Publications by authors named "Eugene O Major"

102 Publications

Identification of circulating CD31CD45 cell populations with the potential to differentiate into erythroid cells.

Stem Cell Res Ther 2021 Apr 13;12(1):236. Epub 2021 Apr 13.

Department of Haematology, Laboratory of Stem Cells, Transfusion Medicine and Biotechnologies, Santo Spirito Hospital, Pescara, Italy.

Erythro-myeloid progenitors (EMP) are found in a population of cells expressing CD31 and CD45 markers (CD31CD45). A recent study indicated that EMPs persist until adulthood and can be a source of endothelial cells. We identified two sub-populations of EMP cells, CD31CD45 and CD31CD45, from peripheral blood that can differentiate into cells of erythroid lineage. Our novel findings add to the current knowledge of hematopoietic lineage commitment, and our sequential, dual-step, in vitro culture model provides a platform for the study of the molecular and cellular mechanisms underlying human hematopoiesis and erythroid differentiation.
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http://dx.doi.org/10.1186/s13287-021-02311-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8042691PMC
April 2021

Treating MS after surviving PML: Discrete strategies for rescue, remission, and recovery patient 2: From the National Multiple Sclerosis Society Case Conference Proceedings.

Neurol Neuroimmunol Neuroinflamm 2021 01 15;8(1). Epub 2020 Dec 15.

From the Department of Neurology (R.A.C., E. Melamed, T.C.V., E. Meltzer), Dell Medical School, University of Texas at Austin; Department of Ophthalmology (N.H.), University of Texas Southwestern, Dallas; Wellness Care Centers and Pediatric Rehabilitation (J.S.), Denton, TX; Ascension Seton Medical Center (M.S.), Austin, TX; National Institutes of Health (E.O.M.), Bethesda, MD; Departments of Neurology, and Biochemistry, Microbiology and Immunology (R.P.L.), Wayne State University, Detroit, MI; Colangelo College of Business (T.C.V.), Grand Canyon University, Phoenix, AZ; Department of Neurology (A.G.), University of Rochester, NY; Department of Computer Science (O.K.), Texas State University, San Marcos; Division of Microbiology and Immunology (M.S.P.), Yerkes National Primate Research Center, and Department of Pathology and Laboratory Medicine (M.S.P.), Emory University, Atlanta, GA; The National Multiple Sclerosis Society (K.C.), New York, NY; Department of Neurology (J.S.G.), University of California at San Diego; Department of Neurology (S.N.), Johns Hopkins Hospital, Bethesda, MD; Department of Neurology and Program in Immunology (S.S.Z.), University of California, San Francisco; andDepartments of Neurology, Ophthalmology & Neurosurgery (E.M.F., T.C.F.), Dell Medical School at the University of Texas at Austin.

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http://dx.doi.org/10.1212/NXI.0000000000000930DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7803334PMC
January 2021

Treating MS after surviving PML: Discrete strategies for rescue, remission, and recovery patient 1: From the National Multiple Sclerosis Society Case Conference Proceedings.

Neurol Neuroimmunol Neuroinflamm 2021 01 15;8(1). Epub 2020 Dec 15.

From the University of Rochester (N.A.), NY. N. Anadani is now with Department of Neurology, University of Oklahoma Health Science Center; Department of Neurology (M.H., A.D.G.), University of Rochester, NY; Department of Neurology (R.A.C., E.M., T.C.V.), Dell Medical School at the University of Texas at Austin; Department of Neurology (R.L.), Wayne State University, Detroit, MI; The National Multiple Sclerosis Society (K.C.), New York, NY; Laboratory of Molecular Medicine and Neuroscience (E.O.M.), Neurological Institute of Neurological Disorder and Stroke (Y.J.), Bethesda, MD. Y. Jassam is now with Department of Neurology, The University of Kansas Health System; Colangelo College of Business (T.C.V.), Grand Canyon University, Phoenix, AZ; Division of Microbiology and Immunology (M.S.P.), Yerkes National Primate Research Center, and Department of Pathology and Laboratory Medicine (M.S.P.), Emory University, Atlanta, GA; Department of Neurosciences (J.S.G.), University of California at San Diego; Department of Neurology (S.N.), Johns Hopkins University School of Medicine, Baltimore, MD; Department of Neurology and Program in Immunology (S.S.Z.), University of California San Francisco; and Department of Neurology, Neurosurgery, and Ophthalmology (E.M.F., T.C.F.), Dell Medical School at the University of Texas at Austin.

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http://dx.doi.org/10.1212/NXI.0000000000000929DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7803340PMC
January 2021

Intranasal anti-caspase-1 therapy preserves myelin and glucose metabolism in a model of progressive multiple sclerosis.

Glia 2021 Jan 3;69(1):216-229. Epub 2020 Sep 3.

Department of Medical Microbiology & Immunology, University of Alberta, Edmonton, Alberta, Canada.

Inflammatory demyelination and axonal injury in the central nervous system (CNS) are cardinal features of progressive multiple sclerosis (MS), and linked to activated brain macrophage-like cells (BMCs) including resident microglia and trafficking macrophages. Caspase-1 is a pivotal mediator of inflammation and cell death in the CNS. We investigated the effects of caspase-1 activation and its regulation in models of MS. Brains from progressive MS and non-MS patients, as well as cultured human oligodendrocytes were examined by transcriptomic and morphological methods. Next generation transcriptional sequencing of progressive MS compared to non-MS patients' normal appearing white matter (NAWM) showed induction of caspase-1 as well as other inflammasome-associated genes with concurrent suppression of neuron-specific genes. Oligodendrocytes exposed to TNFα exhibited upregulation of caspase-1 with myelin gene suppression in a cell differentiation state-dependent manner. Brains from cuprizone-exposed mice treated by intranasal delivery of the caspase-1 inhibitor, VX-765 or its vehicle, were investigated in morphological and molecular studies, as well as by fluorodeoxyglucose-positron emission tomography (FDG-PET) imaging. Cuprizone exposure resulted in BMC and caspase-1 activation accompanied by demyelination and axonal injury, which was abrogated by intranasal VX-765 treatment. FDG-PET imaging revealed suppressed glucose metabolism in the thalamus, hippocampus and cortex of cuprizone-exposed mice that was restored with VX-765 treatment. These studies highlight the caspase-1 dependent interactions between inflammation, demyelination, and glucose metabolism in progressive MS and associated models. Intranasal delivery of an anti-caspase-1 therapy represents a promising therapeutic approach for progressive MS and other neuro-inflammatory diseases.
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http://dx.doi.org/10.1002/glia.23896DOI Listing
January 2021

Productive HIV infection in astrocytes can be established via a nonclassical mechanism.

AIDS 2020 06;34(7):963-978

Section of Infections of the Nervous System.

Objective: Astrocytes are proposed to be a critical reservoir of HIV in the brain. However, HIV infection of astrocytes is inefficient in vitro except for cell-to-cell transmission from HIV-infected cells. Here, we explore mechanisms by which cell-free HIV bypasses entry and postentry barriers leading to a productive infection.

Methods: HIV infection of astrocytes was investigated by a variety of techniques including transfection of CD4-expressing plasmid, treatment with lysosomotropic agents or using a transwell culture system loaded with HIV-infected lymphocytes. Infection was monitored by HIV-1 p24 in culture supernatants and integrated proviral DNA was quantified by Alu-PCR.

Results: Persistent HIV infection could be established in astrocytes by transfection of proviral DNA, transduction with VSV-G-pseudotyped viruses, transient expression of CD4 followed by HIV infection, or simultaneous treatment with lysosomotropic chloroquine or Tat-HA2 peptide with HIV infection. In absence of these treatments, HIV entered via endocytosis as seen by electronmicroscopy and underwent lysosomal degradation without proviral integration, indicating endocytosis is a dead end for HIV in astrocytes. Nevertheless, productive infection was observed when astrocytes were in close proximity but physically separated from HIV-infected lymphocytes in the transwell cultures. This occurred with X4 or dual tropic R5X4 viruses and was blocked by an antibody or antagonist to CXCR4.

Conclusion: A CD4-independent, CXCR4-dependent mechanism of viral entry is proposed, by which immature HIV particles from infected lymphocytes might directly bind to CXCR4 on astrocytes and trigger virus--cell fusion during or after the process of viral maturation. This mechanism may contribute to the formation of brain HIV reservoirs.
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http://dx.doi.org/10.1097/QAD.0000000000002512DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7429268PMC
June 2020

Germline Genetic Risk Variants for Progressive Multifocal Leukoencephalopathy.

Front Neurol 2020 17;11:186. Epub 2020 Mar 17.

Population Bio UK, Inc., Oxfordshire, United Kingdom.

Progressive multifocal leukoencephalopathy (PML) is a rare demyelinating disorder of the brain caused by reactivation of the JC virus (JCV), a polyomavirus that infects at least 60% of the population but is asymptomatic or results in benign symptoms in most people. PML occurs as a secondary disease in a variety of disorders or as a serious adverse event from immunosuppressant agents, but is mainly found in three groups: HIV-infected patients, patients with hematological malignancies, or multiple sclerosis (MS) patients on the immunosuppressant therapy natalizumab. It is severely debilitating and is deadly in ~50% HIV cases, ~90% of hematological malignancy cases, and ~24% of MS-natalizumab cases. A PML risk prediction test would have clinical utility in all at risk patient groups but would be particularly beneficial in patients considering therapy with immunosuppressant agents known to cause PML, such as natalizumab, rituximab, and others. While a JC antibody test is currently used in the clinical decision process for natalizumab, it is suboptimal because of its low specificity and requirement to periodically retest patients for seroconversion or to assess if a patient's JCV index has increased. Whereas a high specificity genetic risk prediction test comprising host genetic risk variants (i.e., germline variants occurring at higher frequency in PML patients compared to the general population) could be administered one time to provide clinicians with additional risk prediction information that is independent of JCV serostatus. Prior PML case reports support the hypothesis that PML risk is greater in patients with a genetically caused immunodeficiency disorder. To identify germline PML risk variants, we performed exome sequencing on 185 PML cases (70 in a discovery cohort and 115 in a replication cohort) and used the gnomAD variant database for interpretation. Our study yielded 19 rare variants (maximum allele frequency of 0.02 in gnomAD ethnically matched populations) that impact 17 immune function genes (10 are known to cause inborn errors of immunity). Modeling of these variants in a PML genetic risk test for MS patients considering natalizumab treatment indicates that at least a quarter of PML cases may be preventable.
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http://dx.doi.org/10.3389/fneur.2020.00186DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7094807PMC
March 2020

Perspectives from the Laboratory at the National Institute of Neurological Disorders and Stroke Assessing JC Virus DNA in Clinical Samples as It Ends Its Operation.

Authors:
Eugene O Major

Ann Neurol 2019 10 27;86(4):477-479. Epub 2019 Aug 27.

CLIA Laboratory of Molecular Medicine and Neuroscience, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD.

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http://dx.doi.org/10.1002/ana.25553DOI Listing
October 2019

Pembrolizumab Treatment for Progressive Multifocal Leukoencephalopathy.

N Engl J Med 2019 04 10;380(17):1597-1605. Epub 2019 Apr 10.

From the Neuroimmunology Clinic (I.C., J.O.), the Viral Immunology Section (Y.E.-A., S.J.), the Section of Infections of the Nervous System (B.S., L.B.R., A.N.), the Laboratory of Molecular Medicine and Neuroscience (M.M., C.R., E.O.M.), and the Translational Neuroradiology Section (S.-K.H., M.K.S., E.B., D.S.R.), National Institute of Neurological Disorders and Stroke, and the Hematology Branch, National Heart, Lung, and Blood Institute (P.M.), National Institutes of Health, Bethesda, MD; and the Herbert Irving Comprehensive Cancer Center, Columbia University Medical Center, New York (P.M.).

Background: Progressive multifocal leukoencephalopathy (PML) is an opportunistic brain infection that is caused by the JC virus and is typically fatal unless immune function can be restored. Programmed cell death protein 1 (PD-1) is a negative regulator of the immune response that may contribute to impaired viral clearance. Whether PD-1 blockade with pembrolizumab could reinvigorate anti-JC virus immune activity in patients with PML was unknown.

Methods: We administered pembrolizumab at a dose of 2 mg per kilogram of body weight every 4 to 6 weeks to eight adults with PML, each with a different underlying predisposing condition. Each patient received at least one dose but no more than three doses.

Results: Pembrolizumab induced down-regulation of PD-1 expression on lymphocytes in peripheral blood and in cerebrospinal fluid (CSF) in all eight patients. Five patients had clinical improvement or stabilization of PML accompanied by a reduction in the JC viral load in the CSF and an increase in in vitro CD4+ and CD8+ anti-JC virus activity. In the other three patients, no meaningful change was observed in the viral load or in the magnitude of antiviral cellular immune response, and there was no clinical improvement.

Conclusions: Our findings are consistent with the hypothesis that in some patients with PML, pembrolizumab reduces JC viral load and increases CD4+ and CD8+ activity against the JC virus; clinical improvement or stabilization occurred in five of the eight patients who received pembrolizumab. Further study of immune checkpoint inhibitors in the treatment of PML is warranted. (Funded by the National Institutes of Health.).
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http://dx.doi.org/10.1056/NEJMoa1815039DOI Listing
April 2019

Activated T cells induce proliferation of oligodendrocyte progenitor cells via release of vascular endothelial cell growth factor-A.

Glia 2018 11;66(11):2503-2513

Translational Neuroscience Center, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland.

Neuroinflammatory diseases such as multiple sclerosis are characterized by infiltration of lymphocytes into the central nervous system followed by demyelination and axonal degeneration. While evidence suggests that activated T lymphocytes induce neurotoxicity and impair function of neural stem cells, the effect of T cells on oligodendrocyte progenitor cells (OPCs) is still uncertain, partly due to the difficulty in obtaining human OPCs. Here we studied the effect of activated T cells on OPCs using OPCs derived from human hematopoietic stem cells or from human fetal brain. OPCs were exposed to supernatants (sups) from activated T cells. Cell proliferation was determined by EdU incorporation and CellQuanti-Blue assays. Surprisingly, we found that sups from activated T cells induced OPC proliferation by regulating cell cycle progression. Vascular endothelial growth factor A (VEGF-A) transcripts were increased in T cells after activation. Immunodepletion of VEGF-A from activated T cell sups significantly attenuated its effect on OPC proliferation. Furthermore, VEGF receptor 2 (VEGFR2) was expressed on OPCs and its inhibition also attenuated activated T cell-induced OPC proliferation. Thus, activated T cells have a trophic role by promoting OPC proliferation via the VEGFR2 pathway.
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http://dx.doi.org/10.1002/glia.23501DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6278606PMC
November 2018

Caspase-1 inhibition prevents glial inflammasome activation and pyroptosis in models of multiple sclerosis.

Proc Natl Acad Sci U S A 2018 06 12;115(26):E6065-E6074. Epub 2018 Jun 12.

Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, AB T6G 2R3, Canada;

Multiple sclerosis (MS) is a progressive inflammatory demyelinating disease of the CNS of unknown cause that remains incurable. Inflammasome-associated caspases mediate the maturation and release of the proinflammatory cytokines IL-1β and IL-18 and activate the pore-forming protein gasdermin D (GSDMD). Inflammatory programmed cell death, pyroptosis, was recently shown to be mediated by GSDMD. Here, we report molecular evidence for GSDMD-mediated inflammasome activation and pyroptosis in both myeloid cells (macrophages/microglia) and, unexpectedly, in myelin-forming oligodendrocytes (ODCs) in the CNS of patients with MS and in the MS animal model, experimental autoimmune encephalomyelitis (EAE). We observed inflammasome activation and pyroptosis in human microglia and ODCs in vitro after exposure to inflammatory stimuli and demonstrate caspase-1 inhibition by the small-molecule inhibitor VX-765 in both cell types. GSDMD inhibition by siRNA transduction suppressed pyroptosis in human microglia. VX-765 treatment of EAE animals reduced the expression of inflammasome- and pyroptosis-associated proteins in the CNS, prevented axonal injury, and improved neurobehavioral performance. Thus, GSDMD-mediated pyroptosis in select glia cells is a previously unrecognized mechanism of inflammatory demyelination and represents a unique therapeutic opportunity for mitigating the disease process in MS and other CNS inflammatory diseases.
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http://dx.doi.org/10.1073/pnas.1722041115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6042136PMC
June 2018

Pathogenesis of progressive multifocal leukoencephalopathy and risks associated with treatments for multiple sclerosis: a decade of lessons learned.

Lancet Neurol 2018 05;17(5):467-480

Department of Neurology, Washington University School of Medicine, St Louis, MO, USA. Electronic address:

Progressive multifocal leukoencephalopathy (PML) is a rare, devastating demyelinating disease of the CNS caused by the JC virus (JCV) that occurs in patients with compromised immune systems. Detection of PML in systemically immunocompetent patients with multiple sclerosis treated with natalizumab points to a role for this drug in the pathophysiology of PML. Emerging knowledge of the cellular and molecular biology of JCV infection and the pathogenesis of PML-including interplay of this common virus with the human immune system and features of natalizumab that might contribute to PML pathogenesis-provides new opportunities to monitor viral status and predict risk of JCV-associated disease. In the absence of an effective treatment for PML, early detection of the disease in patients with multiple sclerosis who are receiving natalizumab or other immunomodulatory treatments is vital to minimize CNS injury and avoid severe disability. Frequent MRI, stratified along a clinical and virus-specific immune risk profile, can be used to detect presymptomatic PML. Improved approaches to PML risk stratification are needed to guide treatment choices and surveillance of patients with multiple sclerosis.
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http://dx.doi.org/10.1016/S1474-4422(18)30040-1DOI Listing
May 2018

Progressive Multifocal Leukoencephalopathy Lesions and JC Virus: The Limits and Value of Imaging.

Authors:
Eugene O Major

JAMA Neurol 2018 07;75(7):789-790

Molecular Medicine and Neuroscience, Division of Neuroimmunology and Neurovirology, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland.

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http://dx.doi.org/10.1001/jamaneurol.2018.0004DOI Listing
July 2018

The emergence of neuroepidemiology, neurovirology and neuroimmunology: the legacies of John F. Kurtzke and Richard 'Dick' T. Johnson.

J Neurol 2017 Apr 12;264(4):817-828. Epub 2016 Oct 12.

Departments of Neurology and Neurotherapeutics, University of Texas Southwestern School of Medicine, 5323 Harry Hines Blvd., Dallas, TX, 75235, USA.

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http://dx.doi.org/10.1007/s00415-016-8293-yDOI Listing
April 2017

A decade of natalizumab and PML: Has there been a tacit transfer of risk acceptance?

Mult Scler 2017 Jun 27;23(7):934-936. Epub 2016 Sep 27.

Division of NeuroImmunology and NeuroVirology, NINDS, NIH, Bethesda, MD, USA.

The interplay between each of the stakeholder's responsibilities and desires clearly has resulted in continued widespread use of natalizumab with substantial risks and an ongoing quest for better risk mitigation. In the United States, regulatory actions codified the process of risk acceptance-and risk transfer-by escalating monitoring and information transfer to physicians and patients. Management of medication-related risks is a core function of regulatory agencies such as the Food and Drug Administration (FDA), European Medicines Agency (EMA), and the medical community. The interaction among stakeholders in medicine, pharma, regulatory bodies, physicians, and patients, sometimes has changed without overt review and discussion. Such is the case for natalizumab, an important and widely used disease-modifying therapy for multiple sclerosis. A rather silent but very considerable shift, effectively transferring increased risk for progressive multifocal leukoencephalopathy (PML) to the physicians and patients, has occurred in the past decade. We believe this changed risk should be clearly recognized and considered by all the stakeholders.
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http://dx.doi.org/10.1177/1352458516670735DOI Listing
June 2017

JC virus granule cell neuronopathy in the setting of chronic lymphopenia treated with recombinant interleukin-7.

J Neurovirol 2017 02 15;23(1):141-146. Epub 2016 Jul 15.

Department of Neurology, Washington University School of Medicine, Campus Box 8111. 660 South Euclid Ave., St. Louis, MO, 63110, USA.

JC virus (JCV) is a human polyomavirus that infects the central nervous system (CNS) of immunocompromised patients. JCV granule cell neuronopathy (JCV-GCN) is caused by infection of cerebellar granule cells, causing ataxia. A 77-year-old man with iatrogenic lymphopenia presented with severe ataxia and was diagnosed with JCV-GCN. His ataxia and cerebrospinal fluid (CSF) improved with intravenous immunoglobulin, high-dose intravenous methylprednisolone, mirtazapine, and mefloquine. Interleukin-7 (IL-7) therapy reconstituted his lymphocytes and reduced his CSF JCV load. One month after IL-7 therapy, he developed worsening ataxia and CSF inflammation, which raised suspicion for immune reconstitution inflammatory syndrome. Steroids were restarted and his ataxia stabilized.
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http://dx.doi.org/10.1007/s13365-016-0465-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5588866PMC
February 2017

BK virus encephalopathy and sclerosing vasculopathy in a patient with hypohidrotic ectodermal dysplasia and immunodeficiency.

Acta Neuropathol Commun 2016 07 13;4(1):73. Epub 2016 Jul 13.

Department of Pathology, Icahn School of Medicine at Mount Sinai, New York, 10029, NY, USA.

Human BK polyomavirus (BKV) is reactivated under conditions of immunosuppression leading most commonly to nephropathy or cystitis; its tropism for the brain is rare and poorly understood. We present a unique case of BKV-associated encephalopathy in a man with hypohidrotic ectodermal dysplasia and immunodeficiency (HED-ID) due to IKK-gamma (NEMO) mutation, who developed progressive neurological symptoms. Brain biopsy demonstrated polyomavirus infection of gray and white matter, with predominant involvement of cortex and distinct neuronal tropism, in addition to limited demyelination and oligodendroglial inclusions. Immunohistochemistry demonstrated polyoma T-antigen in neurons and glia, but expression of VP1 capsid protein only in glia. PCR analysis on both brain biopsy tissue and cerebrospinal fluid detected high levels of BKV DNA. Sequencing studies further identified novel BKV variant and disclosed unique rearrangements in the noncoding control region of the viral DNA (BKVN NCCR). Neuropathological analysis also demonstrated an unusual form of obliterative fibrosing vasculopathy in the subcortical white matter with abnormal lysosomal accumulations, possibly related to the patient's underlying ectodermal dysplasia. Our report provides the first neuropathological description of HED-ID due to NEMO mutation, and expands the diversity of neurological presentations of BKV infection in brain, underscoring the importance of its consideration in immunodeficient patients with unexplained encephalopathy. We also document novel BKVN NCCR rearrangements that may be associated with the unique neuronal tropism in this patient.
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http://dx.doi.org/10.1186/s40478-016-0342-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4944483PMC
July 2016

A link between long-term natalizumab dosing in MS and PML: Putting the puzzle together.

Neurol Neuroimmunol Neuroinflamm 2016 Jun 29;3(3):e235. Epub 2016 Apr 29.

Division of Neuroimmunology and Neurovirology, Intramural Research Program, National Institute of Neurological Disorders and Stroke, NIH, Bethesda, MD.

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http://dx.doi.org/10.1212/NXI.0000000000000235DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4853054PMC
June 2016

Progressive Multifocal Leukoencephalopathy in Primary Immune Deficiencies: Stat1 Gain of Function and Review of the Literature.

Clin Infect Dis 2016 Apr 6;62(8):986-94. Epub 2016 Jan 6.

Laboratory of Clinical Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland.

Background: Progressive multifocal leukoencephalopathy (PML) is a rare, severe, otherwise fatal viral infection of the white matter of the brain caused by the polyomavirus JC virus, which typically occurs only in immunocompromised patients. One patient with dominant gain-of-function (GOF) mutation in signal transducer and activator of transcription 1 (STAT1) with chronic mucocutaneous candidiasis and PML was reported previously. We aim to identify the molecular defect in 3 patients with PML and to review the literature on PML in primary immune defects (PIDs).

Methods: STAT1 was sequenced in 3 patients with PML. U3C cell lines were transfected with STAT1 and assays to search for STAT1 phosphorylation, transcriptional response, and target gene expression were performed.

Results: We identified 3 new unrelated cases of PML in patients with GOF STAT1 mutations, including the novel STAT1 mutation, L400Q. These STAT1 mutations caused delayed STAT1 dephosphorylation and enhanced interferon-gamma-driven responses. In our review of the literature regarding PML in primary immune deficiencies we found 26 cases, only 54% of which were molecularly characterized, the remainder being syndromically diagnosed only.

Conclusions: The occurrence of PML in 4 cases of STAT1 GOF suggests that STAT1 plays a critical role in the control of JC virus in the central nervous system.
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http://dx.doi.org/10.1093/cid/civ1220DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4803104PMC
April 2016

Cell-to-cell contact facilitates HIV transmission from lymphocytes to astrocytes via CXCR4.

AIDS 2015 Apr;29(7):755-66

aSection of Infections of the Nervous System, National Institute of Neurological Diseases and Stroke bLaboratory of Cell Biology, Center for Cancer Research, National Cancer Institute cLaboratory of Molecular Medicine and Neuroscience, National Institute of Neurological Diseases and Stroke, National Institutes of Health, Bethesda, Maryland, USA.

Objectives: HIV reservoir in the brain represents a major barrier for curing HIV infection. As the most abundant, long-lived cell type, astrocytes play a critical role in maintaining the reservoir; however, the mechanism of infection remains unknown. Here, we determine how viral transmission occurs from HIV-infected lymphocytes to astrocytes by cell-to-cell contact.

Design And Methods: Human astrocytes were exposed to HIV-infected lymphocytes and monitored by live-imaging, confocal microscopy, transmission and three-dimensional electron microscopy. A panel of receptor antagonists was used to determine the mechanism of viral entry.

Results: We found that cell-to-cell contact resulted in efficient transmission of X4 or X4R5-using viruses from T lymphocytes to astrocytes. In co-cultures of astrocytes with HIV-infected lymphocytes, the interaction occurred through a dynamic process of attachment and detachment of the two cell types. Infected lymphocytes invaginated into astrocytes or the contacts occurred via filopodial extensions from either cell type, leading to the formation of virological synapses. In the synapses, budding of immature or incomplete HIV particles from lymphocytes occurred directly onto the membranes of astrocytes. This cell-to-cell transmission could be almost completely blocked by anti-CXCR4 antibody and its antagonist, but only partially inhibited by anti-CD4, ICAM1 antibodies.

Conclusion: Cell-to-cell transmission was mediated by a unique mechanism by which immature viral particles initiated a fusion process in a CXCR4-dependent, CD4-independent manner. These observations have important implications for developing approaches to prevent formation of HIV reservoirs in the brain.
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http://dx.doi.org/10.1097/QAD.0000000000000605DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4438861PMC
April 2015

Immune System Involvement in the Pathogenesis of JC Virus Induced PML: What is Learned from Studies of Patients with Underlying Diseases and Therapies as Risk Factors.

Front Immunol 2015 28;6:159. Epub 2015 Apr 28.

Laboratory of Molecular Medicine and Neuroscience, National Institute of Neurological Disorders and Stroke, National Institutes of Health , Bethesda, MD , USA.

The human polyomavirus JC PyV lytic infection of oligodendrocytes in the human brain results in the demyelinating disease progressive multifocal leukoencephalopathy, PML. JCV is a common virus infection in the population that leads to PML in patients with underlying diseases and therapies that cause immune deficiencies or modulate immune system functions. Patients may have high levels of antibody to JCV that neither protect them from PML nor clear the infection once PML is established. Cell-mediated immunity plays a more effective role in clearing initial or reactivated JCV infection before PML occurs. However, patients with underlying diseases and therapies for treatment are at high risk for PML. MS patients on natalizumab are one of the categories with the highest incidence of PML. Natalizumab is a humanized monoclonal antibody targeting α4 integrins that prevents inflammatory cells from entering the brain and it has been used as a treatment for MS. A number of studies have investigated the occurrence of PML in these patients and their cell-mediated immune profile that might gain insight into the mechanism that ties natalizumab with a high risk of developing PML. It seems that cells of the immune system participate in the pathogenesis of PML as well as clearance of JCV infection.
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http://dx.doi.org/10.3389/fimmu.2015.00159DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4412132PMC
May 2015

Direct induction of human neural stem cells from peripheral blood hematopoietic progenitor cells.

J Vis Exp 2015 Jan 28(95):52298. Epub 2015 Jan 28.

Translational Neuroscience Center, National Institute of Neurological Disorders and Stroke, National Institutes of Health.

Human disease specific neuronal cultures are essential for generating in vitro models for human neurological diseases. However, the lack of access to primary human adult neural cultures raises unique challenges. Recent developments in induced pluripotent stem cells (iPSC) provides an alternative approach to derive neural cultures from skin fibroblasts through patient specific iPSC, but this process is labor intensive, requires special expertise and large amounts of resources, and can take several months. This prevents the wide application of this technology to the study of neurological diseases. To overcome some of these issues, we have developed a method to derive neural stem cells directly from human adult peripheral blood, bypassing the iPSC derivation process. Hematopoietic progenitor cells enriched from human adult peripheral blood were cultured in vitro and transfected with Sendai virus vectors containing transcriptional factors Sox2, Oct3/4, Klf4, and c-Myc. The transfection results in morphological changes in the cells which are further selected by using human neural progenitor medium containing basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF). The resulting cells are characterized by the expression for neural stem cell markers, such as nestin and SOX2. These neural stem cells could be further differentiated to neurons, astroglia and oligodendrocytes in specified differentiation media. Using easily accessible human peripheral blood samples, this method could be used to derive neural stem cells for further differentiation to neural cells for in vitro modeling of neurological disorders and may advance studies related to the pathogenesis and treatment of those diseases.
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http://dx.doi.org/10.3791/52298DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4354569PMC
January 2015

JC virus quasispecies analysis reveals a complex viral population underlying progressive multifocal leukoencephalopathy and supports viral dissemination via the hematogenous route.

J Virol 2015 Jan 12;89(2):1340-7. Epub 2014 Nov 12.

Janssen Diagnostics, Beerse, Belgium

Unlabelled: Opportunistic infection of oligodendrocytes by human JC polyomavirus may result in the development of progressive multifocal encephalopathy in immunocompromised individuals. Neurotropic JC virus generally harbors reorganized noncoding control region (NCCR) DNA interspersed on the viral genome between early and late coding genes. By applying 454 sequencing on NCCR DNA amplified from body fluid samples (urine, plasma, and cerebrospinal fluid [CSF]) from 19 progressive multifocal leukoencephalopathy (PML) patients, we attempted to reveal the composition of the JC polyomavirus population (the quasispecies, i.e., the whole of the consensus population and minor viral variants) contained in different body compartments and to better understand intrapatient viral dissemination. Our data demonstrate that in the CSF of PML patients, the JC viral population is often a complex mixture composed of multiple viral variants that contribute to the quasispecies. In contrast, urinary JC virus highly resembled the archetype virus, and urine most often did not contain minor viral variants. It also appeared that archetype JC virus could sporadically be identified in PML patient brain, although selection of rearranged JC virus DNA was favored. Comparison of the quasispecies from different body compartments within a given patient suggested a strong correlation between the viral population in plasma and CSF, whereas the viral population shed in urine appeared to be unrelated. In conclusion, it is shown that the representation of viral DNA in the CSF following the high-level DNA replication in the brain underlying PML has hitherto been much underestimated. Our data also underscore that the hematogenous route might play a pivotal role in viral dissemination from or toward the brain.

Importance: For the first time, the JC polyomavirus population contained in different body compartments of patients diagnosed with progressive multifocal encephalopathy has been studied by deep sequencing. Two main findings came out of this work. First, it became apparent that the complexity of the viral population associated with PML has been highly underestimated so far, suggestive of a highly dynamic process of reorganization of the noncoding control region of JC polyomavirus in vivo, mainly in CSF and blood. Second, evidence showing viral dissemination from and/or toward the brain via the hematogenous route was provided, confirming a hypothesis that was recently put forward in the field.
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http://dx.doi.org/10.1128/JVI.02565-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4300648PMC
January 2015

Contamination of SVG p12 cells with BK polyomavirus occurred after deposit in the American Type Culture Collection.

J Virol 2014 Nov;88(21):12928-9

Laboratory of Molecular Medicine and Neuroscience, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland

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http://dx.doi.org/10.1128/JVI.01600-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4248886PMC
November 2014

Relevance of CD34+ cells as a reservoir for JC virus in patients with multiple sclerosis--reply.

JAMA Neurol 2014 Sep;71(9):1192-3

National Institutes of Health, Bethesda, Maryland.

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http://dx.doi.org/10.1001/jamaneurol.2014.1858DOI Listing
September 2014

JC virus in CD34+ and CD19+ cells in patients with multiple sclerosis treated with natalizumab.

JAMA Neurol 2014 May;71(5):596-602

Laboratory of Molecular Medicine and Neuroscience, National Institutes of Health (NIH), Bethesda, Maryland.

Importance: Infection with JC virus (JCV) may lead to development of demyelinating progressive multifocal leukoencephalopathy in patients with multiple sclerosis (MS) who are treated with natalizumab.

Objective: To determine whether mononuclear cells in circulation from MS patients treated with natalizumab harbor JCV DNA.

Design, Setting, And Participants: In this prospective investigation, we enrolled 49 MS patients from the Clinical Center for Multiple Sclerosis at The University of Texas Southwestern Medical Center and 18 healthy volunteers. We drew 120-mL blood samples from 26 MS patients at baseline and at approximately 3-month intervals to 10 months during the course of natalizumab infusions. One blood sample was drawn from 23 MS patients receiving natalizumab for more than 24 months and from 18 healthy volunteers.

Interventions: Natalizumab treatment of MS.

Main Outcomes And Measures: The blood samples were separated using flow cytometry into CD34+, CD19+, and CD3+ cell subsets; DNA templates were prepared using quantitative polymerase chain reaction for JCV DNA identification. Plasma samples were tested for anti-JCV antibodies by enzyme-linked immunosorbent assays performed at the Laboratory of Molecular Medicine and Neuroscience, National Institute of Neurological and Communicative Diseases and Stroke.

Results: Thirteen of the 26 patients (50%) with baseline and follow-up blood samples had detectable viral DNA in at least 1 cell compartment at 1 or more points. Ten of the 23 patients (44%) receiving treatment for more than 24 months and 3 of the 18 healthy volunteers (17%) also had detectable viral DNA in 1 or more cell compartment. Fifteen of the 49 MS patients (31%) were confirmed to harbor JCV in CD34+ cells and 12 of 49 (24%) in CD19+ cells. Only 1 of 18 healthy volunteers were viremic in CD34+ cells and none in CD19+ cells. Nine patients and 1 healthy volunteer were viremic but had seronegative test results for JCV antibodies.

Conclusions And Relevance: JC virus DNA was detectable within cell compartments of natalizumab-treated MS patients after treatment inception and longer. JC virus DNA may harbor in CD34+ cells in bone marrow that mobilize into the peripheral circulation at high concentrations. Latently infected cells initiate differentiation to CD19+ cells that favors growth of JCV. These data link the mechanism of natalizumab treatment with progressive multifocal leukoencephalopathy.
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http://dx.doi.org/10.1001/jamaneurol.2014.63DOI Listing
May 2014

Cerebellar manifestation of PML under fumarate and after efalizumab treatment of psoriasis.

J Neurol 2014 May 18;261(5):1021-4. Epub 2014 Mar 18.

Department of Neurology, University of Leipzig, Liebigstraße 20, 04103, Leipzig, Germany.

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http://dx.doi.org/10.1007/s00415-014-7311-1DOI Listing
May 2014

Lymphocyte gene expression and JC virus noncoding control region sequences are linked with the risk of progressive multifocal leukoencephalopathy.

J Virol 2014 May 19;88(9):5177-83. Epub 2014 Feb 19.

Laboratory of Molecular Medicine and Neuroscience, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland, USA.

Progressive multifocal leukoencephalopathy (PML)-derived noncoding control region (NCCR) sequences permitted greater early viral gene expression than kidney-associated NCCR sequences. This was driven in part by binding of the transcription factor Spi-B to unique PML-associated Spi-B binding sites. Spi-B is upregulated in developing B cells in response to natalizumab therapy, a known risk factor for PML. Naturally occurring JCV sequence variation, together with drug treatment-induced cellular changes, may synergize to create an environment leading to an increased risk of PML.
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http://dx.doi.org/10.1128/JVI.03221-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3993791PMC
May 2014

Two demyelinating diseases in the brain of a single patient, PML and MS: how to minimize 'one' while treating the 'other'.

Expert Rev Clin Immunol 2013 Oct;9(10):887-90

Laboratory of Molecular Medicine and Neuroscience, NINDS, National Institutes of Health Bethesda, MD 20892, USA.

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http://dx.doi.org/10.1586/1744666X.2013.841559DOI Listing
October 2013