VZV Infection of Primary Human Adrenal Cortical Cells Produces a Proinflammatory Environment without Cell Death.

Virus infection of adrenal glands can disrupt secretion of mineralocorticoids, glucocorticoids, and sex hormones from the cortex and catecholamines from the medulla, leading to a constellation of symptoms such as fatigue, dizziness, weight loss, nausea, and muscle and joint pain. Specifically, varicella zoster virus (VZV) can produce bilateral adrenal hemorrhage and adrenal insufficiency during primary infection or following reactivation. However, the mechanisms by which VZV affects the adrenal glands are not well-characterized. Herein, we determined if primary human adrenal cortical cells (HAdCCs) infected with VZV support viral replication and produce a proinflammatory environment. Quantitative PCR showed VZV DNA increasing over time in HAdCCs, yet no cell death was seen at 3 days post-infection by TUNEL staining or Western Blot analysis with PARP and caspase 9 antibodies. Compared to conditioned supernatant from mock-infected cells, supernatant from VZV-infected cells contained significantly elevated IL-6, IL-8, IL-12p70, IL-13, IL-4, and TNF-α. Overall, VZV can productively infect adrenal cortical cells in the absence of cell death, suggesting that these cells may be a potential reservoir for ongoing viral replication and proinflammatory cytokine production, leading to chronic adrenalitis and dysfunction.

Targeted RNA Sequencing of VZV-Infected Brain Vascular Adventitial Fibroblasts Indicates That Amyloid May Be Involved in VZV Vasculopathy.

BACKGROUND AND OBJECTIVES: Compared with stroke controls, patients with varicella zoster virus (VZV) vasculopathy have increased amyloid in CSF, along with increased amylin (islet amyloid polypeptide [IAPP]) and anti-VZV antibodies. Thus, we examined the gene expression profiles of VZV-infected primary human brain vascular adventitial fibroblasts (HBVAFs), one of the initial arterial cells infected in VZV vasculopathy, to determine whether they are a potential source of amyloid that can disrupt vasculature and potentiate inflammation. METHODS: Mock- and VZV-infected quiescent HBVAFs were harvested at 3 days postinfection. Targeted RNA sequencing of the whole-human transcriptome (BioSpyder Technologies, TempO-Seq) was conducted followed by gene set enrichment and pathway analysis. Selected pathways unique to VZV-infected cells were confirmed by enzyme-linked immunoassays, migration assays, and immunofluorescence analysis (IFA) that included antibodies against amylin and amyloid-beta, as well as amyloid staining by Thioflavin-T. RESULTS: Compared with mock, VZV-infected HBVAFs had significantly enriched gene expression pathways involved in vascular remodeling and vascular diseases; confirmatory studies showed secretion of matrix metalloproteinase-3 and -10, as well increased migration of infected cells and uninfected cells when exposed to conditioned media from VZV-infected cells. In addition, significantly enriched pathways involved in amyloid-associated diseases (diabetes mellitus, amyloidosis, and Alzheimer disease), tauopathy, and progressive neurologic disorder were identified; predicted upstream regulators included amyloid precursor protein, apolipoprotein E, microtubule-associated protein tau, presenilin 1, and IAPP. Confirmatory IFA showed that VZV-infected HBVAFs contained amyloidogenic peptides (amyloid-beta and amylin) and intracellular amyloid. DISCUSSION: Gene expression profiles and pathway enrichment analysis of VZV-infected HBVAFs, as well as phenotypic studies, reveal features of pathologic vascular remodeling (e.g., increased cell migration and changes in the extracellular matrix) that can contribute to cerebrovascular disease. Furthermore, the discovery of amyloid-associated transcriptional pathways and intracellular amyloid deposition in HBVAFs raise the possibility that VZV vasculopathy is an amyloid disease. Amyloid deposition may contribute to cell death and loss of vascular wall integrity, as well as potentiate chronic inflammation in VZV vasculopathy, with disease severity and recurrence determined by the host's ability to clear virus infection and amyloid deposition and by the coexistence of other amyloid-associated diseases (i.e., Alzheimer disease and diabetes mellitus).

Targeted RNA Sequencing of Formalin-Fixed, Paraffin-Embedded Temporal Arteries From Giant Cell Arteritis Cases Reveals Viral Signatures.

BACKGROUND AND OBJECTIVES: Varicella zoster virus (VZV) antigen has been detected in temporal arteries (TAs) of individuals with giant cell arteritis (GCA), the most common systemic vasculitis in older adults. Thus, we explored the contribution of VZV to GCA pathogenesis. METHODS: Formalin-fixed, paraffin-embedded TA sections from biopsy-positive GCA participants with VZV antigen (GCA/VZV-positive; n = 20) and without (GCA/VZV-negative, n = 20) and from normal participants with VZV antigen (control/VZV-positive, n = 11) and without (control/VZV-negative, n = 20) were analyzed by targeted RNA sequencing of the whole human transcriptome (BioSpyder TempO-Seq). Ingenuity pathway analysis and R-computational program were used to identify differentially expressed genes and pathways between groups. RESULTS: Compared with control/VZV-negative TAs, GCA/VZV-negative and GCA/VZV-positive TAs were significantly enriched for human transcripts specific for pathways involved in viral infections, including viral entry, nuclear factor kappa B activation by viruses, and other pathogen-related immune activation pathways. Similarly, human gene sets supporting viral infection were found in control/VZV-positive TAs that showed no morphological signs of inflammation, suggesting that the enriched pathways were not nonspecific signatures of infiltrating immune cells. All GCA TAs and control/VZV-positive TAs showed enrichment of transcripts involved in vascular remodeling, including smooth muscle cell migration. DISCUSSION: The detection of viral and immune activation pathways in GCA TAs supports a role for virus infection in GCA pathogenesis. In addition, the detection of viral pathways in control/VZV-positive TAs, along with vascular remodeling pathways, suggests that these samples may represent early infection with progression to clinical disease, depending on host and other environmental factors.

Amylin, Aß42, and Amyloid in Varicella Zoster Virus Vasculopathy Cerebrospinal Fluid and Infected Vascular Cells.

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: Aß 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; Aß40 was reduced and Aß42 unchanged. Intracellular amylin, Aß42, 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.

Acute zoster plasma contains elevated amyloid, correlating with Aß42 and amylin levels, and is amyloidogenic.

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, Aß40, Aß42, and amylin in 14 zoster and 10 control plasmas. Compared with controls, zoster plasma had significantly elevated amyloid that correlated with Aß42 and amylin levels and increased amyloid aggregation with addition of exogenous Aß42 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.

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

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-ß [Aß]) and amyloid, as well as extracellular amylin, Aß, 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 Aß42 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 Aß, and amyloid. No differences in extracellular amylin, Aß40, or Aß42 were detected, yet only supernatant from VZV-infected cells induced amylin aggregation and, to a lesser extent, Aß42 aggregation into amyloid fibrils. VZV glycoprotein B (gB) peptides assembled into fibrils and catalyzed amylin and Aß42 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.

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

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.

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

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.IMPORTANCE 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.

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

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)-1ß, 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-1ß 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-1ß. 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.

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

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.

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

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.

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.

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.

Varicella zoster virus triggers the immunopathology of giant cell arteritis.

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.

Varicella Zoster Virus Infection in Granulomatous Arteritis of the Aorta.

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.

Frequency of varicella zoster virus DNA in human adrenal glands.

Varicella zoster virus (VZV) becomes latent in ganglionic neurons derived from neural crest cells. Because the adrenal gland also contains medullary chromaffin cells of neural crest origin, we examined human adrenal glands and medullary chromaffin cell tumors (pheochromocytomas) for VZV and herpes simplex virus type 1 (HSV-1). We found VZV, but not HSV-1, DNA in 4/63 (6 %) normal adrenal glands. No VZV transcripts or antigens were detected in the 4 VZV DNA-positive samples. No VZV or HSV-1 DNA was found in 21 pheochromocytomas.

Varicella zoster virus and giant cell arteritis.

PURPOSE OF REVIEW: Giant cell arteritis (GCA) is a serious disease and the most common cause of vasculitis in the elderly. Here, studies describing the recent discovery of varicella zoster virus (VZV) in the temporal arteries of patients with GCA are reviewed. RECENT FINDINGS: GCA is characterized by severe headache/head pain and scalp tenderness. Many patients also have a history of vision loss, jaw claudication, polymyalgia rheumatica, fever, night sweats, weight loss, and fatigue. The erythrocyte sedimentation rate and C-reactive protein 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. The present 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 are pathologically negative for GCA. Virological analysis revealed that VZV is present in most GCA-positive and GCA-negative temporal artery biopsies, particularly in skip areas that correlate with adjacent GCA disease. SUMMARY: The presence of VZV in GCA-positive and GCA-negative temporal arteries reflects the possible role of VZV in triggering the immunopathology of GCA and indicates that both groups of patients should be treated with antivirals in addition to corticosteroids. Whether oral antiviral agents and steroids are as effective as intravenous acyclovir and steroids, and the dosage and duration of treatment, remain to be determined.

Analysis of Varicella-Zoster Virus in Temporal Arteries Biopsy Positive and Negative for Giant Cell Arteritis.

IMPORTANCE: Giant cell arteritis (GCA) is the most common systemic vasculitis in elderly individuals. Diagnosis is confirmed by temporal artery (TA) biopsy, although biopsy results are often negative. Despite the use of corticosteroids, disease may progress. Identification of causal agents will improve outcomes. Biopsy-positive GCA is associated with TA infection by varicella-zoster virus (VZV). OBJECTIVE: To analyze VZV infection in TAs of patients with clinically suspected GCA whose TAs were histopathologically negative and in normal TAs removed post mortem from age-matched individuals. DESIGN, SETTING, AND PARTICIPANTS: A cross-sectional study for VZV antigen was performed from January 2013 to March 2015 using archived, deidentified, formalin-fixed, paraffin-embedded GCA-negative, GCA-positive, and normal TAs (50 sections/TA) collected during the past 30 years. Regions adjacent to those containing VZV were examined by hematoxylin-eosin staining. Immunohistochemistry identified inflammatory cells and cell types around nerve bundles containing VZV. A combination of 17 tertiary referral centers and private practices worldwide contributed archived TAs from individuals older than 50 years. MAIN OUTCOMES AND MEASURES: Presence and distribution of VZV antigen in TAs and histopathological changes in sections adjacent to those containing VZV were confirmed by 2 independent readers. RESULTS: Varicella-zoster virus antigen was found in 45 of 70 GCA-negative TAs (64%), compared with 11 of 49 normal TAs (22%) (relative risk [RR] = 2.86; 95% CI, 1.75-5.31; P < .001). Extension of our earlier study revealed VZV antigen in 68 of 93 GCA-positive TAs (73%), compared with 11 of 49 normal TAs (22%) (RR = 3.26; 95% CI, 2.03-5.98; P < .001). Compared with normal TAs, VZV antigen was more likely to be present in the adventitia of both GCA-negative TAs (RR = 2.43; 95% CI, 1.82-3.41; P < .001) and GCA-positive TAs (RR = 2.03; 95% CI, 1.52-2.86; P < .001). Varicella-zoster virus antigen was frequently found in perineurial cells expressing claudin-1 around nerve bundles. Of 45 GCA-negative participants whose TAs contained VZV antigen, 1 had histopathological features characteristic of GCA, and 16 (36%) showed adventitial inflammation adjacent to viral antigen; no inflammation was seen in normal TAs. CONCLUSIONS AND RELEVANCE: In patients with clinically suspected GCA, prevalence of VZV in their TAs is similar independent of whether biopsy results are negative or positive pathologically. Antiviral treatment may confer additional benefit to patients with biopsy-negative GCA treated with corticosteroids, although the optimal antiviral regimen remains to be determined.

Adventitial fibroblasts induce a distinct proinflammatory/profibrotic macrophage phenotype in pulmonary hypertension.

Macrophage accumulation is not only a characteristic hallmark but is also a critical component of pulmonary artery remodeling associated with pulmonary hypertension (PH). However, the cellular and molecular mechanisms that drive vascular macrophage activation and their functional phenotype remain poorly defined. Using multiple levels of in vivo (bovine and rat models of hypoxia-induced PH, together with human tissue samples) and in vitro (primary mouse, rat, and bovine macrophages, human monocytes, and primary human and bovine fibroblasts) approaches, we observed that adventitial fibroblasts derived from hypertensive pulmonary arteries (bovine and human) regulate macrophage activation. These fibroblasts activate macrophages through paracrine IL-6 and STAT3, HIF1, and C/EBPß signaling to drive expression of genes previously implicated in chronic inflammation, tissue remodeling, and PH. This distinct fibroblast-activated macrophage phenotype was independent of IL-4/IL-13-STAT6 and TLR-MyD88 signaling. We found that genetic STAT3 haplodeficiency in macrophages attenuated macrophage activation, complete STAT3 deficiency increased macrophage activation through compensatory upregulation of STAT1 signaling, and deficiency in C/EBPß or HIF1 attenuated fibroblast-driven macrophage activation. These findings challenge the current paradigm of IL-4/IL-13-STAT6-mediated alternative macrophage activation as the sole driver of vascular remodeling in PH, and uncover a cross-talk between adventitial fibroblasts and macrophages in which paracrine IL-6-activated STAT3, HIF1α, and C/EBPß signaling are critical for macrophage activation and polarization. Thus, targeting IL-6 signaling in macrophages by completely inhibiting C/EBPß or HIF1α or by partially inhibiting STAT3 may hold therapeutic value for treatment of PH and other inflammatory conditions characterized by increased IL-6 and absent IL-4/IL-13 signaling.

Biopsy-negative, varicella zoster virus (VZV)-positive giant cell arteritis, zoster, VZV encephalitis and ischemic optic neuropathy, all in one.

A 72-year-old man developed clinical features of giant cell arteritis (GCA) and ipsilateral ophthalmic-distribution zoster, followed within 2 weeks by VZV encephalitis and 2 months later by ischemic optic neuropathy. Temporal artery biopsy was histopathologically negative for GCA, but contained VZV antigen and VZV DNA in multiple non-contiguous (skip) areas. The collective clinical and laboratory findings revealed a remarkably close temporal association of zoster, multifocal VZV vasculopathy with temporal artery infection, biopsy-negative VZV-positive GCA and VZV encephalitis.