Publications by authors named "Maria A Nagel"

92 Publications

Transcriptional profiling reveals potential involvement of microvillous TRPM5-expressing cells in viral infection of the olfactory epithelium.

BMC Genomics 2021 Mar 30;22(1):224. Epub 2021 Mar 30.

Neuroscience Graduate Program, University of Colorado Anschutz Medical Campus, Aurora, CO, 80045, USA.

Background: Understanding viral infection of the olfactory epithelium is essential because the olfactory nerve is an important route of entry for viruses to the central nervous system. Specialized chemosensory epithelial cells that express the transient receptor potential cation channel subfamily M member 5 (TRPM5) are found throughout the airways and intestinal epithelium and are involved in responses to viral infection.

Results: Herein we performed deep transcriptional profiling of olfactory epithelial cells sorted by flow cytometry based on the expression of mCherry as a marker for olfactory sensory neurons and for eGFP in OMP-H2B::mCherry/TRPM5-eGFP transgenic mice (Mus musculus). We find profuse expression of transcripts involved in inflammation, immunity and viral infection in TRPM5-expressing microvillous cells compared to olfactory sensory neurons.

Conclusion: Our study provides new insights into a potential role for TRPM5-expressing microvillous cells in viral infection of the olfactory epithelium. We find that, as found for solitary chemosensory cells (SCCs) and brush cells in the airway epithelium, and for tuft cells in the intestine, the transcriptome of TRPM5-expressing microvillous cells indicates that they are likely involved in the inflammatory response elicited by viral infection of the olfactory epithelium.
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http://dx.doi.org/10.1186/s12864-021-07528-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8007386PMC
March 2021

Detection of varicella zoster virus antigen and DNA in two cases of cerebral amyloid angiopathy.

J Neurol Sci 2021 Mar 9;422:117315. Epub 2021 Jan 9.

Department of Neurology, University of Colorado School of Medicine, Aurora, CO 80045, United States; Department of Ophthalmology, University of Colorado School of Medicine, Aurora, CO 80045, United States. Electronic address:

Objective: Varicella zoster virus (VZV) vasculopathy and cerebral amyloid angiopathy (CAA) have similar clinical presentations: both affect cerebrovasculature in the elderly, produce hemorrhage, and can have a protracted course of cognitive decline and other neurological deficits. The cause of CAA is unknown, but amyloid-beta (Aβ) is found within arterial walls. Recent studies show that VZV induces Aβ and amylin expression and an amyloid-promoting environment. Thus, we determined if VZV was present in CAA-affected arteries.

Methods: Two subjects with pathologically-verified CAA were identified postmortem and frontal lobes analyzed by immunohistochemistry for arteries containing VZV, Aβ, and amylin and H&E for pathological changes. VZV antigen detection was confirmed by PCR for VZV DNA in the same region.

Results: In both CAA cases, sections with cerebral arteries containing VZV antigen with corresponding VZV DNA were identified; VZV antigen co-localized with Aβ in media of arteries with histological changes characteristic of CAA. Amylin was also seen in the intima of a VZV-positive artery in the diabetic subject. Not all Aβ-containing arteries had VZV, but all VZV-positive arteries contained Aβ.

Conclusions: VZV antigen co-localized with Aβ in some affected arteries from two CAA cases, suggesting a possible association between VZV infection and CAA.
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http://dx.doi.org/10.1016/j.jns.2021.117315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7935758PMC
March 2021

Elevated serum substance P during simian varicella virus infection in rhesus macaques: implications for chronic inflammation and adverse cerebrovascular events.

J Neurovirol 2020 12 22;26(6):945-951. Epub 2020 Sep 22.

Department of Neurology, University of Colorado School of Medicine, 12700 E. 19th Avenue, Mail Stop B182, Aurora, CO, 80045, USA.

Varicella and zoster, produced by varicella-zoster virus (VZV), are associated with an increased risk of stroke that may be due to persistent inflammation and hypercoagulability. Because substance P is associated with inflammation, hypercoagulability, and atherosclerotic plaque rupture that may contribute to increased stroke risk after VZV infection, we measured serum substance P in simian varicella virus-infected rhesus macaques. We found significantly increased and persistent serum substance P concentrations during varicella and zoster compared with pre-inoculation, supporting the hypothesis that VZV-induced increases in serum substance P may contribute to increased stroke risk associated with VZV infection.
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http://dx.doi.org/10.1007/s13365-020-00907-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7718397PMC
December 2020

Amylin, Aβ42, and Amyloid in Varicella Zoster Virus Vasculopathy Cerebrospinal Fluid and Infected Vascular Cells.

J Infect Dis 2021 Apr;223(7):1284-1294

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, USA.

Background: Varicella zoster virus (VZV) vasculopathy is characterized by persistent arterial inflammation leading to stroke. Studies show that VZV induces amyloid formation that may aggravate vasculitis. Thus, we determined if VZV central nervous system infection produces amyloid.

Methods: Aβ peptides, amylin, and amyloid were measured in cerebrospinal fluid (CSF) from 16 VZV vasculopathy subjects and 36 stroke controls. To determine if infection induced amyloid deposition, mock- and VZV-infected quiescent primary human perineurial cells (qHPNCs), present in vasculature, were analyzed for intracellular amyloidogenic transcripts/proteins and amyloid. Supernatants were assayed for amyloidogenic peptides and ability to induce amyloid formation. To determine amylin's function during infection, amylin was knocked down with small interfering RNA and viral complementary DNA (cDNA) was quantitated.

Results: Compared to controls, VZV vasculopathy CSF had increased amyloid that positively correlated with amylin and anti-VZV antibody levels; Aβ40 was reduced and Aβ42 unchanged. Intracellular amylin, Aβ42, and amyloid were seen only in VZV-infected qHPNCs. VZV-infected supernatant formed amyloid fibrils following addition of amyloidogenic peptides. Amylin knockdown decreased viral cDNA.

Conclusions: VZV infection increased levels of amyloidogenic peptides and amyloid in CSF and qHPNCs, indicating that VZV-induced amyloid deposition may contribute to persistent arterial inflammation in VZV vasculopathy. In addition, we identified a novel proviral function of amylin.
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http://dx.doi.org/10.1093/infdis/jiaa513DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8030705PMC
April 2021

Transcriptional profiling reveals TRPM5-expressing cells involved in viral infection in the olfactory epithelium.

bioRxiv 2020 May 15. Epub 2020 May 15.

Understanding viral infection of the olfactory epithelium is essential because smell loss can occur with coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus clade 2 (SARS-CoV-2), and because the olfactory nerve is an important route of entry for viruses to the central nervous system. Specialized chemosensory epithelial cells that express the transient receptor potential cation channel subfamily M member 5 (TRPM5) are found throughout the airways and intestinal epithelium and are involved in responses to viral infection. Herein we performed deep transcriptional profiling of olfactory epithelial cells sorted by flow cytometry based on the expression of fluorescent protein markers for olfactory sensory neurons and TRPM5. We find profuse expression of transcripts involved in inflammation, immunity and viral infection in TRPM5-expressing microvillous cells and olfactory sensory neurons. These cells express the transcript that encodes for a serine protease that primes the SARS-CoV-2 spike protein before entry into host cells. Our study provides new insights into a potential role for TRPM5-expressing cells in viral infection of the olfactory epithelium.
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http://dx.doi.org/10.1101/2020.05.14.096016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7263547PMC
May 2020

Acute zoster plasma contains elevated amyloid, correlating with Aβ42 and amylin levels, and is amyloidogenic.

J Neurovirol 2020 06 8;26(3):422-428. Epub 2020 May 8.

Department of Neurology, University of Colorado School of Medicine, 12700 E. 19th Avenue, Mail Stop B182, Aurora, CO, 80045, USA.

Herpes zoster is associated with an increased dementia and neovascular macular degeneration risk and a decline in glycemic control in diabetes mellitus. Because amyloid is present and pathogenic in these diseases, we quantified amyloid, Aβ40, Aβ42, and amylin in 14 zoster and 10 control plasmas. Compared with controls, zoster plasma had significantly elevated amyloid that correlated with Aβ42 and amylin levels and increased amyloid aggregation with addition of exogenous Aβ42 or amylin. These results suggest that zoster plasma contains factor(s) that promotes aggregation of amyloidogenic peptides, potentially contributing to the toxic amyloid burden and explaining accelerated disease progression following zoster.
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http://dx.doi.org/10.1007/s13365-020-00830-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7528582PMC
June 2020

Central nervous system infections produced by varicella zoster virus.

Curr Opin Infect Dis 2020 06;33(3):273-278

Department of Neurology.

Purpose Of Review: Varicella zoster virus (VZV) causes varicella, establishes latency, then reactivates to produce herpes zoster. VZV reactivation can also cause central nervous system (CNS) disease with or without rash. Herein, we review these CNS diseases, pathogenesis, diagnosis, and treatment.

Recent Findings: The most common CNS manifestation of VZV infection is vasculopathy that presents as headache, cognitive decline, and/or focal neurological deficits. VZV vasculopathy has also been associated with cerebral amyloid angiopathy and moyamoya syndrome. Rarely, VZV will produce a meningitis, encephalitis, cerebellitis, and myelopathy. Pathogenic mechanisms include direct VZV infection of affected tissue, persistent inflammation, and/or virus-induced hypercoagulability. Diagnosis is confirmed by the temporal association of rash to disease onset, intrathecal synthesis of anti-VZV antibodies, and/or the presence of VZV DNA in CSF. Most cases respond to intravenous acyclovir with corticosteroids.

Summary: VZV produces a wide spectrum of CNS disorders that may be missed as some cases do not have an associated rash or a CSF pleocytosis. Clinicians must be vigilant in including VZV in their differential diagnosis of CNS infections as VZV is a ubiquitous pathogen; importantly, VZV CNS infections are treatable with intravenous acyclovir therapy and corticosteroids.
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http://dx.doi.org/10.1097/QCO.0000000000000647DOI Listing
June 2020

Varicella-Zoster Virus Infection of Primary Human Spinal Astrocytes Produces Intracellular Amylin, Amyloid-β, and an Amyloidogenic Extracellular Environment.

J Infect Dis 2020 03;221(7):1088-1097

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, USA.

Background: Herpes zoster is linked to amyloid-associated diseases, including dementia, macular degeneration, and diabetes mellitus, in epidemiological studies. Thus, we examined whether varicella-zoster virus (VZV)-infected cells produce amyloid.

Methods: Production of intracellular amyloidogenic proteins (amylin, amyloid precursor protein [APP], and amyloid-β [Aβ]) and amyloid, as well as extracellular amylin, Aβ, and amyloid, was compared between mock- and VZV-infected quiescent primary human spinal astrocytes (qHA-sps). The ability of supernatant from infected cells to induce amylin or Aβ42 aggregation was quantitated. Finally, the amyloidogenic activity of viral peptides was examined.

Results: VZV-infected qHA-sps, but not mock-infected qHA-sps, contained intracellular amylin, APP, and/or Aβ, and amyloid. No differences in extracellular amylin, Aβ40, or Aβ42 were detected, yet only supernatant from VZV-infected cells induced amylin aggregation and, to a lesser extent, Aβ42 aggregation into amyloid fibrils. VZV glycoprotein B (gB) peptides assembled into fibrils and catalyzed amylin and Aβ42 aggregation.

Conclusions: VZV-infected qHA-sps produced intracellular amyloid and their extracellular environment promoted aggregation of cellular peptides into amyloid fibrils that may be due, in part, to VZV gB peptides. These findings suggest that together with host and other environmental factors, VZV infection may increase the toxic amyloid burden and contribute to amyloid-associated disease progression.
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http://dx.doi.org/10.1093/infdis/jiz560DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7075411PMC
March 2020

Herpes Zoster, a Rash of Cerebrovascular Events.

Mayo Clin Proc 2019 05;94(5):742-744

Department of Neurology, University of Colorado School of Medicine, Aurora.

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http://dx.doi.org/10.1016/j.mayocp.2019.03.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6822385PMC
May 2019

Varicella zoster virus productively infects human peripheral blood mononuclear cells to modulate expression of immunoinhibitory proteins and blocking PD-L1 enhances virus-specific CD8+ T cell effector function.

PLoS Pathog 2019 03 14;15(3):e1007650. Epub 2019 Mar 14.

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, United States of America.

Varicella zoster virus (VZV) is a lymphotropic alpha-herpesvirinae subfamily member that produces varicella on primary infection and causes zoster, vascular disease and vision loss upon reactivation from latency. VZV-infected peripheral blood mononuclear cells (PBMCs) disseminate virus to distal organs to produce clinical disease. To assess immune evasion strategies elicited by VZV that may contribute to dissemination of infection, human PBMCs and VZV-specific CD8+ T cells (V-CD8+) were mock- or VZV-infected and analyzed for immunoinhibitory protein PD-1, PD-L1, PD-L2, CTLA-4, LAG-3 and TIM-3 expression using flow cytometry. All VZV-infected PBMCs (monocytes, NK, NKT, B cells, CD4+ and CD8+ T cells) and V-CD8+ showed significant elevations in PD-L1 expression compared to uninfected cells. VZV induced PD-L2 expression in B cells and V-CD8+. Only VZV-infected CD8+ T cells, NKT cells and V-CD8+ upregulated PD-1 expression, the immunoinhibitory receptor for PD-L1/PD-L2. VZV induced CTLA-4 expression only in V-CD8+ and no significant changes in LAG-3 or TIM-3 expression were observed in V-CD8+ or PBMC T cells. To test whether PD-L1, PD-L2 or CTLA-4 regulates V-CD8+ effector function, autologous PBMCs were VZV-infected and co-cultured with V-CD8+ cells in the presence of blocking antibodies against PD-L1, PD-L2 or CTLA-4; ELISAs revealed significant elevations in IFNγ only upon blocking of PD-L1. Together, these results identified additional immune cells that are permissive to VZV infection (monocytes, B cells and NKT cells); along with a novel mechanism for inhibiting CD8+ T cell effector function through induction of PD-L1 expression.
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http://dx.doi.org/10.1371/journal.ppat.1007650DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6435197PMC
March 2019

Varicella Zoster Virus Alters Expression of Cell Adhesion Proteins in Human Perineurial Cells via Interleukin 6.

J Infect Dis 2019 09;220(9):1453-1461

Department of Neurology, University of Colorado School of Medicine, Aurora.

Background: In temporal arteries (TAs) from patients with giant cell arteritis, varicella zoster virus (VZV) is seen in perineurial cells that surround adventitial nerve bundles and form the peripheral nerve-extrafascicular tissue barrier (perineurium). We hypothesized that during VZV reactivation from ganglia, virus travels transaxonally and disrupts the perineurium to infect surrounding cells.

Methods: Mock- and VZV-infected primary human perineurial cells (HPNCs) were examined for alterations in claudin-1, E-cadherin, and N-cadherin. Conditioned supernatant was analyzed for a soluble factor(s) mediating these alterations and for the ability to increase cell migration. To corroborate in vitro findings, a VZV-infected TA was examined.

Results: In VZV-infected HPNCs, claudin-1 redistributed to the nucleus; E-cadherin was lost and N-cadherin gained, with similar changes seen in VZV-infected perineurial cells in a TA. VZV-conditioned supernatant contained increased interleukin 6 (IL-6) that induced E-cadherin loss and N-cadherin gain and increased cell migration when added to uninfected HPNCs; anti-IL-6 receptor antibody prevented these changes.

Conclusions: IL-6 secreted from VZV-infected HPNCs facilitated changes in E- and N-cadherin expression and cell migration, reminiscent of an epithelial-to-mesenchymal cell transition, potentially contributing to loss of perineurial cell barrier integrity and viral spread. Importantly, an anti-IL-6 receptor antibody prevented virus-induced perineurial cell disruption.
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http://dx.doi.org/10.1093/infdis/jiz095DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6761973PMC
September 2019

Varicella Zoster Virus Induces Differential Cell-Type Specific Responses in Human Corneal Epithelial Cells and Keratocytes.

Invest Ophthalmol Vis Sci 2019 02;60(2):704-711

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, United States.

Purpose: While VZV DNA and antigen have been detected in acute and chronic VZV keratitis, it is unclear whether productive infection of corneal cells is ongoing or whether residual, noninfectious VZV antigens elicit inflammation. Herein, we examined VZV-infected primary human corneal epithelial cells (HCECs) and keratocytes (HKs) to elucidate the pathogenesis of VZV keratitis.

Methods: HCECs and HKs were mock- or VZV infected. Seven days later, cells were examined for morphology, proinflammatory cytokine and matrix metalloproteinase (MMP) release, ability to recruit peripheral blood mononuclear cells (PBMCs) and neutrophils, and MMP substrate cleavage.

Results: Both cell types synthesized infectious virus. VZV-infected HCECs proliferated, whereas VZV-infected HKs died. Compared to mock-infected cells, VZV-infected HCECs secreted significantly more IL-6, IL-8, IL-10, and IL-12p70 that were confirmed at the transcript level, and MMP-1 and MMP-9; conditioned supernatant attracted PBMCs and neutrophils and cleaved MMP substrates. In contrast, VZV-infected HKs suppressed cytokine secretion except for IL-8, which attracted neutrophils, and suppressed MMP release and substrate cleavage.

Conclusions: Overall, VZV-infected HCECs recapitulate findings of VZV keratitis with respect to epithelial cell proliferation, pseudodendrite formation and creation of a proinflammatory environment, providing an in vitro model for VZV infection of corneal epithelial cells. Furthermore, the proliferation and persistence of VZV-infected HCECs suggest that these cells may serve as viral reservoirs if immune clearance is incomplete. Finally, the finding that VZV-infected HKs die and suppress most proinflammatory cytokines and MMPs may explain the widespread death of these cells with unchecked viral spread due to ineffective recruitment of PBMCs.
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http://dx.doi.org/10.1167/iovs.18-25801DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6383726PMC
February 2019

Varicella zoster virus differentially alters morphology and suppresses proinflammatory cytokines in primary human spinal cord and hippocampal astrocytes.

J Neuroinflammation 2018 Nov 15;15(1):318. Epub 2018 Nov 15.

Department of Neurology, University of Colorado School of Medicine, 4200 E. 19th Avenue, Mail Stop B182, Aurora, CO, 80045, USA.

Background: Varicella zoster virus (VZV) is a ubiquitous alphaherpesvirus that produces varicella and zoster. VZV can infect multiple cell types in the spinal cord and brain, including astrocytes, producing myelopathy and encephalopathy. While studies of VZV-astrocyte interactions are sparse, a recent report showed that quiescent primary human spinal cord astrocytes (qHA-sps) did not appear activated morphologically during VZV infection. Since astrocytes play a critical role in host defenses during viral infections of the central nervous system, we examined the cytokine responses of qHA-sps and quiescent primary human hippocampal astrocytes (qHA-hps) to VZV infection in vitro, as well as the ability of conditioned supernatant to recruit immune cells.

Methods: At 3 days post-infection, mock- and VZV-infected qHA-sps and qHA-hps were examined for morphological changes by immunofluorescence antibody assay using antibodies directed against glial fibrillary acidic protein and VZV. Conditioned supernatants were analyzed for proinflammatory cytokines [interleukin (IL)-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, interferon-gamma, and tumor necrosis factor-α] using the Meso Scale Discovery multiplex ELISA platform. Finally, the ability of conditioned supernatants to attract peripheral blood mononuclear cells (PBMCs) was determined using a chemotaxis assay. Quiescent primary human perineurial cells (qHPNCs) served as a control for VZV-induced cytokine production and PBMC migration. To confirm that the astrocytes have the ability to increase cytokine secretion, qHA-sps and qHA-hps were treated with IL-1β and examined for morphological changes and IL-6 secretion.

Results: VZV-infected qHA-sps displayed extensive cellular processes, whereas VZV-infected qHA-hps became swollen and clustered together. Astrocytes had the capacity to secrete IL-6 in response to IL-1β. Compared to mock-infected cells, VZV-infected qHA-sps showed significantly reduced secretion of IL-2, IL-4, IL-6, IL-12p70, and IL-13, while VZV-infected qHA-hps showed significantly reduced IL-8 secretion. In contrast, levels of all 10 cytokines examined were significantly increased in VZV-infected qHPNCs. Consistent with these results, conditioned supernatant from VZV-infected qHPNCs, but not that from VZV-infected qHA-sps and qHA-hps, recruited PBMCs.

Conclusions: VZV-infected qHA-sps and qHA-hps have distinct morphological alterations and patterns of proinflammatory cytokine suppression that could contribute to ineffective viral clearance in VZV myelopathy and encephalopathy, respectively.
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http://dx.doi.org/10.1186/s12974-018-1360-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6236967PMC
November 2018

Reactivation of Simian Varicella Virus in Rhesus Macaques after CD4 T Cell Depletion.

J Virol 2019 02 17;93(3). Epub 2019 Jan 17.

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, USA

Rhesus macaques intrabronchially inoculated with simian varicella virus (SVV), the counterpart of human varicella-zoster virus (VZV), developed primary infection with viremia and rash, which resolved upon clearance of viremia, followed by the establishment of latency. To assess the role of CD4 T cell immunity in reactivation, monkeys were treated with a single 50-mg/kg dose of a humanized monoclonal anti-CD4 antibody; within 1 week, circulating CD4 T cells were reduced from 40 to 60% to 5 to 30% of the total T cell population and remained low for 2 months. Very low viremia was seen only in some of the treated monkeys. Zoster rash developed after 7 days in the monkey with the most extensive CD4 T cell depletion (5%) and in all other monkeys at 10 to 49 days posttreatment, with recurrent zoster in one treated monkey. SVV DNA was detected in the lung from two of five monkeys, in bronchial lymph nodes from one of the five monkeys, and in ganglia from at least two dermatomes in three of five monkeys. Immunofluorescence analysis of skin rash, lungs, lymph nodes, and ganglia revealed SVV ORF63 protein at the following sites: sweat glands in skin; type II cells in lung alveoli, macrophages, and dendritic cells in lymph nodes; and the neuronal cytoplasm of ganglia. Detection of SVV antigen in multiple tissues upon CD4 T cell depletion and virus reactivation suggests a critical role for CD4 T cell immunity in controlling varicella virus latency. Reactivation of latent VZV in humans can result in serious neurological complications. VZV-specific cell-mediated immunity is critical for the maintenance of latency. Similar to VZV in humans, SVV causes varicella in monkeys, establishes latency in ganglia, and reactivates to produce shingles. Here, we show that depletion of CD4 T cells in rhesus macaques results in SVV reactivation, with virus antigens found in zoster rash and SVV DNA and antigens found in lungs, lymph nodes, and ganglia. These results suggest the critical role of CD4 T cell immunity in controlling varicella virus latency.
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http://dx.doi.org/10.1128/JVI.01375-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6340024PMC
February 2019

Varicella Zoster Virus Vasculopathy.

J Infect Dis 2018 09;218(suppl_2):S107-S112

Department of Neurology, University of Colorado School of Medicine, Aurora.

Varicella zoster virus (VZV) is a ubiquitous, exclusively human alphaherpesvirus that produces varicella then becomes latent in ganglionic neurons. In elderly and immunocompromised individuals, VZV reactivates and typically produces herpes zoster. Studies of patients with VZV vasculopathy have identified key clinical, imaging, and laboratory features to assist in diagnosis and treatment. Complementary studies have further expanded the spectrum of VZV vasculopathy to include the extracranial circulation and identified mechanisms contributing to its pathogenesis. Given our increasing aging population and recognition that VZV reactivation manifesting as zoster is a risk factor for stroke and myocardial infarction, recognition of VZV as a potential cause of vascular disease with or without associated zoster rash is essential to decrease associated morbidity and mortality because VZV vasculopathy can be treated with antiviral therapy.
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http://dx.doi.org/10.1093/infdis/jiy425DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6151079PMC
September 2018

Nanoparticle uptake by circulating leukocytes: A major barrier to tumor delivery.

J Control Release 2018 09 17;286:85-93. Epub 2018 Jul 17.

Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Anschutz Medical Campus, Aurora, CO, United States. Electronic address:

Decades of research into improving drug delivery to tumors has documented uptake of particulate delivery systems by resident macrophages in the lung, liver, and spleen, and correlated short circulation times with reduced tumor accumulation. An implicit assumption in these studies is that nanoparticles present in the blood are available for distribution to the tumor. This study documents significant levels of lipoplex uptake by circulating leukocytes, and its effect on distribution to the tumor and other organs. In agreement with previous studies, PEGylation dramatically extends circulation times and enhances tumor delivery. However, our studies suggest that this relationship is not straightforward, and that particle sequestration by leukocytes can significantly alter biodistribution, especially with non-PEGylated nanoparticle formulations. We conclude that leukocyte uptake should be considered in biodistribution studies, and that delivery to these circulating cells may present opportunities for treating viral infections and leukemia.
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http://dx.doi.org/10.1016/j.jconrel.2018.07.031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6936323PMC
September 2018

Varicella Zoster Virus Induces Nuclear Translocation of the Neurokinin-1 Receptor, Promoting Lamellipodia Formation and Viral Spread in Spinal Astrocytes.

J Infect Dis 2018 09;218(8):1324-1335

Department of Neurology, University of Colorado School of Medicine, Aurora.

Background: Varicella zoster virus (VZV) can present as a myelopathy with spinal astrocyte infection. Recent studies support a role for the neurokinin-1 receptor (NK-1R) in virus infections, as well as for cytoskeletal alterations that may promote viral spread. Thus, we examined the role of NK-1R in VZV-infected primary human spinal astrocytes (HA-sps) to shed light on the pathogenesis of VZV myelopathy.

Methods: Mock- and VZV-infected HA-sps were examined for substance P (subP) production, NK-1R localization, morphological changes, and viral spread in the presence or absence of the NK-1R antagonists aprepitant and rolapitant.

Results: VZV infection of HA-sps induced nuclear localization of full-length and truncated NK-1R in the absence of the endogenous ligand, subP, and was associated with extensive lamellipodia formation and viral spread that was inhibited by NK-1R antagonists.

Conclusions: We have identified a novel, subP-independent, proviral function of nuclear NK-1R associated with lamellipodia formation and viral spread that is distinct from subP-induced NK-1R cell membrane/cytoplasmic localization without lamellipodia formation. These results suggest that binding of a putative viral ligand to NK-1R produces a dramatically different NK-1R downstream effect than binding of subP. Finally, the NK-1R antagonists aprepitant and rolapitant provide promising alternatives to nucleoside analogs in treating VZV infections, including myelopathy.
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http://dx.doi.org/10.1093/infdis/jiy297DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6129113PMC
September 2018

Attenuation of Simian Varicella Virus Infection by Enhanced Green Fluorescent Protein in Rhesus Macaques.

J Virol 2018 04 14;92(7). Epub 2018 Mar 14.

Division of Microbiology, Tulane University, Tulane National Primate Research Center, Covington, Louisiana, USA.

Simian varicella virus (SVV), the primate counterpart of varicella-zoster virus, causes varicella (chickenpox), establishes latency in ganglia, and reactivates to produce zoster. We previously demonstrated that a recombinant SVV expressing enhanced green fluorescent protein (rSVV.eGFP) is slightly attenuated both in culture and in infected monkeys. Here, we generated two additional recombinant SVVs to visualize infected cells and One harbors eGFP fused to the N terminus of open reading frame 9 (ORF9) (rSVV.eGFP-2a-ORF9), and another harbors eGFP fused to the C terminus of ORF66 (rSVV.eGFP-ORF66). Both recombinant viruses efficiently expressed eGFP in cultured cells. Both recombinant SVV infections in culture were comparable to that of wild-type SVV (SVV.wt). Unlike SVV.wt, eGFP-tagged SVV did not replicate in rhesus cells in culture. Intratracheal (i.t.) or i.t. plus intravenous (i.v.) inoculation of rhesus macaques with these new eGFP-tagged viruses resulted in low viremia without varicella rash, although SVV DNA was abundant in bronchoalveolar lavage (BAL) fluid at 10 days postinoculation (dpi). SVV DNA was also found in trigeminal ganglia of one monkey inoculated with rSVV.eGFP-ORF66. Intriguingly, a humoral response to both SVV and eGFP was observed. In addition, monkeys inoculated with the eGFP-expressing viruses were protected from superinfection with SVV.wt, suggesting that the monkeys had mounted an efficient immune response. Together, our results show that eGFP expression could be responsible for their reduced pathogenesis. SVV infection in nonhuman primates has served as an extremely useful animal model to study varicella-zoster virus (VZV) pathogenesis. eGFP-tagged viruses are a great tool to investigate their pathogenesis. We constructed and tested two new recombinant SVVs with eGFP inserted into two different locations in the SVV genome. Both recombinant SVVs showed robust replication in culture but reduced viremia compared to that with SVV.wt during primary infection in rhesus macaques. Our results indicate that conclusions on eGFP-tagged viruses based on results should be handled with care, since eGFP expression could result in attenuation of the virus.
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http://dx.doi.org/10.1128/JVI.02253-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5972883PMC
April 2018

Varicella zoster virus-infected cerebrovascular cells produce a proinflammatory environment.

Neurol Neuroimmunol Neuroinflamm 2017 Sep 13;4(5):e382. Epub 2017 Jul 13.

Department of Neurology (D.J., M.A.N.), Department of Medicine (C.P.N., B.E.P.), and Department of Pediatrics (K.S.), University of Colorado School of Medicine, Aurora.

Objective: To test whether varicella zoster virus (VZV) infection of human brain vascular cells and of lung fibroblasts directly increases proinflammatory cytokine levels, consistent with VZV as a causative agent in intracerebral VZV vasculopathy and giant-cell arteritis (GCA).

Methods: Conditioned supernatant from mock- and VZV-infected human brain vascular adventitial fibroblasts (HBVAFs), human perineurial cells (HPNCs), human brain vascular smooth muscle cells (HBVSMCs), and human fetal lung fibroblasts (HFLs) were collected at 72 hours postinfection and analyzed for levels of 30 proinflammatory cytokines using the Meso Scale Discovery Multiplex ELISA platform.

Results: Compared with mock infection, VZV infection led to significantly increased levels of the following: interleukin-8 (IL-8) in all cell lines examined; IL-6 in HBVAFs, HPNCs, and HFLs, with no change in HBVSMCs; and vascular endothelial growth factor A in HBVAFs, HBVSMCs, and HFLs, with a significant decrease in HPNCs. Other cytokines, including IL-2, IL-4, IL-15, IL-16, TGF-b, Eotaxin-1, Eotaxin-3, IP-10, MCP-1, and granulocyte macrophage colony-stimulating factor, were also significantly altered upon VZV infection in a cell type-specific manner.

Conclusions: VZV infection of vascular cells can directly produce a proinflammatory environment that may potentially lead to prolonged arterial wall inflammation and vasculitis. The VZV-mediated increase in IL-8 and IL-6 is consistent with that seen in the CSF of patients with intracerebral VZV vasculopathy, and the VZV-mediated increase in IL-6 is consistent with the cytokine's elevated levels in temporal arteries and plasma of patients with GCA.
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http://dx.doi.org/10.1212/NXI.0000000000000382DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5682918PMC
September 2017

Varicella zoster virus vasculopathy: The expanding clinical spectrum and pathogenesis.

J Neuroimmunol 2017 07 18;308:112-117. Epub 2017 Mar 18.

Department of Neurology, University of Colorado School of Medicine, Aurora, CO 80045, USA. Electronic address:

Varicella zoster virus (VZV) is a ubiquitous, human alphaherpesvirus that produces varicella on primary infection then becomes latent in ganglionic neurons along the entire neuraxis. In elderly and immunocompromised individuals, VZV reactivates and travels along nerve fibers peripherally resulting in zoster. However, VZV can also spread centrally and infect cerebral and extracranial arteries (VZV vasculopathy) to produce transient ischemic attacks, stroke, aneurysm, sinus thrombosis and giant cell arteritis, as well as granulomatous aortitis. The mechanisms of virus-induced pathological vascular remodeling are not fully elucidated; however, recent studies suggest that inflammation and dysregulation of programmed death ligand-1 play a significant role.
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http://dx.doi.org/10.1016/j.jneuroim.2017.03.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5489071PMC
July 2017

Donald H. Gilden, M.D.

J Neuroimmunol 2017 07 6;308:2-5. Epub 2017 Jan 6.

Department of Neurology, University of Colorado School of Medicine, Aurora, CO, USA; Department of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, CO, USA. Electronic address:

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http://dx.doi.org/10.1016/j.jneuroim.2017.01.002DOI Listing
July 2017

Varicella-Zoster Virus Downregulates Programmed Death Ligand 1 and Major Histocompatibility Complex Class I in Human Brain Vascular Adventitial Fibroblasts, Perineurial Cells, and Lung Fibroblasts.

J Virol 2016 Dec 14;90(23):10527-10534. Epub 2016 Nov 14.

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, USA

Varicella-zoster virus (VZV) vasculopathy produces stroke, giant cell arteritis, and granulomatous aortitis, and it develops after virus reactivates from ganglia and spreads transaxonally to arterial adventitia, resulting in persistent inflammation and pathological vascular remodeling. The mechanism(s) by which inflammatory cells persist in VZV-infected arteries is unknown; however, virus-induced dysregulation of programmed death ligand 1 (PD-L1) may play a role. Specifically, PD-L1 can be expressed on virtually all nucleated cells and suppresses the immune system by interacting with the programmed cell death protein receptor 1, found exclusively on immune cells; thus, downregulation of PD-L1 may promote inflammation, as seen in some autoimmune diseases. Both flow cytometry and immunofluorescence analyses to test whether VZV infection of adventitial cells downregulates PD-L1 showed decreased PD-L1 expression in VZV-infected compared to mock-infected human brain vascular adventitial fibroblasts (HBVAFs), perineural cells (HPNCs), and fetal lung fibroblasts (HFLs) at 72 h postinfection. Quantitative RT-PCR analyses showed no change in PD-L1 transcript levels between mock- and VZV-infected cells, indicating a posttranscriptional mechanism for VZV-mediated downregulation of PD-L1. Flow cytometry analyses showed decreased major histocompatibility complex class I (MHC-I) expression in VZV-infected cells and adjacent uninfected cells compared to mock-infected cells. These data suggest that reduced PD-L1 expression in VZV-infected adventitial cells contribute to persistent vascular inflammation observed in virus-infected arteries from patients with VZV vasculopathy, while downregulation of MHC-I prevents viral clearance.

Importance: Here, we provide the first demonstration that VZV downregulates PD-L1 expression in infected HBVAFs, HPNCs, and HFLs, which, together with the noted VZV-mediated downregulation of MHC-I, might foster persistent inflammation in vessels, leading to pathological vascular remodeling during VZV vasculopathy and persistent inflammation in infected lungs to promote subsequent infection of T cells and hematogenous virus spread. Identification of a potential mechanism by which persistent inflammation in the absence of effective viral clearance occurs in VZV vasculopathy and VZV infection of the lung is a step toward targeted therapy of VZV-induced disease.
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http://dx.doi.org/10.1128/JVI.01546-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5110195PMC
December 2016

Successful antiviral treatment after 6years of chronic progressive neurological disease attributed to VZV brain infection.

J Neurol Sci 2016 Sep 15;368:240-2. Epub 2016 Jul 15.

Department of Neurology, University of Colorado School of Medicine, Aurora, CO, USA.

We describe an extraordinary case of an immunocompetent patient who developed sacral-distribution zoster, followed 3months later by neurological disease that progressed for 6years and was attributed to varicella zoster virus (VZV) infection of the brain. Despite the prolonged infection, neurologic symptoms and signs resolved rapidly and completely after treatment with intravenous acyclovir.
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http://dx.doi.org/10.1016/j.jns.2016.07.035DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4996351PMC
September 2016

Burning mouth syndrome associated with varicella zoster virus.

BMJ Case Rep 2016 Jul 5;2016. Epub 2016 Jul 5.

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, USA.

We present two cases of burning mouth syndrome (BMS)-of 8-month duration in a 61-year-old woman and of 2-year duration in a 63-year-old woman-both associated with increased levels of antivaricella zoster virus (VZV) IgM antibodies in serum and with pain that improved with antiviral treatment. Combined with our previous finding of BMS due to herpes simplex virus type 1 (HSV-1) infection, we recommend evaluation of patients with BMS not only for VZV or HSV-1 DNA in the saliva, but also for serum anti-VZV and anti-HSV-1 IgM antibodies. Both infections are treatable with oral antiviral agents.
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http://dx.doi.org/10.1136/bcr-2016-215953DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4956955PMC
July 2016

Proinflammatory cytokines and matrix metalloproteinases in CSF of patients with VZV vasculopathy.

Neurol Neuroimmunol Neuroinflamm 2016 Aug 13;3(4):e246. Epub 2016 Jun 13.

Departments of Neurology (D.J., E.A., S.S., D.G., M.A.N.) and Immunology and Microbiology (D.G.), University of Colorado School of Medicine, Aurora.

Objective: To determine the levels of proinflammatory cytokines and matrix metalloproteinases (MMPs) in the CSF of patients with virologically verified varicella zoster virus (VZV) vasculopathy.

Methods: CSF from 30 patients with virologically verified VZV vasculopathy was analyzed for levels of proinflammatory cytokines and MMPs using the Meso Scale Discovery multiplex ELISA platform. Positive CNS inflammatory disease controls were provided by CSF from 30 patients with multiple sclerosis. Negative controls were provided by CSF from 20 healthy controls.

Results: Compared to multiple sclerosis CSF and CSF from healthy controls, levels of interleukin (IL)-8, IL-6, and MMP-2 were significantly elevated in VZV vasculopathy CSF.

Conclusions: CSF of patients with VZV vasculopathy revealed a unique profile of elevated proinflammatory cytokines, IL-8 and IL-6, along with elevated MMP-2. The relevance of these cytokines to the pathogenesis of VZV vasculopathy requires further study.
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http://dx.doi.org/10.1212/NXI.0000000000000246DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4907802PMC
August 2016

SUNCT headaches after ipsilateral ophthalmic-distribution zoster.

J Neurol Sci 2016 Jul 14;366:207-208. Epub 2016 May 14.

Department of Neurology, University of Colorado School of Medicine, Aurora, CO, USA; Department of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, CO, USA. Electronic address:

Nine days after left ophthalmic-distribution zoster, a 47-year-old man developed SUNCT headaches (short-lasting unilateral neuralgiform headache with conjunctival injection and tearing). In contrast to two prior cases of SUNCT that developed after varicella zoster virus (VZV) meningoencephalitis without rash, this case describes an association of SUNCT with overt zoster, thus adding to the spectrum of headache and facial pain syndromes caused by VZV reactivation.
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http://dx.doi.org/10.1016/j.jns.2016.05.027DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4903154PMC
July 2016

Varicella zoster virus triggers the immunopathology of giant cell arteritis.

Curr Opin Rheumatol 2016 Jul;28(4):376-82

aDepartment of NeurologybDepartment of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, Colorado, USA.

Purpose Of Review: Giant cell arteritis (GCA) is a severe form of vasculitis in the elderly. The recent discovery of varicella zoster virus (VZV) in the temporal arteries and adjacent skeletal muscle of patients with GCA, and the rationale and strategy for antiviral and corticosteroid treatment for GCA are reviewed.

Recent Findings: The clinical features of GCA include excruciating headache/head pain, often with scalp tenderness, a nodular temporal arteries and decreased temporal artery pulsations. Jaw claudication, night sweats, fever, malaise, and a history of polymyalgia rheumatica (aching and stiffness of large muscles primarily in the shoulder girdle, upper back, and pelvis without objective signs of weakness) are common. ESR and CRP are usually elevated. Diagnosis is confirmed by temporal artery biopsy which reveals vessel wall damage and inflammation, with multinucleated giant cells and/or epithelioid macrophages. Skip lesions are common. Importantly, temporal artery biopsies are pathologically negative in many clinically suspect cases. This review highlights recent virological findings in temporal arteries from patients with pathologically verified GCA and in temporal arteries from patients who manifest clinical and laboratory features of GCA, but whose temporal artery biopsies (Bx) are pathologically negative for GCA (Bx-negative GCA). Virological analysis revealed that VZV is present in most GCA-positive and GCA-negative temporal artery biopsies, mostly in skip areas that correlate with adjacent GCA pathology.

Summary: The presence of VZV in Bx-positive and Bx-negative GCA temporal arteries indicates that VZV triggers the immunopathology of GCA. However, the presence of VZV in about 20% of temporal artery biopsies from non-GCA postmortem controls also suggests that VZV alone is not sufficient to produce disease. Treatment trials should be performed to determine if antiviral agents confer additional benefits to corticosteroids in both Bx-positive and Bx-negative GCA patients. These studies should also examine whether oral antiviral agents and corticosteroids are as effective as intravenous acyclovir and corticosteroids. Appropriate dosage and duration of treatment also remain to be determined.
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http://dx.doi.org/10.1097/BOR.0000000000000292DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4896311PMC
July 2016

Blinded search for varicella zoster virus in giant cell arteritis (GCA)-positive and GCA-negative temporal arteries.

J Neurol Sci 2016 May 19;364:141-3. Epub 2016 Mar 19.

Department of Neurology, University of Colorado School of Medicine, Aurora, CO, USA.

Recent analysis of archived temporal arteries (TAs) acquired from 13 pathology laboratories in the US, Canada, Iceland, France, Germany and Israel from patients with pathologically-verified giant cell arteritis (GCA-positive) and TAs from patients with clinical features and laboratory abnormalities of GCA but whose TAs were pathologically negative (GCA-negative) revealed VZV antigen in most TAs from both groups. Despite formalin-fixation, VZV DNA was also found in many VZV-antigen positive sections that were scraped, subjected to DNA extraction, and examined by PCR with VZV-specific primers. Importantly, in past studies, the pathological diagnosis (GCA-positive or -negative) was known to the neurovirology laboratory. Herein, GCA-positive and GCA-negative TAs were provided by an outside institution and examined by 4 investigators blinded to the pathological diagnoses. VZV antigen was found in 3/3 GCA-positive TAs and in 4/6 GCA-negative TAs, and VZV DNA in 1/3 VZV antigen-positive, GCA-positive TAs and in 3/4 VZV antigen-positive, GCA-negative TAs. VZV DNA was also detected in one GCA-negative, VZV-antigen negative TA. Overall, the detection of VZV antigen in 78% of GCA-positive and GCA-negative TAs is consistent with previous reports on the prevalence of VZV antigen in patients with clinically suspect GCA.
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http://dx.doi.org/10.1016/j.jns.2016.03.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4834150PMC
May 2016

Varicella Zoster Virus Infection in Granulomatous Arteritis of the Aorta.

J Infect Dis 2016 06 31;213(12):1866-71. Epub 2016 Mar 31.

Department of Neurology.

Granulomatous arteritis characterizes the pathology of giant cell arteritis, granulomatous aortitis, and intracerebral varicella zoster virus (VZV) vasculopathy. Because intracerebral VZV vasculopathy and giant cell arteritis are strongly associated with productive VZV infection in cerebral and temporal arteries, respectively, we evaluated human aortas for VZV antigen and VZV DNA. Using 3 different anti-VZV antibodies, we identified VZV antigen in 11 of 11 aortas with pathologically verified granulomatous arteritis, in 1 of 1 cases of nongranulomatous arteritis, and in 5 of 18 control aortas (28%) obtained at autopsy. The presence of VZV antigen in granulomatous aortitis was highly significant (P = .0001) as compared to control aortas, in which VZV antigen was never associated with pathology, indicating subclinical reactivation. VZV DNA was found in most aortas containing VZV antigen. The frequent clinical, radiological, and pathological aortic involvement in patients with giant cell arteritis correlates with the significant detection of VZV in granulomatous aortitis.
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http://dx.doi.org/10.1093/infdis/jiw101DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4878728PMC
June 2016

VZV in biopsy-positive and -negative giant cell arteritis: Analysis of 100+ temporal arteries.

Neurol Neuroimmunol Neuroinflamm 2016 Apr 9;3(2):e216. Epub 2016 Mar 9.

University of Colorado School of Medicine (D.G., T.W., N.K., M.A.N.), Aurora; and East Carolina University (P.J.B.), Greenville, NC.

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http://dx.doi.org/10.1212/NXI.0000000000000216DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4794807PMC
April 2016