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.

No Evidence of Varicella-Zoster Virus Infection in Temporal Artery Biopsies of Anterior Ischemic Optic Neuropathy Patients With and Without Giant Cell Arteritis.

BACKGROUND: To test the hypothesis that varicella-zoster virus (VZV) infection contributes to temporal arteritis pathogenesis, comprehensive in situ analysis was performed on temporal artery biopsies of 38 anterior ischemic optic neuropathy (AION) patients, including 14 (37%) with giant cell arteritis. METHODS: Biopsies were completely sectioned, and, on average, 146 serial sections per patient were stained for VZV glycoprotein E. RESULTS: Four of 38 AION patients showed VZV glycoprotein E staining, but VZV infection was not confirmed by staining for VZV IE63 protein and VZV-specific polymerase chain reaction on adjacent sections. CONCLUSIONS: This study refutes the premise that VZV is casually related to AION with and without giant cell arteritis.

Assessment for varicella zoster virus in patients newly suspected of having giant cell arteritis.

OBJECTIVES: There is uncertainty if varicella zoster virus (VZV) triggers GCA. This is based on discordant reports of VZV detection in GCA temporal artery biopsies. We conducted a multimodal evaluation for VZV in the inception Giant Cell Arteritis and PET Scan (GAPS) cohort. METHODS: Consecutive patients who underwent temporal artery biopsy for suspected GCA were clinically reviewed for active and past VZV infection and followed for 6 months. Serum was tested for VZV IgM and IgG. Temporal artery biopsy (TAB) sections were stained for VZV antigen using the VZV Mouse Cocktail Antibody (Cell Marque, Rocklin, CA, USA). A selection of GCA and control tissues were stained with the VZV gE antibody (Santa Cruz Biotechnology, Dallas, TX, USA), which was used in previous studies. RESULTS: A total of 58 patients met inclusion criteria, 12 (21%) had biopsy-positive GCA and 20 had clinically positive GCA. None had herpes zoster at enrolment and only one patient developed a VZV clinical syndrome (zoster ophthalmicus) on follow-up. There was no difference in VZV exposure between GCA and non-GCA patients. None of the 53 patients who had VZV serology collected had positive VZV IgM antibodies. VZV antigen was not convincingly demonstrated in any of the TAB specimens; 57 TABs stained negative and 1 stained equivocally positive. The Santa Cruz Biotechnology VZV antibody exhibited positive staining in a range of negative control tissues, questioning its specificity for VZV antigen. CONCLUSION: The absence of active infection markers argues against VZV reactivation being the trigger for GCA. Non-specific immunohistochemistry staining may account for positive findings in previous studies.

Investigating the Association of Giant Cell Arteritis with Varicella Zoster Virus in Temporal Artery Biopsies or Ascending Aortic Resections.

OBJECTIVE: A variety of infectious agents, including varicella zoster virus (VZV), have been hypothesized to play a role in the pathogenesis of giant cell arteritis (GCA). The detectability of the virus in patients with GCA is debatable. To further investigate an association between GCA and VZV infection, 10 years of GCA cases were evaluated for VZV by immunohistochemistry (IHC). METHODS: All temporal artery biopsies and ascending aortic resections positive for GCA from 2007 to 2017 at Brigham and Women's Hospital were immunostained using a VZV antibody cocktail (SG1-1, SG1-SG4, NCP-1, and IE-62). RESULTS: Forty-one temporal artery biopsies and 47 ascending aortic resections positive for GCA were identified, all of which were found to be negative for VZV by IHC. Twelve temporal artery biopsies in this cohort were previously analyzed by unbiased metagenomics sequencing and were negative for VZV DNA. CONCLUSION: These results argue against a clinically relevant association between VZV infection and GCA, and support neither routine testing for VZV nor treatment with antiviral drugs.

Identification of Herpes Zoster-Associated Temporal Arteritis Among Cases of Giant Cell Arteritis.

PURPOSE: To examine whether herpes zoster antigen (also called varicella-zoster virus antigen) was detectable in temporal artery biopsies taken from individuals with giant cell arteritis (GCA). DESIGN: Retrospective comparative case series. METHODS: Sections of formalin-fixed paraffin-embedded temporal arteries were examined first by hematoxylin-eosin (H&E) staining to establish the diagnosis of GCA. Adjacent sections of the same biopsy were then examined by immunohistochemistry, using 2 different monoclonal antibodies against a major antigen of varicella-zoster virus called gE. Pathologic specimens were obtained from patients cared for at the University of Iowa and Washington University in St. Louis ophthalmology clinics. RESULTS: The study included biopsies from 25 patients with symptoms of GCA as well as positive H&E pathology and 25 patients with symptoms compatible with GCA but negative H&E pathology. Among the GCA-positive group, 3 patients had positive staining for herpes zoster antigen. Among the GCA-negative group, herpes zoster antigen was not detected in any biopsy. In both groups of patients, false-positive staining for herpes zoster antigen was detected in the presence of calcifications in the arteries. False-positive staining was also detected on some extra-arterial skeletal muscle and erythrocytes. CONCLUSION: Herpes zoster antigen was detected in 3 of 25 temporal arteries from patients with biopsy-proven GCA. One of the 3 positive cases was noteworthy because the patient had had herpes zoster ophthalmicus diagnosed 3 weeks before the onset of GCA symptoms. False-positive staining for herpes zoster antigen was detected on several temporal artery biopsies.

Herpes Zoster as a Risk Factor for Incident Giant Cell Arteritis.

OBJECTIVE: Histopathologic studies have implicated herpes zoster (HZ) as a causative organism of giant cell arteritis (GCA). The purpose of this study was to assess the epidemiologic association of HZ events with incident GCA. METHODS: We performed a retrospective cohort study in 2 large independent US administrative data sets: Medicare 5% and Truven Health Analytics MarketScan. Eligible subjects had 12 months of continuous coverage, were >50 years old, and had no history of GCA or polymyalgia rheumatica. HZ events (complicated and uncomplicated) and GCA were identified by the presence of International Classification of Diseases, Ninth Revision, Clinical Modification codes from physician visit or hospital discharge records. Antiviral therapies and vaccinations were identified from prescription claims and drug codes. Risk of incident GCA was calculated using multivariable Cox proportional hazards regression. RESULTS: Among 16,686,345 subjects, a total of 5,942 GCA cases occurred, with 3.1% (MarketScan) and 6.0% (Medicare) having preceding HZ events. Unadjusted GCA incidence rates were highest in the groups with complicated and uncomplicated HZ. After multivariable adjustment, complicated HZ was associated with an increased risk of GCA (hazard ratio [HR] 1.99 [95% confidence interval (95% CI) 1.32-3.02] in the Medicare cohort and 2.16 [95% CI 1.46-3.18] in the MarketScan cohort), as was uncomplicated HZ (HR 1.42 [95% CI 1.02-1.99] and HR 1.45 [95% CI 1.05-2.01] in the respective cohorts). Vaccination and antiviral treatment were not consistently associated with GCA risk, although antiviral treatment was marginally associated with a decreased risk of GCA in the Medicare cohort (HR 0.67 [95% CI 0.46-0.99]). CONCLUSION: HZ is associated with an increased risk of GCA. The infrequency of HZ in GCA patients suggests that it is only one potential trigger for GCA. Antivirals and vaccination did not consistently mitigate this risk.

No detection of varicella-zoster virus in temporal arteries of patients with giant cell arteritis.

OBJECTIVE: Data on the presence of varicella-zoster virus (VZV) in temporal arteries of patients with giant cell arteritis (GCA) are controversial. We analyzed VZV infection in temporal arteries from Italian patients with temporal artery biopsy (TAB)-positive GCA, TAB-negative GCA, and controls. METHODS: A total of 79 formalin-fixed, paraffin-embedded (FFPE) TABs performed between 2009 and 2012 at a single institution from 34 TAB-positive GCA patients, 15 TAB-negative GCA patients, and 30 controls were retrieved. Six 5-µm sections of all FFPE TABs were cut. The first section was analyzed by immunohistochemistry using mouse monoclonal anti-VZVgE IgG1 antibody. DNA was extracted from the remaining five sections and analyzed by real-time polymerase chain reaction (PCR) for the presence of VZV DNA. For 10 of the 34 TAB-positive GCA patients, an additional 2-mm piece of frozen TAB was available. DNA was extracted from the entire 2-mm length frozen specimen and analyzed by PCR for the presence of VZV DNA. Thirty additional 5-µm sections were cut from the FFPE TABs of these 10 patients and analyzed by immunohistochemistry for the presence of VZV antigen. RESULTS: Immunohistochemical analysis detected VZV antigen in 1/34 (3%) TAB-positive GCA, 0/15 TAB-negative GCA, and 0/30 controls, and in none of the 300 sections cut from the 10 FFPE TABs positive for GCA for which the frozen specimens were available. DNA obtained from all TABs was amplifiable. VZV DNA was neither found in any of the FFPE TABs nor found in frozen TABs. CONCLUSION: Our data do not support in Italian patients a possible role for VZV infection in the etiopathogenesis of GCA.

Giant cell arteritis following varicella zoster vaccination.

BACKGROUND: Giant cell arteritis (GCA) is a form of large-vessel vasculitis affecting patients older than 50years old. Recent reports have suggested that the condition is caused by reactivation of varicella zoster virus (VZV). If that is indeed the case, a vaccine that prevents VZV reactivation should reduce the incidence of GCA. To further test that hypothesis, we assessed the incidence of GCA among patients older than 50years of age who were vaccinated against VZV and compared it to GCA incidence in the general population. METHODS: Using a centralized electronic medical database of the major medical insurer in Israel, Clalit Health Services (CHS), we have calculated the incidence of newly-diagnosed GCA patients in the general population, as well as the incidence of GCA among patients previously vaccinated against VZV between 1.1 2014-31.10.2016. RESULTS: The mean incidence of newly-diagnosed GCA among non-vaccinated patients older than 50years of age was 41.6/100,000/year, while the mean incidence of GCA among patients previously vaccinated against VZV in the same time period was 75.2/100,000/year (P=0.07). CONCLUSIONS: These findings do not support the hypothesis that GCA is due to VZV reactivation.

Varicella Zoster Virus: A Common Cause of Stroke in Children and Adults.

BACKGROUND: Varicella zoster virus (VZV) is a neurotropic, exclusively human herpesvirus. Primary infection causes varicella (chickenpox), after which the virus becomes latent in ganglionic neurons along the entire neuraxis. As cell-mediated immunity to VZV declines with advancing age and immunosuppression, VZV reactivates to produce zoster (shingles). One of the most serious complications of zoster is VZV vasculopathy. METHODS: We reviewed recent studies of stroke associated with varicella and zoster, how VZV vasculopathy is verified virologically, vaccination to prevent varicella and immunization to prevent zoster, and VZV in giant cell arteritis (GCA). FINDINGS: We report recent epidemiological studies revealing an increased risk of stroke after zoster; the clinical, laboratory, and imaging features of VZV vasculopathy; that VZV vasculopathy is confirmed by the presence of either VZV DNA or anti-VZV IgG antibody in cerebrospinal fluid; special features of VZV vasculopathy in children; vaccination to prevent varicella and immunization to prevent zoster; and the latest evidence linking VZV to GCA. CONCLUSION: In children and adults, VZV is a common cause of stroke.

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.

Issues in the Treatment of Neurological Conditions Caused by Reactivation of Varicella Zoster Virus (VZV).

Varicella zoster virus (VZV) is a ubiquitous neurotropic human herpesvirus. Primary infection usually causes varicella (chicken pox), after which virus becomes latent in ganglia along the entire neuraxis. Decades later, virus reactivates to produce herpes zoster (shingles), a painful dermatomally distributed vesicular eruption. Zoster may be further complicated by postherpetic neuralgia, VZV vasculopathy, myelitis, and segmental motor weakness. VZV reactivation has also been associated with giant cell arteritis. This overview discusses treatment of various conditions that often require both corticosteroids and antiviral drugs. Treatment for VZV-associated disease is often based on case reports and small studies rather than large-scale clinical trials. Issues that require resolution include the optimal duration of such combined therapy, more effective treatment for postherpetic neuralgia, whether some treatments should be given orally or intravenously, the widening spectrum of zoster sine herpete, and the role of antiviral therapy in giant cell arteritis.

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

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.

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.

Varicella Zoster Virus in Temporal Arteries of Patients With Giant Cell Arteritis.

Giant cell arteritis (GCA) is an immune-mediated disease of unknown etiology. Varicella zoster virus (VZV) antigen was found in all of 4 GCA-positive temporal arteries (TAs) but was not present in any of 13 normal TAs. All 4 GCA-positive TAs contained viral antigen in skip areas, mostly in the adventitia and media and least in the intima. Despite formalin fixation, VZV DNA was detected in 2 of 4 GCA-positive, VZV antigen-positive TAs. Skeletal muscle was attached to 3 of 4 TAs, and VZV antigen was found in 2 and VZV DNA in 1. VZV may cause GCA.

Prevalence and distribution of VZV in temporal arteries of patients with giant cell arteritis.

OBJECTIVE: Varicella-zoster virus (VZV) infection may trigger the inflammatory cascade that characterizes giant cell arteritis (GCA). METHODS: Formalin-fixed, paraffin-embedded GCA-positive temporal artery (TA) biopsies (50 sections/TA) including adjacent skeletal muscle and normal TAs obtained postmortem from subjects >50 years of age were examined by immunohistochemistry for presence and distribution of VZV antigen and by ultrastructural examination for virions. Adjacent regions were examined by hematoxylin & eosin staining. VZV antigen-positive slides were analyzed by PCR for VZV DNA. RESULTS: VZV antigen was found in 61/82 (74%) GCA-positive TAs compared with 1/13 (8%) normal TAs (p < 0.0001, relative risk 9.67, 95% confidence interval 1.46, 63.69). Most GCA-positive TAs contained viral antigen in skip areas. VZV antigen was present mostly in adventitia, followed by media and intima. VZV antigen was found in 12/32 (38%) skeletal muscles adjacent to VZV antigen-positive TAs. Despite formalin fixation, VZV DNA was detected in 18/45 (40%) GCA-positive VZV antigen-positive TAs, in 6/10 (60%) VZV antigen-positive skeletal muscles, and in one VZV antigen-positive normal TA. Varicella-zoster virions were found in a GCA-positive TA. In sections adjacent to those containing VZV, GCA pathology was seen in 89% of GCA-positive TAs but in none of 18 adjacent sections from normal TAs. CONCLUSIONS: Most GCA-positive TAs contained VZV in skip areas that correlated with adjacent GCA pathology, supporting the hypothesis that VZV triggers GCA immunopathology. Antiviral treatment may confer additional benefit to patients with GCA treated with corticosteroids, although the optimal antiviral regimen remains to be determined.