Exploring neuron-specific steroid synthesis and DHEAS therapy in Alzheimer's disease.

Alzheimer's Disease (AD) is a neurodegenerative disorder characterized by cognitive decline and memory loss. Recent studies have suggested a potential role for steroid synthesis in AD pathology. This study investigated the co-localization of steroidogenic enzymes in neuronal cells, changes in enzyme expression in an AD mouse model, and steroid expressions in human AD samples. Additionally, we conducted a steroidomic metabolomics analysis and evaluated the effects of dehydroepiandrosterone sulfate (DHEAS) treatment in an AD mouse model. Immunofluorescence analysis revealed significant co-localization of cytochrome P450 family 17 subfamily A member 1 (CYP17A1) and steroidogenic acute regulatory protein (StAR) proteins with α-synuclein in presynaptic neurons, suggesting active steroid synthesis in these cells. Conversely, such co-localization was absent in astrocytes. In the AD mouse model, a marked decrease in the expression of steroidogenic enzymes (Cyp11a1, Cyp17a1, Star) was observed, especially in areas with amyloid beta plaque accumulation. Human AD and MS brain tissues showed similar reductions in StAR and CYP17A1 expressions. Steroidomic analysis indicated a downregulation of key steroids in the serum of AD patients. DHEAS treatment in AD mice resulted in improved cognitive function and reduced Aß accumulation. Our findings indicate a neuron-specific pathway for steroid synthesis, potentially playing a crucial role in AD pathology. The reduction in steroidogenic enzymes and key steroids in AD models and human samples suggests that impaired steroid synthesis is a feature of neurodegenerative diseases. The therapeutic potential of targeting steroid synthesis pathways, as indicated by the positive effects of DHEAS treatment, warrants further investigation.

Astrocytic Cebpd Regulates Pentraxin 3 Expression to Promote Fibrotic Scar Formation After Spinal Cord Injury.

Astroglial-fibrotic scars resulted from spinal cord injury affect motor and sensory function, leading to paralysis. In particular, the fibrotic scar is a main barrier that disrupts neuronal regeneration after spinal cord injury. However, the association between astrocytes and fibrotic scar formation is not yet understood. We have previously demonstrated that the transcriptional factor Cebpd contributes to astrogliosis, which promotes glial scar formation after spinal cord injury. Herein, we show that fibrotic scar formation was decreased in the epicenter region in Cebpd-/- mice after contusive spinal cord injury and astrocytic Cebpd promoted fibroblast migration through secretion of Ptx3. Furthermore, the expression of Mmp3 was increased under recombinant protein Ptx3 treatment in fibroblasts by observing microarray data, resulting in fibroblast migration. In addition, regulation of Mmp3 occurs through the NFκB signaling pathway by using an irreversible inhibitor of IκBα phosphorylation in pretreated fibroblasts. Of note, we used the synthetic peptide RI37, which blocks fibroblast migration and decreases fibroblast Mmp3 expression in IL-1ß-treated astrocyte conditioned media. Collectively, our data suggest that fibroblast migration can be affected by astrocytic Cebpd through the Ptx3/NFκB/Mmp3 axis pathway and that the RI37 peptide may act as a therapeutic medicine to inhibit fibrotic scar formation after spinal cord injury.

Inhibition of MZF1/c-MYC Axis by Cantharidin Impairs Cell Proliferation in Glioblastoma.

Myeloid zinc finger 1 (MZF1), also known as zinc finger protein 42, is a zinc finger transcription factor, belonging to the Krüppel-like family that has been implicated in several types of malignancies, including glioblastoma multiforme (GBM). MZF1 is reportedly an oncogenic gene that promotes tumor progression. Moreover, higher expression of MZF1 has been associated with a worse overall survival rate among patients with GBM. Thus, MZF1 may be a promising target for therapeutic interventions. Cantharidin (CTD) has been traditionally used in Chinese medicine to induce apoptosis and inhibit cancer cell proliferation; however, the mechanism by which CTD inhibits cell proliferation remains unclear. In this study, we found that the expression of MZF1 was higher in GBM tissues than in adjacent normal tissues and low-grade gliomas. Additionally, the patient-derived GBM cells and GBM cell lines presented higher levels of MZF1 than normal human astrocytes. We demonstrated that CTD had greater anti-proliferative effects on GBM than a derivative of CTD, norcantharidin (NCTD). MZF1 expression was strongly suppressed by CTD treatment. Furthermore, MZF1 enhanced the proliferation of GBM cells and upregulated the expression of c-MYC, whereas these effects were reversed by CTD treatment. The results of our study suggest that CTD may be a promising therapeutic agent for patients with GBM and suggest a promising direction for further investigation.

Blockade of the pentraxin 3/CD44 interaction attenuates lung injury-induced fibrosis.

BACKGROUND: Fibrosing interstitial lung diseases (fILD) are potentially fatal with limited therapeutic options and no effective strategies to reverse fibrogenesis. Myofibroblasts are chief effector cells in fibrosis that excessively deposit collagen in the pulmonary interstitium and lead to progressive impairment of gaseous exchange. METHODS: Plasma and lung specimens from patients with fILD were applied for detecting pentraxin 3 (PTX3) abundance by ELISA and Immunohistochemistry. Masson's trichrome and Sirius red stains and hydroxyproline assay were performed for assessing collagen accumulation in the lungs of bleomycin-exposed conditional Ptx3-deficient and PTX3-neutralizing antibody (αPTX3i)-treated mice. Downstream effectors including signaling pathways and fibrotic genes were examined for assessing CD44-involved PTX3-induced fibrosis in HFL1 and primary mouse fibroblasts. RESULTS: PTX3 was upregulated in the lungs and plasma of bleomycin-exposed mice and correlated with disease severity and adverse outcomes in fILD patients. Decreased collagen accumulation, attenuation of alveolar fibrosis and fibrotic markers, and improved lung function were observed in bleomycin-exposed conditional Ptx3-deficient mice. PTX3 activates lung fibroblasts to differentiate towards migrative and highly collagen-expressing myofibroblasts. Lung fibroblasts with CD44 inactivation attenuated the PI3K-AKT1, NF-κB, and JNK signaling pathways and fibrotic markers. αPTX3i mimic-based therapeutic studies demonstrated abrogation of the migrative fibroblast phenotype and myofibroblast activation in vitro. Notably, αPTX3i inhibited lung fibrosis, reduced collagen deposition, increased mouse survival, and improved lung function in bleomycin-induced pulmonary fibrosis. CONCLUSIONS: The present study reveals new insights into the involvement of the PTX3/CD44 axis in fibrosis and suggests PTX3 as a promising therapeutic target in fILD patients.

CCAAT/Enhancer-Binding Protein Delta Regulates Glioblastoma Survival through Catalase-Mediated Hydrogen Peroxide Clearance.

It has long been documented that cancer cells show increased and persistent oxidative stress due to increased reactive oxygen species (ROS), which is necessary for their increased proliferative rate. Due to the high levels of ROS, cancer cells also stimulate the antioxidant system, which includes the enzymes superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX), to eliminate ROS. However, overexpressed antioxidant enzymes often lead to drug resistance and therapeutic failure. Glioblastoma (GBM) is the most aggressive brain tumor and has the poorest prognosis. The transcription factor CCAAT/enhancer-binding protein delta (CEBPD) is highly expressed in GBM and correlates with drug resistance, prompting us to elucidate its role in GBM cell survival. In this study, we first demonstrated that loss of CEBPD significantly inhibited GBM cell viability and increased cell apoptosis. Furthermore, the expression of CAT was attenuated through promoter regulation following CEBPD knockdown, accelerating intracellular hydrogen peroxide (H2O2) accumulation. In addition, mitochondrial function was impaired in CEBPD knockdown cells. Together, we revealed the mechanism by which CEBPD-mediated CAT expression regulates H2O2 clearance for GBM cell survival.

Reprogramming of arachidonate metabolism confers temozolomide resistance to glioblastoma through enhancing mitochondrial activity in fatty acid oxidation.

BACKGROUND: Sp1 is involved in the recurrence of glioblastoma (GBM) due to the acquirement of resistance to temozolomide (TMZ). Particularly, the role of Sp1 in metabolic reprogramming for drug resistance remains unknown. METHODS: RNA-Seq and mass spectrometry were used to analyze gene expression and metabolites amounts in paired GBM specimens (primary vs. recurrent) and in paired GBM cells (sensitive vs. resistant). ω-3/6 fatty acid and arachidonic acid (AA) metabolism in GBM patients were analyzed by targeted metabolome. Mitochondrial functions were determined by Seahorse XF Mito Stress Test, RNA-Seq, metabolome and substrate utilization for producing ATP. Therapeutic options targeting prostaglandin (PG) E2 in TMZ-resistant GBM were validated in vitro and in vivo. RESULTS: Among the metabolic pathways, Sp1 increased the prostaglandin-endoperoxide synthase 2 expression and PGE2 production in TMZ-resistant GBM. Mitochondrial genes and metabolites were obviously increased by PGE2, and these characteristics were required for developing resistance in GBM cells. For inducing TMZ resistance, PGE2 activated mitochondrial functions, including fatty acid ß-oxidation (FAO) and tricarboxylic acid (TCA) cycle progression, through PGE2 receptors, E-type prostanoid (EP)1 and EP3. Additionally, EP1 antagonist ONO-8713 inhibited the survival of TMZ-resistant GBM synergistically with TMZ. CONCLUSION: Sp1-regulated PGE2 production activates FAO and TCA cycle in mitochondria, through EP1 and EP3 receptors, resulting in TMZ resistance in GBM. These results will provide us a new strategy to attenuate drug resistance or to re-sensitize recurred GBM.

Allopregnanolone suppresses glioblastoma survival through decreasing DPYSL3 and S100A11 expression.

Allopregnanolone (allo) is a physiological regulator of neuronal activity that treats multiple neurological disorders. Allo penetrates the blood-brain barrier with very high efficiency, implying that allo can treat CNS-related diseases, including glioblastoma (GBM), which always recurs after standard therapy. Hence, this study aimed to determine whether allo has a therapeutic effect on GBM. We found that allo enhanced temozolomide (TMZ)-suppressed cell survival and proliferation of TMZ-resistant cells. In particular, allo enhanced TMZ-inhibited cell migration and TMZ-induced apoptosis. Additionally, allo strongly induced DNA damage characterized by γH2Ax. Furthermore, quantitative proteomic analysis, iTRAQ, showed that allo significantly decreased the levels of DPYSL3, S100A11, and S100A4, reflecting the poor prognosis of patients with GBM confirmed by differential gene expression and survival analysis. Moreover, single-cell RNA-Seq revealed that S100A11, expressed in malignant cells, oligodendrocytes, and macrophages, was significantly associated with immune cell infiltration. Furthermore, overexpression of DPYSL3 or S100A11 prevented allo-induced cell death. In conclusion, allo suppresses GBM cell survival by decreasing DPYSL3/S100A11 expression and inducing DNA damage.

Histone deacetylase 6 acts upstream of DNA damage response activation to support the survival of glioblastoma cells.

DNA repair promotes the progression and recurrence of glioblastoma (GBM). However, there remain no effective therapies for targeting the DNA damage response and repair (DDR) pathway in the clinical setting. Thus, we aimed to conduct a comprehensive analysis of DDR genes in GBM specimens to understand the molecular mechanisms underlying treatment resistance. Herein, transcriptomic analysis of 177 well-defined DDR genes was performed with normal and GBM specimens (n = 137) from The Cancer Genome Atlas and further integrated with the expression profiling of histone deacetylase 6 (HDAC6) inhibition in temozolomide (TMZ)-resistant GBM cells and patient-derived tumor cells. The effects of HDAC6 inhibition on DDR signaling were examined both in vitro and intracranial mouse models. We found that the expression of DDR genes, involved in repair pathways for DNA double-strand breaks, was upregulated in highly malignant primary and recurrent brain tumors, and their expression was related to abnormal clinical features. However, a potent HDAC6 inhibitor, MPT0B291, attenuated the expression of these genes, including RAD51 and CHEK1, and was more effective in blocking homologous recombination repair in GBM cells. Interestingly, it resulted in lower cytotoxicity in primary glial cells than other HDAC6 inhibitors. MPT0B291 reduced the growth of both TMZ-sensitive and TMZ-resistant tumor cells and prolonged survival in mouse models of GBM. We verified that HDAC6 regulated DDR genes by affecting Sp1 expression, which abolished MPT0B291-induced DNA damage. Our findings uncover a regulatory network among HDAC6, Sp1, and DDR genes for drug resistance and survival of GBM cells. Furthermore, MPT0B291 may serve as a potential lead compound for GBM therapy.

17ß-estradiol induces temozolomide resistance through NRF2-mediated redox homeostasis in glioblastoma.

Glioblastoma multiforme (GBM) is the most fatal cancer among brain tumors, and the standard treatment of GBM patients is surgical tumor resection followed by radiotherapy and temozolomide (TMZ) chemotherapy. However, tumors always recur due to the developing drug resistance. It has been shown that neurosteroids, including dehydroepiandrosterone and 17ß-estradiol, are synthesized in TMZ-resistant GBM tumors. Therefore, we sought to explore the possible role of 17ß-estradiol in the development of drug resistance in GBM. Bioinformatics analysis revealed that aromatase/cytochrome P450 19A1 expression was gradually increased in the development from normal, astrocytoma to GBM. The level of 17ß-estradiol was significantly increased in TMZ-resistant cells characterized by ultra performance liquid chromatography-tandem mass spectrometry. Furthermore, 17ß-estradiol attenuated TMZ-induced cell death and reduced reactive oxygen species production by mitochondria. In addition, 17ß-estradiol attenuated oxidative stress by increasing the expression of superoxide dismutase 1/2, catalase, and nuclear factor erythroid 2-related factor (NRF) 2. We found that NRF2 expression was essential for the induction of drug resistance by 17ß-estradiol through the reduction of oxidative stress in GBM.

Correlation between the expression of cancer stem cell marker BMI1 and glioma prognosis.

B-cell-specific Moloney murine leukemia virus integration site 1 (BMI1) appears to be essential for promoting certain types of cancer, and its inhibition effectively reduced the stemness of cancer cells. Therefore, this study aimed to investigate the potential role of BMI1 in glioma. To this end, we first investigated BMI1 expression in brain tumors using microarray datasets in ONCOMINE, which indicated that BMI1 levels were not commonly increased in clinical brain tumors. Moreover, survival plots in PROGgeneV2 also showed that BMI1 expression was not significantly associated with reduced survival in glioma patients. Interestingly, stressful serum deprivation and anchorage independence growth conditions led to an increased BMI1 expression in glioma cells. A stress-responsive pathway, HDAC/Sp1, was further identified to regulate BMI1 expression. The HDAC inhibitor vorinostat (SAHA) prevented Sp1 binding to the BMI1 promoter, leading to a decreased expression of BMI1 and attenuating tumor growth of TMZ-resistant glioma xenografts. Importantly, we further performed survival analysis using PROGgeneV2 and found that an elevated expression of HDAC1,3/Sp1/BMI1 but not BMI1 alone showed an increased risk of death in both high- and low-grade glioma patients. Thus, HDAC-mediated Sp1 deacetylation is critical for BMI1 regulation to attenuate stress- and therapy-induced death in glioma cells, and the HDAC/Sp1 axis is more important than BMI1 and appears as a therapeutic target to prevent recurrence of malignant glioma cells persisting after primary therapy.

CCAAT/Enhancer-binding protein delta mediates glioma stem-like cell enrichment and ATP-binding cassette transporter ABCA1 activation for temozolomide resistance in glioblastoma.

Glioblastoma (GBM) is the most aggressive brain tumor and relapses after chemo- or radiotherapy in a short time. The anticancer drug temozolamide (TMZ) is commonly used for GBM treatment, but glioma stem-like cells (GSCs) often lead to drug resistance and therapeutic failure. To date, the mechanism of GSC formation in TMZ-treated GBM remains largely unknown. CCAAT/Enhancer-binding protein delta (CEBPD) is an inflammation-responsive transcription factor and is proposed to be oncogenic in the context of drug resistance, prompting us to clarify its role in TMZ-resistant GBM. In this study, we first found that the CEBPD protein levels in GBM patients were significantly increased and further contributed to TMZ resistance by promoting GSC formation. Accordingly, the protein levels of stemness transcription factors, namely, SRY-box transcription factor 2 (SOX2), octamer-binding transcription factor 4 (OCT4), NANOG, and ATP-binding cassette subfamily A member 1 (ABCA1), were increased in GSCs and TMZ-treated GBM cells. Increased binding of CEBPD to promoter regions was observed in GSCs, indicating the direct regulation of these GSC-related genes by CEBPD. In addition, an ABCA1 inhibitor increased the caspase 3/7 activity of TMZ-treated GSCs, suggesting that TMZ efflux is controlled by ABCA1 activity and that the expression levels of the ABCA1 gene are an indicator of the efficiency of TMZ treatment. Together, we revealed the mechanism of CEBPD-mediated GSC drug resistance and proposed ABCA1 inhibition as a potential strategy for the treatment of TMZ-resistant GBM.

AR ubiquitination induced by the curcumin analog suppresses growth of temozolomide-resistant glioblastoma through disrupting GPX4-Mediated redox homeostasis.

Drug resistance is the main obstacle in the improvement of chemotherapeutic efficacy in glioblastoma. Previously, we showed that dehydroepiandrosterone (DHEA), one kind of androgen/neurosteroid, potentiates glioblastoma to acquire resistance through attenuating DNA damage. Androgen receptor (AR) activated by DHEA or other types of androgen was reported to promote drug resistance in prostate cancer. However, in DHEA-enriched microenvironment, the role of AR in acquiring resistance of glioblastoma remains unknown. In this study, we found that AR expression is significantly correlated with poor prognosis, and AR obviously induced the resistance to temozolomide (TMZ) treatment. Herein, we observed that ALZ003, a curcumin analog, induces FBXL2-mediated AR ubiquitination, leading to degradation. Importantly, ALZ003 significantly inhibited the survival of TMZ-sensitive and -resistant glioblastoma in vitro and in vivo. The accumulation of reactive oxygen species (ROS), lipid peroxidation and suppression of glutathione peroxidase (GPX) 4, which are characteristics of ferroptosis, were observed in glioblastoma cell after treatment of ALZ003. Furthermore, overexpression of AR prevented ferroptosis in the presence of GPX4. To evaluate the therapeutic effect in vivo, we transplanted TMZ-sensitive or -resistant U87MG cells into mouse brain followed by intravenous administration with ALZ003. In addition to inhibiting the growth of glioblastoma, ALZ003 significantly extended the survival period of transplanted mice, and significantly decreased AR expression in the tumor area. Taken together, AR potentiates TMZ resistance for glioblastoma, and ALZ003-mediated AR ubiquitination might open a new insight into therapeutic strategy for TMZ resistant glioblastoma.

CYP17A1 Maintains the Survival of Glioblastomas by Regulating SAR1-Mediated Endoplasmic Reticulum Health and Redox Homeostasis.

Cytochrome P450 (CYP) 17A1 is an important steroidogenic enzyme harboring 17α-hydroxylase and performing 17,20 lyase activities in multiple steps of steroid hormone synthesis, including dehydroepiandrosterone (DHEA) biosynthesis. Previously, we showed that CYP17A1-mediated DHEA production clearly protects glioblastomas from temozolomide-induced apoptosis, leading to drug resistance. Herein, we attempt to clarify whether the inhibition of CYP17A1 has a tumor-suppressive effect, and to determine the steroidogenesis-independent functions of CYP17A1 in glioblastomas. Abiraterone, an inhibitor of CYP17A1, significantly inhibits the proliferation of A172, T98G, and PT#3 (the primary glioblastoma cells) by inducing apoptosis. In parallel, abiraterone potently suppresses tumor growth in mouse models through transplantation of PT#3 cells to the back or to the brain. Based on evidence that abiraterone induces endoplasmic reticulum (ER) stress, followed by the accumulation of reactive oxygen species (ROS), CYP17A1 is important for ER health and redox homeostasis. To confirm our hypothesis, we showed that CYP17A1 overexpression prevents the initiation of ER stress and attenuates ROS production by regulating SAR1a/b expression. Abiraterone dissociates SAR1a/b from ER-localized CYP17A1, and induces SAR1a/b ubiquitination, leading to degradation. Furthermore, SAR1 overexpression rescues abiraterone-induced apoptosis and impairs redox homeostasis. In addition to steroid hormone synthesis, CYP17A1 associates with SAR1a/b to regulate protein processing and maintain ER health in glioblastomas.

CCAAT/enhancer-binding protein delta regulates the stemness of glioma stem-like cells through activating PDGFA expression upon inflammatory stimulation.

BACKGROUND: The small population of glioma stem-like cells (GSCs) contributes to tumor initiation, malignancy, and recurrence in glioblastoma. However, the maintenance of GSC properties in the tumor microenvironment remains unclear. In glioma, non-neoplastic cells create an inflammatory environment and subsequently mediate tumor progression and maintenance. Transcriptional factor CCAAT/enhancer-binding protein delta (CEBPD) is suggested to regulate various genes responsive to inflammatory cytokines, thus prompting us to investigate its role in regulating GSCs stemness after inflammatory stimulation. METHODS: Stemness properties were analyzed by using spheroid formation. Oncomine and TCGA bioinformatic databases were used to analyze gene expression. Western blotting, quantitative real-time polymerase chain reaction, luciferase reporter assay, and chromatin immunoprecipitation assay were used to analyze proteins and gene transcript levels. The glioma tissue microarrays were used for CEBPD and PDGFA expression by immunohistochemistry staining. RESULTS: We first found that IL-1ß promotes glioma spheroid formation and is associated with elevated CEBPD expression. Using microarray analysis, platelet-derived growth factor subunit A (PDGFA) was confirmed as a CEBPD-regulated gene that mediates IL-1ß-enhanced GSCs self-renewal. Further analysis of the genomic database and tissue array revealed that the expression levels between CEBPD and PDGFA were coincident in glioma patient samples. CONCLUSION: This is the first report showing the activation of PDGFA expression by CEBPD through IL-1ß treatment and a novel CEBPD function in maintaining the self-renewal feature of GSCs.

E2f1 regulates the induction of promyelocytic leukemia zinc finger transcription in neuronal differentiation of pluripotent P19 embryonal carcinoma cells.

During brain development, the expression of promyelocytic leukemia zinc finger (Plzf) in neural stem cells is precisely controlled to maintain the balance between neural stem cell self-renewal and differentiation. However, the mechanism underlying transcriptional regulation of Plzf in neural stem cell is still unclear. Herein, using P19 embryonal carcinoma cells as a model, we observed that Plzf expression was induced in the P19-derived embryonic bodies, which enrich neural stem-like cell populations, as demonstrated by the expression of neural stem cell markers, Nestin and Sox2. We then characterized the Plzf promoter and identified two E2f1 binding sites (-755/-751 and -53/-49, the transcription start site was designated as +1) are important for the activation of Plzf promoter. Finally, we found that the induction of Plzf in the neural stem-like cells derived from pluripotent P19 cells is decrease by E2f1 knockdown. Taken together, we conclude that E2f1 is an important transcription factor that regulates Plzf transcription and may involve in maintaining the self-renewal ability of neural stem cells.

Dehydroepiandrosterone Induces Temozolomide Resistance Through Modulating Phosphorylation and Acetylation of Sp1 in Glioblastoma.

Glioblastoma is the most malignant type of brain tumor for which there are currently no effective treatments. Patient prognosis is improved by radiation combined with temozolomide (TMZ) therapy but only for a short period of time due to the high prevalence of recurrence. Although O6-methylguanine-DNA methyltransferase (MGMT)-mediated DNA repair is a well-defined characteristic of TMZ resistance, the mechanism by which MGMT-deficient glioblastoma counteracts TMZ-induced DNA damage, leading to apoptosis, still remains unclear. Previously, we determined that aberrantly activated cytochrome P450 17A1 causes TMZ resistance in MGMT-deficient glioblastoma by increasing the secretion of dehydroepiandrosterone (DHEA), a neurosteroid that maintains the health of neurons and astrocytes. However, the precise mechanism by which DHEA alters the response of glioblastoma to TMZ has not been studied. In the present study, we found that DHEA prevents TMZ-induced apoptosis by attenuating DNA damage in MGMT-deficient glioblastoma. In addition, DHEA activated the LYN-AKT cascade to induce Sp1 phosphorylation. Phospho-Sp1 localized in TMZ-damaged DNA, prevented further DNA damage, and was deacetylated through the recruitment of HDAC1/2. Deacetylated Sp1 recruited proliferating cell nuclear antigen (PCNA) to attenuate DNA damage. To confirm whether the DHEA-induced cellular process contributes to TMZ resistance, we established a TMZ-resistant glioblastoma cell line, A172R, and isolated primary resistant tumor cells, PtR#1, from a glioblastoma patient exhibiting chemotherapeutic resistance. Sp1 exhibited phosphorylated and deacetylated status, and associated with HDAC1/2 and PCNA in TMZ-resistant cells. Based on these findings, we conclude that DHEA induces TMZ resistance in glioblastoma via the induction of phospho-Sp1-mediated DNA repair.

Highly sensitive/selective 3D nanostructured immunoparticle-based interface on a multichannel sensor array for detecting amyloid-beta in Alzheimer's disease.

Accumulation of ß-amyloid (Aß) peptides is highly associated with Alzheimer's disease (AD) progression in prevailing studies. The successful development of an ultrasensitive detection assay for Aß is a challenging task, especially from blood-based samples. Methods: We have developed a one-step electrophoresis/electropolymerization strategy for preparing a CSIP hierarchical immunoelectrochemical interface that is easily integrated into a PoCT device. The interface includes conductive silk fibroin-based immunoparticles (CSIPs) via electropolymerized Poly(3,4-ethylenedioxythiophene) (PEDOT) bridging to enable on-site electrochemical detection of serum amyloid-ß42 (Aß42) and -ß40 (Aß40) peptides from an AD blood test. In addition, micro-positron emission tomography (microPET) neuroimaging and behavioral tests were simultaneously performed. Results: This nanostructured conductive interface favors penetration of water-soluble biomolecules and catalyzes a redox reaction, providing limits of detection (LOD) of 6.63 pg/mL for Aß40 and 3.74 pg/mL for Aß42. Our proof-of-concept study confirms that the multi-sensing electrochemical immunosensor array (MEIA) platform enables simultaneous measurement of serum Aß42 and Aß40 peptide levels and is more informative in early stage AD animals than amyloid-labeling Aß plaque PET imaging and behavioral tests. Conclusion: We believe this study greatly expands the applications of silk fibroin-based materials, is an important contribution to the advancement of biomaterials, and would also be valuable in the design of new types of multichannel electrochemical immunosensor arrays for the detection of other diseases.

Astrocytic CCAAT/Enhancer-binding protein delta contributes to reactive oxygen species formation in neuroinflammation.

Excessive reactive oxygen species (ROS) can form an oxidative stress and an associated neuroinflammation. However, the contribution of astrocytes to ROS formation, the cause of the resistance of astrocytes to oxidative stress, and the consequences on neurons remain largely uninvestigated. The transcription factor CCAAT/enhancer-binding protein delta (CEBPD) is highly expressed in astrocytes and has been suggested to contribute to the progress of Alzheimer's disease (AD). In this study, we found that ROS formation and expression of p47phox and p67phox, subunits of NADPH oxidase, were increased in AppTg mice but attenuated in AppTg/Cebpd-/- mice. Cebpd can up-regulate p47phox and p67phox transcription via a direct binding on their promoters, which results in an increase in intracellular oxidative stress. In addition, Cebpd also up-regulated Cu/Zn superoxide dismutase (Sod1) in astrocytes. Inactivation of Sod1 increased the sensitization to oxidative stress, which provides a reason for the resistance of astrocytes in an oxidative stress environment. Taken together, the study first revealed and dissected the involvement of astrocytic Cebpd in the promotion of oxidative stress and the contribution of CEBPD to the resistance of astrocytes in an oxidative stress environment.

MDM2 Degrades Deacetylated Nucleolin Through Ubiquitination to Promote Glioma Stem-Like Cell Enrichment for Chemotherapeutic Resistance.

Glioblastoma multiforme (GBM) is the most fatal of all brain cancers, and the standard care protocol for GBM patients is surgical tumor resection followed by radiotherapy and temozolomide (TMZ)-mediated chemotherapy. However, tumor recurrence frequently occurs, and recurrent GBM exhibits more malignancy and less sensitivity in response to chemotherapy. The malignancy and drug resistance primarily reflect the small population of glioma stem-like cells (GSC). Therefore, understanding the mechanism that controls GSC enrichment is important to benefit the prognosis of GBM patients. Nucleolin (NCL), which is responsible for ribosome biogenesis and RNA maturation, is overexpressed in gliomas. However, the role of NCL in GSC development and drug resistance is still unclear. In this study, we demonstrate that NCL attenuated GSC enrichment to enhance the sensitivity of GBM cells in response to TMZ. In GSC enrichment, NCL was significantly reduced at the protein level as a result of decreased protein stability. In particular, the inhibition of HDAC activity by suberoylanilide hydroxamic acid rescued NCL acetylation accompanied by the loss of mouse double minute 2 homolog (MDM2)-mediated ubiquitination. In addition, we found that NCL ubiquitination resulted from the activation of STAT3- and JNK-mediated signaling in GSC. Moreover, NCL inhibited the formation of stem-like spheres by attenuating the expression of Sox2, Oct4, and Bmi1. Furthermore, NCL sensitized the response of GBM cells to TMZ. Based on these findings, NCL expression is a potential indicator to predict chemotherapeutic efficiency in GBM patients.

Bortezomib-induced miRNAs direct epigenetic silencing of locus genes and trigger apoptosis in leukemia.

MicroRNAs (miRNAs) have been suggested to repress transcription via binding the 3'-untranslated regions of mRNAs. However, the involvement and details of miRNA-mediated epigenetic regulation, particularly in targeting genomic DNA and mediating epigenetic regulation, remain largely uninvestigated. In the present study, transcription factor CCAAT/enhancer binding protein delta (CEBPD) was responsive to the anticancer drug bortezomib, a clinical and highly selective drug for leukemia treatment, and contributed to bortezomib-induced cell death. Interestingly, following the identification of CEBPD-induced miRNAs, we found that miR-744, miR-3154 and miR-3162 could target CpG islands in the 5'-flanking region of the CEBPD gene. We previously demonstrated that the Yin Yang 1 (YY1)/polycomb group (PcG) protein/DNA methyltransferase (DNMT) complex is important for CCAAT/enhancer binding protein delta (CEBPD) gene inactivation; we further found that Argonaute 2 (Ago2) interacts with YY1 and binds to the CEBPD promoter. The miRNA/Ago2/YY1/PcG group protein/DNMT complex linked the inactivation of CEBPD and genes adjacent to its 5'-flanking region, including protein kinase DNA-activated catalytic polypeptide (PRKDC), minichromosome maintenance-deficient 4 (MCM4) and ubiquitin-conjugating enzyme E2 variant 2 (UBE2V2), upon bortezomib treatment. Moreover, we revealed that miRNA binding is necessary for YY1/PcG group protein/DNMT complex-mediated epigenetic gene silencing and is associated with bortezomib-induced methylation on genomic DNA. The present study successfully characterized the interactions of the miRNA/Ago2/YY1/PcG group protein/DNMT complex and provided new insights for miRNA-mediated epigenetic regulation in bortezomib-induced leukemic cell arrest and cell death.