Manganese is essential for several metabolic pathways but becomes toxic in excessive amounts. Manganese levels in the body are therefore tightly regulated, but the responsible protein(s) remain ...incompletely known. We studied two consanguineous families with neurologic disorders including juvenile-onset dystonia, adult-onset parkinsonism, severe hypermanganesemia, polycythemia, and chronic hepatic disease, including steatosis and cirrhosis. We localized the genetic defect by homozygosity mapping and then identified two different homozygous frameshift SLC30A10 mutations, segregating with disease. SLC30A10 is highly expressed in the liver and brain, including in the basal ganglia. Its encoded protein belongs to a large family of membrane transporters, mediating the efflux of divalent cations from the cytosol. We show the localization of SLC30A10 in normal human liver and nervous system, and its depletion in liver from one affected individual. Our in silico analyses suggest that SLC30A10 possesses substrate specificity different from its closest (zinc-transporting) homologs. We also show that the expression of SLC30A10 and the levels of the encoded protein are markedly induced by manganese in vitro. The phenotype associated with SLC30A10 mutations is broad, including neurologic, hepatic, and hematologic disturbances. Intrafamilial phenotypic variability is also present. Chelation therapy can normalize the manganesemia, leading to marked clinical improvements. In conclusion, we show that SLC30A10 mutations cause a treatable recessive disease with pleomorphic phenotype, and provide compelling evidence that SLC30A10 plays a pivotal role in manganese transport. This work has broad implications for understanding of the manganese biology and pathophysiology in multiple human organs.
Successful myelin repair in the adult CNS requires the robust and timely production of myelin proteins to generate new myelin sheaths. The underlying regulatory mechanisms and complex molecular basis ...of myelin regeneration, however, remain poorly understood. Here, we investigate the role of ERK MAP kinase signaling in this process. Conditional deletion of Erk2 from cells of the oligodendrocyte lineage resulted in delayed remyelination following demyelinating injury to the adult mouse corpus callosum. The delayed repair occurred as a result of a specific deficit in the translation of the major myelin protein, MBP. In the absence of ERK2, activation of the ribosomal protein S6 kinase (p70S6K) and its downstream target, ribosomal protein S6 (S6RP), was impaired at a critical time when premyelinating oligodendrocytes were transitioning to mature cells capable of generating new myelin sheaths. Thus, we have described an important link between the ERK MAP kinase signaling cascade and the translational machinery specifically in remyelinating oligodendrocytes in vivo. These results suggest an important role for ERK2 in the translational control of MBP, a myelin protein that appears critical for ensuring the timely generation of new myelin sheaths following demyelinating injury in the adult CNS.
Medulloblastoma (MB) is the most common brain malignancy in children, and is still responsible for significant mortality and morbidity. The aim of this study was to assess the safety and efficacy of ...Disulfiram (DSF), an FDA-approved inhibitor of Aldehyde-Dehydrogenase (ALDH), and Copper (Cu++) in human SSH-driven and Group 3 MB. The molecular mechanisms, effect on cancer-stem-cells (CSC) and DNA damage were investigated in xenograft models.
The cytotoxic and anti-CSC effects of DSF/Cu++ were evaluated with clonogenic assays, flow-cytometry, immunofluorescence, western-blotting. ONS76, UW228 (SHH-driven with Tp53m), D425med, D283 and D341 (Group 3) cell-lines were used. In vivo survival and nuclear protein localization protein-4 (NPL4), Ki67, Cleaved-Caspase-3, GFAP and NeuN expression were assessed in two Group 3 MB xenografts with immunohistochemistry and western-blotting.
Significant in vitro cytotoxicity was demonstrated at nanomolar concentrations. DSF/Cu++ induced cell-death through NPL4 accumulation in cell-nucleus and buildup of poly-ubiquitylated proteins. Flow-cytometry demonstrated a significant decrease in ALDH+, Nestin+ and CD133+ following treatment, anti-CSC effect was confirmed in vitro and in vivo. DSF/Cu++ prolonged survival, and increased nuclear NPL4 expression in vivo.
Our data suggest that this combination may serve as a novel treatment, as monotherapy or in combination with existing therapies, for aggressive subtypes of pediatric MB.
Data representation is of significant importance in minimizing multi-label ambiguity. While most researchers intensively investigate label correlation, the research on enhancing model robustness is ...preliminary. Low-quality data is one of the main reasons that model robustness degrades. Aiming at the cases with noisy features and missing labels, we develop a novel method called robust global and local label correlation (RGLC). In this model, subspace learning reconstructs intrinsic latent features immune from feature noise. The manifold learning ensures that outputs obtained by matrix factorization are similar in the low-rank latent label if the latent features are similar. We examine the co-occurrence of global and local label correlation with the constructed latent features and the latent labels. Extensive experiments demonstrate that the classification performance with integrated information is statistically superior over a collection of state-of-the-art approaches across numerous domains. Additionally, the proposed model shows promising performance on multi-label when noisy features and missing labels occur, demonstrating the robustness of multi-label classification.
Multi-label classification deals with the determination of instance-label associations for unseen instances. Although many margin-based approaches are delicately developed, the uncertainty ...classifications for those with smaller separation margins remain unsolved. The intuitionistic fuzzy set is an effective tool to characterize the concept of uncertainty, yet it has not been examined for multi-label cases. This paper proposed a novel model called intuitionistic fuzzy three-way label enhancement (IFTWLE) for multi-label classification. The IFTWLE combines label enhancement with an intuitionistic fuzzy set under the framework of three-way decisions. For unseen instances, we generated the pseudo-label for label uncertainty evaluation from a logical label-based model. An intuitionistic fuzzy set-based instance selection principle seamlessly bridges logical label learning and numerical label learning. The principle is hierarchically developed. At the label level, membership and non-membership functions are pair-wisely defined to measure the local uncertainty and generate candidate uncertain instances. After upgrading to the instance level, we select instances from the candidates for label enhancement, whereas they remained unchanged for the remaining. To the best of our knowledge, this is the first attempt to combine logical label learning with numerical label learning into a unified framework for minimizing classification uncertainty. Extensive experiments demonstrate that, with the selectively reconstructed label importance, IFTWLE achieves statistically superior over the state-of-the-art multi-label classification algorithms in terms of classification accuracy. The computational complexity of this algorithm is On2mk, where n, m, and k denote the unseen instances count, label count, and average label-specific feature size, respectively.
Olfactory neuroblastoma (ONB) is a rare malignant neoplasm arising in the upper portion of the sinonasal cavity. To better understand the genetic bases for ONB, here we perform whole exome and whole ...genome sequencing as well as single nucleotide polymorphism array analyses in a series of ONB patient samples. Deletions involving the dystrophin (DMD) locus are found in 12 of 14 (86%) tumors. Interestingly, one of the remaining tumors has a deletion in LAMA2, bringing the number of ONBs with deletions of genes involved in the development of muscular dystrophies to 13 or 93%. This high prevalence implicates an unexpected functional role for genes causing hereditary muscular dystrophies in ONB.
Clinical trials involving anti-programmed cell death protein-1 (anti-PD-1) failed to demonstrate improved overall survival in glioblastoma (GBM) patients. This may be due to the expression of ...alternative checkpoints such as B- and T- lymphocyte attenuator (BTLA) on several immune cell types including regulatory T cells. Murine GBM models indicate that there is significant upregulation of BTLA in the tumor microenvironment (TME) with associated T cell exhaustion. We investigate the use of antibodies against BTLA and PD-1 on reversing immunosuppression and increasing long-term survival in a murine GBM model. C57BL/6 J mice were implanted with the murine glioma cell line GL261 and randomized into 4 arms: (i) control, (ii) anti-PD-1, (iii) anti-BTLA, and (iv) anti-PD-1 + anti-BTLA. Kaplan-Meier curves were generated for all arms. Flow cytometric analysis of blood and brains were done on days 11 and 16 post-tumor implantation. Tumor-bearing mice treated with a combination of anti-PD-1 and anti-BTLA therapy experienced improved overall long-term survival (60%) compared to anti-PD-1 (20%) or anti-BTLA (0%) alone (P = .003). Compared to monotherapy with anti-PD-1, mice treated with combination therapy also demonstrated increased expression of CD4+ IFN-γ (P < .0001) and CD8+ IFN-γ (P = .0365), as well as decreased levels of CD4+ FoxP3+ regulatory T cells on day 16 in the brain (P = .0136). This is the first preclinical investigation into the effects of combination checkpoint blockade with anti-PD-1 and anti-BTLA treatment in GBM. We also show a direct effect on activated immune cell populations such as CD4+ and CD8 + T cells and immunosuppressive regulatory T cells through this combination therapy.
Recessive mutations in the F-box only protein 7 gene (FBXO7) cause PARK15, a mendelian form of early-onset, levodopa-responsive parkinsonism with severe loss of nigrostriatal dopaminergic neurons. ...However, the function of the protein encoded by FBXO7, and the pathogenesis of PARK15 remain unknown. No animal models of this disease exist. Here, we report the generation of a vertebrate model of PARK15 in zebrafish. We first show that the zebrafish Fbxo7 homolog protein (zFbxo7) is expressed abundantly in the normal zebrafish brain. Next, we used two zFbxo7-specific morpholinos (targeting protein translation and mRNA splicing, respectively), to knock down the zFbxo7 expression. The injection of either of these zFbxo7-specific morpholinos in the fish embryos induced a marked decrease in the zFbxo7 protein expression, and a range of developmental defects. Furthermore, whole-mount in situ mRNA hybridization showed abnormal patterning and significant decrease in the number of diencephalic tyrosine hydroxylase-expressing neurons, corresponding to the human nigrostriatal or ventral tegmental dopaminergic neurons. Of note, the number of the dopamine transporter-expressing neurons was much more severely depleted, suggesting dopaminergic dysfunctions earlier and larger than those due to neuronal loss. Last, the zFbxo7 morphants displayed severe locomotor disturbances (bradykinesia), which were dramatically improved by the dopaminergic agonist apomorphine. The severity of these morphological and behavioral abnormalities correlated with the severity of zFbxo7 protein deficiency. Moreover, the effects of the co-injection of zFbxo7- and p53-specific morpholinos were similar to those obtained with zFbxo7-specific morpholinos alone, supporting further the contention that the observed phenotypes were specifically due to the knock down of zFbxo7. In conclusion, this novel vertebrate model reproduces pathologic and behavioral hallmarks of human parkinsonism (dopaminergic neuronal loss and dopamine-dependent bradykinesia), representing therefore a valid tool for investigating the mechanisms of selective dopaminergic neuronal death, and screening for modifier genes and therapeutic compounds.
Atypical teratoid/rhabdoid tumors (AT/RT) are highly aggressive, malignant tumors and are the most common malignant brain tumor in children under 6 months of age. Currently, there is no standard ...treatment for AT/RT. Recent studies have reported potential anti-tumoral properties of ribavirin, a guanosine analog and anti-viral molecule approved by the Food and Drug Administration for treatment of hepatitis C. We previously demonstrated that ribavirin inhibited glioma cell growth
and
. Based on these results and the fact that no pre-clinical model of ribavirin in AT/RT exists, we decided to investigate the effect of ribavirin on several human AT/RT cell lines (BT12, BT16, and BT37) both
and
. We provide evidence that ribavirin has a significant impact on AT/RT cell growth and increases cell cycle arrest and cell death, potentially through modulation of the eIF4E and/or EZH2 pathways. Interestingly, using scratch wound and transwell Boyden chamber assays, we observed that ribavirin also impairs AT/RT cell migration, invasion, and adhesion. Finally, we demonstrate that ribavirin significantly improves the survival of mice orthotopically implanted with BT12 cells. Our work establishes that ribavirin is effective against AT/RT by decreasing tumoral cell growth and dissemination and could represent a new therapeutic option for children with this deadly disease.
Mutations in the F-box only protein 7 gene (FBXO7) cause PARK15, an autosomal recessive neurodegenerative disease presenting with severe levodopa-responsive parkinsonism and pyramidal disturbances. ...Understanding the PARK15 pathogenesis might thus provide clues on the mechanisms of maintenance of brain dopaminergic neurons, the same which are lost in Parkinson's disease. The protein(s) encoded by FBXO7 remain very poorly characterized. Here, we show that two protein isoforms are expressed from the FBXO7 gene in normal human cells. The isoform 1 is more abundant, particularly in primary skin fibroblasts. Both isoforms are undetectable in cell lines from the PARK15 patient of an Italian family; the isoform 1 is undetectable and the isoform 2 is severely decreased in the patients from a Dutch PARK15 family. In human cell lines and mouse primary neurons, the endogenous or over-expressed, wild type FBXO7 isoform 1 displays mostly a diffuse nuclear localization. An intact N-terminus is needed for the nuclear FBXO7 localization, as N-terminal modification by PARK15-linked missense mutation, or N-terminus tag leads to cytoplasmic mislocalization. Furthermore, the N-terminus of wild type FBXO7 (but not of mutant FBXO7) is able to confer nuclear localization to profilin (a cytoplasmic protein). Our data also suggest that overexpressed mutant FBXO7 proteins (T22M, R378G and R498X) have decreased stability compared to their wild type counterpart. In human brain, FBXO7 immunoreactivity was highest in the nuclei of neurons throughout the cerebral cortex, intermediate in the globus pallidum and the substantia nigra, and lowest in the hippocampus and cerebellum. In conclusion, the common cellular abnormality found in the PARK15 patients from the Dutch and Italian families is the depletion of the FBXO7 isoform 1, which normally localizes in the cell nucleus. The activity of FBXO7 in the nucleus appears therefore crucial for the maintenance of brain neurons and the pathogenesis of PARK15.
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