Glioma-derived cancer stem cells are hypersensitive to proteasomal inhibition.

Although proteasome inhibitors (PIs) are used as anticancer drugs to treat various cancers, their relative therapeutic efficacy on stem cells vs. bulk cancers remains unknown. Here, we show that stem cells derived from gliomas, GSCs, are up to 1,000-fold more sensitive to PIs (IC50, 27-70 nM) compared with their differentiated controls (IC50, 47 to ¼100 µM). The stemness of GSCs correlates to increased ubiquitination, whose misregulation readily triggers apoptosis. PI-induced apoptosis of GSCs is independent of NF-κB but involves the phosphorylation of c-Jun N-terminal kinase as well as the transcriptional activation of endoplasmic reticulum (ER) stress-associated proapoptotic mediators. In contrast to the general notion that ER stress-associated apoptosis is signaled by prolonged unfolded protein response (UPR), GSC-selective apoptosis is instead counteracted by the UPR ATF3 is a key mediator in GSC-selective apoptosis. Pharmaceutical uncoupling of the UPR from its downstream apoptosis sensitizes GSCs to PIs in vitro and during tumorigenesis in mice. Thus, a combinational treatment of a PI with an inhibitor of UPR-coupled apoptosis may enhance targeting of stem cells in gliomas.

Role of Cigarette Smoke-Induced Aggresome Formation in Chronic Obstructive Pulmonary Disease-Emphysema Pathogenesis.

Cigarette smoke (CS) exposure is known to induce proteostasis imbalance that can initiate accumulation of ubiquitinated proteins. Therefore, the primary goal of this study was to determine if first- and secondhand CS induces localization of ubiquitinated proteins in perinuclear spaces as aggresome bodies. Furthermore, we sought to determine the mechanism by which smoke-induced aggresome formation contributes to chronic obstructive pulmonary disease (COPD)-emphysema pathogenesis. Hence, Beas2b cells were treated with CS extract (CSE) for in vitro experimental analysis of CS-induced aggresome formation by immunoblotting, microscopy, and reporter assays, whereas chronic CS-exposed murine model and human COPD-emphysema lung tissues were used for validation. In preliminary analysis, we observed a significant (P < 0.01) increase in ubiquitinated protein aggregation in the insoluble protein fraction of CSE-treated Beas2b cells. We verified that CS-induced ubiquitin aggregrates are localized in the perinuclear spaces as aggresome bodies. These CS-induced aggresomes (P < 0.001) colocalize with autophagy protein microtubule-associated protein 1 light chain-3B(+) autophagy bodies, whereas U.S. Food and Drug Administration-approved autophagy-inducing drug (carbamazepine) significantly (P < 0.01) decreases their colocalization and expression, suggesting CS-impaired autophagy. Moreover, CSE treatment significantly increases valosin-containing protein-p62 protein-protein interaction (P < 0.0005) and p62 expression (aberrant autophagy marker; P < 0.0001), verifying CS-impaired autophagy as an aggresome formation mechanism. We also found that inhibiting protein synthesis by cycloheximide does not deplete CS-induced ubiquitinated protein aggregates, suggesting the role of CS-induced protein synthesis in aggresome formation. Next, we used an emphysema murine model to verify that chronic CS significantly (P < 0.0005) induces aggresome formation. Moreover, we observed that autophagy induction by carbamazepine inhibits CS-induced aggresome formation and alveolar space enlargement (P < 0.001), confirming involvement of aggresome bodies in COPD-emphysema pathogenesis. Finally, significantly higher p62 accumulation in smokers and severe COPD-emphysema lungs (Global Initiative for Chronic Obstructive Lung Disease Stage III/IV) as compared with normal nonsmokers (Global Initiative for Chronic Obstructive Lung Disease Stage 0) substantiates the pathogenic role of autophagy impairment in aggresome formation and COPD-emphysema progression. In conclusion, CS-induced aggresome formation is a novel mechanism involved in COPD-emphysema pathogenesis.

Sparse Graph Representation Learning based on Reinforcement Learning for Personalized Mild Cognitive Impairment (MCI) Diagnosis.

Resting-state functional magnetic resonance imaging (rs-fMRI) has gained attention as a reliable technique for investigating the intrinsic function patterns of the brain. It facilitates the extraction of functional connectivity networks (FCNs) that capture synchronized activity patterns among regions of interest (ROIs). Analyzing FCNs enables the identification of distinctive connectivity patterns associated with mild cognitive impairment (MCI). For MCI diagnosis, various sparse representation techniques have been introduced, including statistical- and deep learningbased methods. However, these methods face limitations due to their reliance on supervised learning schemes, which restrict the exploration necessary for probing novel solutions. To overcome such limitation, prior work has incorporated reinforcement learning (RL) to dynamically select ROIs, but effective exploration remains challenging due to the vast search space during training. To tackle this issue, in this study, we propose an advanced RL-based framework that utilizes a divide-and-conquer approach to decompose the FCN construction task into smaller subproblems in a subject-specific manner, enabling efficient exploration under each sub-problem condition. Additionally, we leverage the learned value function to determine the sparsity level of FCNs, considering individual characteristics of FCNs. We validate the effectiveness of our proposed framework by demonstrating its superior performance in MCI diagnosis on publicly available cohort datasets.

Graph-Based Conditional Generative Adversarial Networks for Major Depressive Disorder Diagnosis With Synthetic Functional Brain Network Generation.

Major Depressive Disorder (MDD) is a pervasive disorder affecting millions of individuals, presenting a significant global health concern. Functional connectivity (FC) derived from resting-state functional Magnetic Resonance Imaging (rs-fMRI) serves as a crucial tool in revealing functional connectivity patterns associated with MDD, playing an essential role in precise diagnosis. However, the limited data availability of FC poses challenges for robust MDD diagnosis. To tackle this, some studies have employed Deep Neural Networks (DNN) architectures to construct Generative Adversarial Networks (GAN) for synthetic FC generation, but this tends to overlook the inherent topology characteristics of FC. To overcome this challenge, we propose a novel Graph Convolutional Networks (GCN)-based Conditional GAN with Class-Aware Discriminator (GC-GAN). GC-GAN utilizes GCN in both the generator and discriminator to capture intricate FC patterns among brain regions, and the class-aware discriminator ensures the diversity and quality of the generated synthetic FC. Additionally, we introduce a topology refinement technique to enhance MDD diagnosis performance by optimizing the topology using the augmented FC dataset. Our framework was evaluated on publicly available rs-fMRI datasets, and the results demonstrate that GC-GAN outperforms existing methods. This indicates the superior potential of GCN in capturing intricate topology characteristics and generating high-fidelity synthetic FC, thus contributing to a more robust MDD diagnosis.

The Function of FK506-Binding Protein 13 in Protein Quality Control Protects Plasma Cells from Endoplasmic Reticulum Stress-Associated Apoptosis.

Plasma cells (PCs) are exposed to intense endoplasmic reticulum (ER) stress imposed by enormous rates of immunoglobulin (Ig) synthesis and secretion. Therefore, protein homeostasis is crucial for the survival of PCs, but its molecular mechanism remains largely unknown. Here, we found marked overexpression of FK506-binding protein 13 (FKBP13) in long-lived PCs from autoimmune mice and investigated its function using a plasmacytoma cell line secreting IgA. FKBP13 expression was induced largely in the lumen of ER in response to treatment with an ER stressor tunicamycin or overexpression of an adaptive unfolded protein response (UPR) protein X-box binding protein 1 (XBP1). Silencing of FKBP13 expression led to induction of molecules involved in the terminal UPR and ER stress-associated apoptosis. FKBP13 interacted with Ig, facilitated its ubiquitination, and lowered the extent of ER stress. FKBP13 overexpression caused a significant reduction in secreted IgA in plasmacytoma cells, and FKBP13 knockdown exerted an opposite effect. Rapamycin interfered with the interaction between FKBP13 and IgA and enhanced the amount of secreted IgA. Importantly, the level of FKBP13 was inversely correlated with the amount of secreted antibody in long-lived PCs from autoimmune mice. These results suggest that FKBP13 is a marker of long-lived PCs and a component of XBP1-dependent ER protein homeostasis. FKBP13 is likely to act as a molecular chaperone that delivers misfolded ER clients, including Ig, to ER-associated degradation, so reducing proteotoxic stress on the PC. Our data reveal a novel cytoprotective role for FKBP13 in long-lived PCs occurring at the expense of antibody production.

Second-hand cigarette smoke impairs bacterial phagocytosis in macrophages by modulating CFTR dependent lipid-rafts.

INTRODUCTION: First/Second-hand cigarette-smoke (FHS/SHS) exposure weakens immune defenses inducing chronic obstructive pulmonary disease (COPD) but the underlying mechanisms are not fully understood. Hence, we evaluated if SHS induced changes in membrane/lipid-raft (m-/r)-CFTR (cystic fibrosis transmembrane conductance regulator) expression/activity is a potential mechanism for impaired bacterial phagocytosis in COPD. METHODS: RAW264.7 murine macrophages were exposed to freshly prepared CS-extract (CSE) containing culture media and/or Pseudomonas-aeruginosa-PA01-GFP for phagocytosis (fluorescence-microscopy), bacterial survival (colony-forming-units-CFU), and immunoblotting assays. The CFTR-expression/activity and lipid-rafts were modulated by transient-transfection or inhibitors/inducers. Next, mice were exposed to acute/sub-chronic-SHS or room-air (5-days/3-weeks) and infected with PA01-GFP, followed by quantification of bacterial survival by CFU-assay. RESULTS: We investigated the effect of CSE treatment on RAW264.7 cells infected by PA01-GFP and observed that CSE treatment significantly (p<0.01) inhibits PA01-GFP phagocytosis as compared to the controls. We also verified this in murine model, exposed to acute/sub-chronic-SHS and found significant (p<0.05, p<0.02) increase in bacterial survival in the SHS-exposed lungs as compared to the room-air controls. Next, we examined the effect of impaired CFTR ion-channel-activity on PA01-GFP infection of RAW264.7 cells using CFTR172-inhibitor and found no significant change in phagocytosis. We also similarly evaluated the effect of a CFTR corrector-potentiator compound, VRT-532, and observed no significant rescue of CSE impaired PA01-GFP phagocytosis although it significantly (p<0.05) decreases CSE induced bacterial survival. Moreover, induction of CFTR expression in macrophages significantly (p<0.03) improves CSE impaired PA01-GFP phagocytosis as compared to the control. Next, we verified the link between m-/r-CFTR expression and phagocytosis using methyl-ß-cyclodextran (CD), as it is known to deplete CFTR from membrane lipid-rafts. We observed that CD treatment significantly (p<0.01) inhibits bacterial phagocytosis in RAW264.7 cells and adding CSE further impairs phagocytosis suggesting synergistic effect on CFTR dependent lipid-rafts. CONCLUSION: Our data suggest that SHS impairs bacterial phagocytosis by modulating CFTR dependent lipid-rafts.