinical outcome. These studies argue that HCMV gene expression in gliomas is regulated by SOX2 through its regulation of PML expression and that targeting molecules in this SOX2-PML pathway could identify therapies for glioma treatment.

Sp1 induced gene TIMP1 is related to immune cell infiltration in glioblastoma.

Tumor immune microenvironment exerts a profound effect on the population of infiltrating immune cells. Tissue inhibitor of matrix metalloproteinase 1 (TIMP1) is frequently overexpressed in a variety of cells, particularly during inflammation and tissue injury. However, its function in cancer and immunity remains enigmatic. In this study, we find that TIMP1 is substantially up-regulated during tumorigenesis through analyzing cancer bioinformatics databases, which is further confirmed by IHC tissue microarrays of clinical samples. The TIMP1 level is significantly increased in lymphocytes infiltrating the tumors and correlated with cancer progression, particularly in GBM. Notably, we find that the transcriptional factor Sp1 binds to the promoter of TIMP1 and triggers its expression in GBM. Together, our findings suggest that the Sp1-TIMP1 axis can be a potent biomarker for evaluating immune cell infiltration at the tumor sites and therefore, the malignant progression of GBM.

The identification of miRNA and mRNA expression profiles associated with pediatric atypical teratoid/rhabdoid tumor.

BACKGROUND: Atypical teratoid/rhabdoid tumor (AT/RT) is a malignant pediatric tumor of the central nervous system (CNS) with high recurrence and low survival rates that is often misdiagnosed. MicroRNAs (miRNAs) are involved in the tumorigenesis of numerous pediatric cancers, but their roles in AT/RT remain unclear. METHODS: In this study, we used miRNA sequencing and gene expression microarrays from patient tissue to study both the miRNAome and transcriptome traits of AT/RT. RESULTS: Our findings demonstrate that 5 miRNAs were up-regulated, 16 miRNAs were down-regulated, 179 mRNAs were up-regulated and 402 mRNAs were down-regulated in AT/RT. qPCR revealed that hsa-miR-17-5p and MAP7 mRNA were the most significantly differentially expressed miRNA and mRNA in AT/RT tissues. Furthermore, the results from analyses using the miRTarBase database identified MAP7 mRNA as a target gene of hsa-miR-17-5p. CONCLUSIONS: Our findings suggest that the dysregulation of hsa-miR-17-5p may be a pivotal event in AT/RT and miRNAs that may represent potential therapeutic targets and diagnostic biomarkers.

Molecular landscape of pediatric type IDH wildtype, H3 wildtype hemispheric glioblastomas.

The WHO (2021) Classification classified a group of pediatric-type high-grade gliomas as IDH wildtype, H3 wildtype but as of currently, they are characterized only by negative molecular features of IDH and H3. We recruited 35 cases of pediatric IDH wildtype and H3 wildtype hemispheric glioblastomas. We evaluated them with genome-wide methylation profiling, targeted sequencing, RNAseq, TERT promoter sequencing, and FISH. The median survival of the cohort was 27.6 months. With Capper et al.'s36 methylation groups as a map, the cases were found to be epigenetically heterogeneous and were clustered in proximity or overlay of methylation groups PXA-like (n = 8), LGG-like (n = 10), GBM_MYCN (n = 9), GBM_midline (n = 5), and GBM_RTKIII (n = 3). Histology of the tumors in these groups was not different from regular glioblastomas. Methylation groups were not associated with OS. We were unable to identify groups specifically characterized by EGFR or PDGFRA amplification as proposed by other authors. EGFR, PDGFRA, and MYCN amplifications were not correlated with OS. 4/9 cases of the GBM_MYCN cluster did not show MYCN amplification; the group was also enriched for EGFR amplification (4/9 cases) and the two biomarkers overlapped in two cases. Overall, PDGFRA amplification was found in only four cases and they were not restricted to any groups. Cases in proximity to GBM_midline were all hemispheric and showed loss of H3K27me3 staining. Fusion genes ALK/NTRK/ROS1/MET characteristic of infantile glioblastomas were not identified in 17 cases successfully sequenced. BRAF V600E was only found in the PXA group but CDKN2A deletion could be found in other methylation groups. PXA-like cases did not show PXA histological features similar to findings by other authors. No case showed TERT promoter mutation. Mutations of mismatch repair (MMR) genes were poor prognosticators in single (p ≤ 0.001) but not in multivariate analyses (p = 0.229). MGMT had no survival significance in this cohort. Of the other common biomarkers, only TP53 and ATRX mutations were significant poor prognosticators and only TP53 mutation was significant after multivariate analyses (p = 0.024). We conclude that IDH wildtype, H3 wildtype pediatric hemispheric glioblastomas are molecularly heterogeneous and in routine practice, TP53, ATRX, and MMR status could profitably be screened for risk stratification in laboratories without ready access to methylation profiling.

Mismatch repair proteins PMS2 and MLH1 can further refine molecular stratification of IDH-mutant lower grade astrocytomas.

The diagnostic role of Isocitrate Dehydrogenase (IDH) mutation status in adult lower grade astrocytomas was first formally presented within the WHO Classification of Tumours of the Central Nervous System (2016). IDH-mutant astrocytomas are not as common as IDH-wildtype astrocytomas but are of better prognosis. Our previous study provided an evident that IDH-mutant lower grade astrocytomas is not a homogeneous group and could be further stratified by PDGFRA amplification, CDK4 amplification and CDKN2A deletion. In this study, we detected the expressions of DNA mismatch repair (MMR) proteins (PMS2, MLH1, MSH2, MSH6) and PD-L1 by immunohistochemistry in 147 IDH-mutant lower grade astrocytomas and explored their clinical relevance. The loss of was identified in 28.6%, 1.4%, 8.8% and 13.6%, respectively. PD-L1 expression was detected in 1.4% of this cohort. Survival analysis revealed that loss of PMS2 was correlated with shorter OS (p < 0.001) and PFS (p = 0.005). Loss of PMS2 or MLH1 was associated with shorter OS (p < 0.001) and PFS (p = 0.008). In IDH-mutant lower grade astrocytomas without CDKN2A deletion, loss of PMS2 was associated with poorer OS (p < 0.001) and PFS (p = 0.001). Furthermore, among IDH-mutant lower grade astrocytomas lacking the three biomarkers (PDGFRA, CDK4 and CDKN2A), loss of PMS2 was also associated with a poorer OS (p < 0.001) and PFS (p = 0.003). Our data illustrated the potential application of MMR genes in stratification of IDH-mutant lower grade astrocytomas without PDGFRA, CDK4 and CDKN2A copy number alterations.

Nogo­66 promotes ß­amyloid protein secretion via NgR/ROCK­dependent BACE1 activation.

The generation of ß­amyloid protein (Aß) is considered a key step in the pathogenesis of Alzheimer's disease (AD) and the regulation of its production is an important therapeutic strategy. It was hypothesized in the present study that Nogo­A may be involved in AD and may regulate the generation of Aß. Nogo­A is known to act as a major inhibitor of neuron regeneration in the adult central nervous system. A recent study indicated that Nogo­A is associated with AD; however, the underlying effect and molecular mechanisms remain largely elusive. In the present study, the potential effects of Nogo­A on AD were investigated. ELISA was used to detect the levels of Aß, enzymatic activity detection kits were used to determine the activity of secretase enzymes in amyloid precursor protein (APP) metabolism, and western blot analysis was used to detect the expression levels of proteins associated with the APP processing and Nogo­A/Nogo­66 receptor (NgR) signaling pathways. The results revealed that Nogo­66, the major inhibitory region of Nogo­A, promoted neuronal Aß secretion by increasing the activity of ß­secretase 1 via the NgR/Rho­associated coiled­coil containing kinases pathway in a dose­dependent manner. The present data suggested that Nogo­A may facilitate the onset and development of AD by promoting Aß secretion, providing information on a potential novel target for AD therapy.

Self-Assembly Behavior of Diacetylenic Acid Molecules upon Vapor Deposition: Odd-Even Effect on the Film Morphology.

A series of linear carboxylic acids containing diacetylenic units at different positions along the chain (C12 H25 (C≡C)2 (CH2 )n COOH, n=7-11) were vacuum-deposited on clean silica substrates. The morphologies of the initial films after UV irradiation were studied. A clear odd-even effect on the morphology of the initial film was observed in that, depending on the spacer length between the diacetylenic unit and carboxyl head group, rings or dendrites of acid dimer layers were obtained. A molecular dynamic simulation of the aggregation process suggests that two competing intermolecular interactions and thus aggregation directions are involved and modulated by the odd or even carbon chain length. Further modulation of the interaction by substitution of a phenyl group at the terminus of the chain or by changing the carboxyl head group to an amidobenzoic acid head group led to a similar odd-even effect but with different dimensions or trends, which can be rationalized similarly. These results give the opportunity to create aligned conjugated polymer chains of different dimensions through self-assembly for applications in molecular/organic electronics.

The CXCL12/CXCR4 axis confers temozolomide resistance to human glioblastoma cells via up-regulation of FOXM1.

Glioblastoma multiforme (GBM) is the most common primary malignancy in the adult central nervous, and is characterized by high aggressiveness and a high mortality rate. The high mortality rate is largely due to the development of drug resistance. Temozolomide (TMZ) resistance is considered to be one of the major reasons responsible for GBM therapy failure. CXCL12/CXCR4 has been demonstrated to be involved in cell proliferation, migration, invasion, angiogenesis, and radioresistance in GBM. However, its role in TMZ resistance in GBM is unknown. In this study, we aimed to evaluate the role of CXCL12/CXCR4 in mediating the TMZ resistance to GBM cells and explore the underlying mechanisms. We found that the CXCL12/CXCR4 axis enhanced TMZ resistance in GBM cells. Further study showed that CXCL12/CXCR4 conferred TMZ resistance and promoted the migration and invasion of GBM cells by up-regulating FOXM1. This resistance was partially reversed by suppressing CXCL12/CXCR4 and FOXM1 silencing. Our study revealed the vital role of CXCL12/CXCR4 in mediating the resistance of GBM cells to TMZ, and suggested that targeting CXCL12/CXCR4 axis may attenuate the resistance to TMZ in GBM.

Integrative genomic analyses identify WDR12 as a novel oncogene involved in glioblastoma.

Glioblastoma (GBM) is the most malignant primary brain tumor in adults. Due to its invasive nature, it cannot be thoroughly eliminated. WD repeat domain 12 (WDR12) processes the 32S precursor rRNA but cannot affect the synthesis of the 45S/47S primary transcript. In this study, we found that WDR12 is highly expressed in GBM according to the analysis results of mRNA expression by The Cancer Genome Atlas database. The high expression level of WDR12 is dramatically related to shorter overall survival and reduced disease-free survival. Next, we knocked down WDR12 and found that knockdown of WDR12 promoted the apoptosis and inhibited the proliferation by cell biology experiments. Differential expression genes in gene-chip revealed that WDR12 knockdown mainly inhibited cell cycle. Finally, we also found that WDR12 is associated with PLK1 and EZH2 in cell proliferation of GBM. Resumptively, this report showed a possible evidence that WDR12 drove malignant behavior of GBM, whose expression may present a neoteric independent prognostic biomarker in GBM.

Pediatric primary diffuse leptomeningeal melanomatosis: Case report and review of the literature.

RATIONALE: Primary melanocytic tumors of central nerve system (CNS) are rare, primary diffuse leptomeningeal melanomatosis (PDLM), a subtype of malignant melanomas of CNS, is extremely rare,especially in pediatrics. As the clinical manifestation of PDLM is not characteristic, It is often misdiagnosed as tubercular meningitis and hemorrhage. PATIENT CONCERNS: A 13-year-old boy was admitted to our department with symptoms of recurrent headache and vomiting twice. As the brain imaging revealed a lesion located in the left temporal lobe mimicked hemorrhage, so there was a misdiagnosis of hemorrhage in first hospitalization. He was admitted again for the recurrence of the headache and vomiting. Detailed physical examination showed multiple melanin changes in the skin of the whole body which were ignored in last hospitalization. Brain imaging showed the significantly enlarged lesion in the left temporal lobe and several smaller lesions in the left parietal lobe and cerebellum which indicated metastasis. DIAGNOSIS: According to the history,physical examination and the radiological finding, the patient was diagnosed with malignant melanoma of central never system possibly. INTERVENTIONS: The patient underwent left temporal and parietal lesions total resection with a craniotomy. OUTCOMES: The diagnosis of PDLM was established according to pathological characteristics and the negative finding of positron emission tomography (PET)-computed tomography (CT) outside CNS. The patient got no further treatment for economic reasons and experienced the progression and died 5 months after operation. LESSONS: PDLM is extremely rare in CNS, as the clinical manifestation, radiological changes are not special, early diagnosis is difficult. The confirmed diagnosis is established by leptomeningeal biospy or surgical tissue. PET-CT can help differential diagnosis with metastastic leptomeningeal melanomas. The prognosis is dismal due to the inefficiency of chemotherapy or radiotherapy.

Multiple Cavernous Malformations of Brain, Chest, and Skin: A Rare Case of an Infant and Literature Review.

BACKGROUND: Cerebral cavernous malformations (CCMs) are vascular malformations that account for 5%-15% of all central nervous system vascular malformations. However, multiple CCMs, which can be sporadic or familial, are rare, with a prevalence of 0.1%-0.5%. CASE DESCRIPTION: Here, we presented a rare case of sporadic multiple CCMs in an infant, which were accompanied with multiple cavernous malformations of the chest and skin. CONCLUSIONS: CCMs were pathologically diagnosed through the total resection of the pineal regional lesion. We also observed a spontaneous regression of the remaining lesions during a follow-up period of 2 years. To our knowledge, this is the first case of CCMs in an infant in the English-language literature.

The PDK1/c­Jun pathway activated by TGF­ß induces EMT and promotes proliferation and invasion in human glioblastoma.

Glioblastoma multiforme (GBM) is the most common primary malignant tumor affecting the human brain. Despite improvements in therapeutic technologies, patients with GBM have a poor clinical result and the molecular mechanisms responsible for the development of GBM have not yet been fully elucidated. 3-phosphoinositide dependent protein kinase 1 (PDK1) is upregulated in various tumors and promotes tumor invasion. In glioma, transforming growth factor-ß (TGF­ß) promotes cell invasion; however, whether TGF­ß directly regulates PDK1 protein and promotes proliferation and invasion is not yet clear. In this study, PDK1 levels were measured in glioma tissues using tissue microarray (TMA) by immunohistochemistry (IHC) and RT­qPCR. Kaplan-Meier analyses were used to calculate the survival rate of patients with glioma. In vitro, U251 and U87 glioma cell lines were used for functional analyses. Cell proliferation and invasion were analyzed using siRNA transfection, MTT assay, RT­qPCR, western blot analysis, flow cytometry and invasion assay. In vivo, U251 glioma cell xenografts were established. The results revealed that PDK1 protein was significantly upregulated in glioma tissues compared with non-tumorous tissues. Furthermore, the higher PDK1 levels were associated with a large tumor size (>5.0 cm), a higher WHO grade and a shorter survival of patients with GBM. Univariate and multivariate analyses indicated that PDK1 was an independent prognostic factor. In vivo, PDK1 promoted glioma tumor xenograft growth. In vitro, functional analyses confirmed that TGF­ß upregulated PDK1 protein expression and PDK1 promoted cell migration and invasion, and functioned as an oncogene in GBM, by upregulating c­Jun protein and inducing epithelial-mesenchymal transition (EMT). c­Jun protein were overexpressed in glioma tissues and positively correlated with PDK1 levels. Moreover, our findings were further validated by the online Oncomine database. On the whole, the findings of this study indicate that in GBM, PDK1 functions as an oncogene, promoting proliferation and invasion.

Underlying mechanism of the photodynamic activity of hematoporphyrin­induced apoptosis in U87 glioma cells.

Photodynamic therapy (PDT) is a relatively novel type of tumor therapy method with low toxicity and limited side­effects. The aim of the present study was to investigate the underlying mechanism and potential microRNAs (miRNAs) involved in the treatment of glioma by PDT with hematoporphyrin, a clinical photosensitizer. The photodynamic activity of hematoporphyrin on the cell viability and apoptosis of gliomas was investigated by MTT, and flow cytometry and fluorescence microscopy, respectively. Alterations in singlet oxygen and mitochondrial membrane potential were detected. The differentially expressed miRNAs and proteins were evaluated by miRNA gene chip and apoptosis­associated protein chip, respectively. The results demonstrated that cell viability significantly decreased with hematoporphyrin concentration. PDT with hematoporphyrin significantly increased cell apoptosis at a later stage, induced the content of reactive oxygen species (ROS) and decreased the mitochondrial membrane potential, indicating that PDT with hematoporphyrin inhibited cell growth via induction of radical oxygen, decreased the mitochondrial membrane potential and induced apoptosis. The upregulated miRNAs, including hsa­miR­7641, hsa­miR­9500, hsa­miR­4459, hsa­miR­21­5p, hsa­miR­663a and hsa­miR­205­5p may be important in PDT­induced cell apoptosis in glioma. Transporter 1, ATP binding cassette subfamily B member­ and nuclear factor­κB­mediated apoptosis signaling pathways were the most significant pathways. Thus, the current study presents PDT as a potential therapeutic approach for the treatment of malignant glioma, and identified miRNAs for the molecular design and development of a third­generation photosensitizer (PS).

Downregulation of miR­205 is associated with glioblastoma cell migration, invasion, and the epithelial-mesenchymal transition, by targeting ZEB1 via the Akt/mTOR signaling pathway.

Glioblastoma (GBM) is the most common type of malignant brain tumor. In spite of recent advancements in surgical techniques, chemotherapy, and radiation therapy, patients with GBM often face a dire prognosis. MicroRNAs have been shown to modulate the aggressiveness of various cancers, and have emerged as possible therapeutic agents for the management of GBM. miR­205 is dysregulated in glioma and act as a prognostic indicator. However, the role of miR­205 in the development of GBM has not been elucidated. To better understand the pathogenesis of GBM, we examine the biological significance and molecular mechanisms of miR­205 in GBM cells. Zinc finger E-box binding homeobox 1 (ZEB1) has been shown to regulate the epithelial-mesenchymal transition (EMT), which is strongly associated with GBM malignancy. In the present study, we show miR­205 expression is reduced in GBM tissues and cell lines, and ZEB1 expression is inversely correlated with miR­205 expression. We also show ZEB1 is a downstream target of miR­205 and the Akt/mTOR signaling pathway is activated when miR­205 interacts with ZEB1. Increased activity of miR­205 in GBM cells significantly inhibits migration and invasion, and prevents EMT. Furthermore, overexpression of ZEB1 partially abolishes these inhibitory effects of miR­205. We show that miR­205 negatively regulates the expression of ZEB1 in GBM, inhibits cell migration and invasion, and prevents EMT, at least in part through the inhibition of the activation of the Akt/mTOR signaling pathway. Our results indicate miR­205 may be an efficacious therapeutic agent in the treatment of GBM.

The effect of allogeneic cardiac stem cells in left ventricular geometry and function in a rat model of myocardial infarction.

BACKGROUND: The development of heart failure after acute myocardial infarction (MI) was related to left ventricular (LV) pathological remodeling and dysfunction. Cardiac stem cells (CSCs) provided a new option to treat acute MI. This study was to investigate the effects of CSCs on structural and functional alteration in acute MI. METHODS: Acute MI was induced in 20 Sprague-Dawley rats by ligation of the left anterior descending coronary artery. Two weeks after MI, animals were randomized into CSCs or control group. LV geometry and function were echocardiographically measured at baseline, 2, 4, and 6 weeks after MI. After measuring hemodynamics at 6 weeks after MI, hearts were harvested for tracing CSCs stained by PKH26 and testing expression of VEGF-α/TGF-ß1 by RT-PCR and ELISA. RESULTS: Two weeks after MI, there were significant decreases in interventricular septal systolic and diastolic thickness (IVSTs/d), while increases in LV systolic and diastolic dimension (LVDs/d). Consequently, this contributed to decreases in ejection fraction (EF) and fractional shorting (FS). With the treatment of CSCs for 4 weeks, significant better ejection fraction (EF) and fractional shorting (FS) were achieved in CSCs group accompanied by the reduction in LV systole and diastole dimension (LVDs/d). Besides, a significant improvement in the maximal rate of LV pressure development and decline (peak +dP/dt and -dP/dt, respectively) was observed. Moreover, significantly higher VEGF-α was expressed in CSCs group rather than TGF-ß1. CONCLUSION: CSCs significantly prevented structural and functional deterioration after MI with increasing expression of VEGF-α.