Ocular Herpes Simplex Virus Infection Research Articles

Metabolic signatures of tear extracellular vesicles caused by herpes simplex keratitis

PurposeHerpes simplex keratitis (HSK), caused by type 1 herpes simplex virus (HSV) reactivation, is a severe infectious disease that leads to vision loss. HSV can trigger metabolic reprogramming in the host cell and change the extracellular vesicles (EV) cargos; however, little is known about the EV metabolic signatures during ocular HSV infection. Here, we aimed to depict the EV-associated metabolic landscape in HSK patients’ tears. MethodsWe collected 82 samples from 41 participants with unilateral HSK (contralateral unaffected tears were set as negative control), including subtype cohorts of 13 epithelial, 20 stromal, and 8 endothelial HSK. We isolated tear EVs via our previously established platform and conducted metabolic analysis using LC-MS/MS. The metabolic signatures for recognizing HSK and subtypes were assessed through differential analysis and machine learning algorithms. ResultsHypopsia and increased extracellular CD63 levels were observed in affected eyes. We identified 339 metabolites based on sEVs isolated from tears. Differential analysis revealed alterations in energy and amino acid metabolism, as well as the infectious microenvironment. Furthermore, we observed dysregulated metabolite such as methyldopa, which is associated with inappropriate neovascularization and corneal sensation loss, contributing to the HSK severity particularly in the stromal subtype. Moreover, machine learning classification also suggested a set of EV metabolic signatures that have potential for pan-keratitis detection. ConclusionsOur findings demonstrate that tear EV metabolites can serve as valuable indicators for comprehending the underlying pathological mechanisms. This knowledge is expected to facilitate the development of liquid biopsy means and therapeutic target discovery.

Whole exome sequencing of patients with varicella-zoster virus and herpes simplex virus induced acute retinal necrosis reveals rare disease-associated genetic variants.

Herpes simplex virus (HSV) and varicella-zoster virus (VZV) are neurotropic human alphaherpesviruses endemic worldwide. Upon primary infection, both viruses establish lifelong latency in neurons and reactivate intermittently to cause a variety of mild to severe diseases. Acute retinal necrosis (ARN) is a rare, sight-threatening eye disease induced by ocular VZV or HSV infection. The virus and host factors involved in ARN pathogenesis remain incompletely described. We hypothesize an underlying genetic defect in at least part of ARN cases. We collected blood from 17 patients with HSV-or VZV-induced ARN, isolated DNA and performed Whole Exome Sequencing by Illumina followed by analysis in Varseq with criteria of CADD score > 15 and frequency in GnomAD < 0.1% combined with biological filters. Gene modifications relative to healthy control genomes were filtered according to high quality and read-depth, low frequency, high deleteriousness predictions and biological relevance. We identified a total of 50 potentially disease-causing genetic variants, including missense, frameshift and splice site variants and on in-frame deletion in 16 of the 17 patients. The vast majority of these genes are involved in innate immunity, followed by adaptive immunity, autophagy, and apoptosis; in several instances variants within a given gene or pathway was identified in several patients. We propose that the identified variants may contribute to insufficient viral control and increased necrosis ocular disease presentation in the patients and serve as a knowledge base and starting point for the development of improved diagnostic, prophylactic, and therapeutic applications.

Neutrophil Extracellular Traps and Their Possible Implications in Ocular Herpes Infection.

Neutrophil extracellular traps (NETs) are net-like structures released from neutrophils. NETs predominantly contain cell-free deoxyribonucleic acid (DNA) decorated with histones and neutrophil granule proteins. Numerous extrinsic and intrinsic stimuli can induce the formation of NETs such as pathogens, cytokines, immune complexes, microcrystals, antibodies, and other physiological stimuli. The mechanism of NETosis induction can either be ROS-dependent or independent based on the catalase producing activity of the pathogen. NADPH is the source of ROS production, which in turn depends on the upregulation of Ca2+ production in the cytoplasm. ROS-independent induction of NETosis is regulated through toll-like receptors (TLRs). Besides capturing and eliminating pathogens, NETs also aggravate the inflammatory response and thus act as a double-edged sword. Currently, there are growing reports of NETosis induction during bacterial and fungal ocular infections leading to different pathologies, but there is no direct report suggesting its role during herpes simplex virus (HSV) infection. There are innumerable independent reports showing that the major effectors of NETosis are also directly affected by HSV infection, and thus, there is a strong possibility that HSV interacts with these facilitators that can either result in virally mediated modulation of NETosis or NETosis-mediated suppression of ocular HSV infection. This review focuses on the mechanism of NETs formation during different ocular pathologies, with its prime focus on highlighting their potential implications during HSV ocular infections and acting as prospective targets for the treatment of ocular diseases.

Herpes Simplex Virus Keratitis.

Herpes simplex virus (HSV) is a highly prevalent infection in the United States. One complication of HSV is HSV keratitis, an ocular HSV infection thought to be the leading cause of corneal blindness in the United States. Home care clinicians with knowledge of the signs and symptoms of HSV and HSV keratitis can aid in early detection and treatment of this potentially serious infection. This article discusses signs and symptoms of HSV keratitis, preventive measures, and treatment.

Blockade of PD-1 and LAG-3 Immune Checkpoints Combined with Vaccination Restores the Function of Antiviral Tissue-Resident CD8+ TRM Cells and Reduces Ocular Herpes Simplex Infection and Disease in HLA Transgenic Rabbits.

Chronic viruses such as herpes simplex virus 1 (HSV-1) evade the hosts' immune system by inducing the exhaustion of antiviral T cells. In the present study, we found that exhausted HSV-specific CD8+ T cells, with elevated expression of programmed death ligand-1 (PD-1) and lymphocyte activation gene-3 (LAG-3) receptors were frequent in symptomatic patients, with a history of numerous episodes of recurrent corneal herpetic disease, compared to asymptomatic patients who never had corneal herpetic disease. Subsequently, using a rabbit model of recurrent ocular herpes, we found that the combined blockade of PD-1 and LAG-3 pathways with antagonist antibodies significantly restored the function of tissue-resident antiviral CD8+ TRM cells in both the cornea and the trigeminal ganglia (TG). An increased number of functional tissue-resident HSV-specific CD8+ TRM cells in latently infected rabbits was associated with protection against recurrent herpes infection and disease. Compared to the PD-1 or LAG-3 blockade alone, the combined blockade of PD-1 and LAG-3 appeared to have a synergistic effect in generating frequent polyfunctional Ki-67+, IFN-γ+, CD107+, and CD8+ T cells. Moreover, using the human leukocyte antigen (HLA) transgenic rabbit model, we found that dual blockade of PD-1 and LAG-3 reinforced the effect of a multiepitope vaccine in boosting the frequency of HSV-1-specific CD8+ TRM cells and reducing disease severity. Thus, both the PD-1 and the LAG-3 exhaustion pathways play a fundamental role in ocular herpes T cell immunopathology and provide important immune checkpoint targets to combat ocular herpes.IMPORTANCE HSV-specific tissue-resident memory CD8+ TRM cells play a critical role in preventing virus reactivation from latently infected TG and subsequent virus shedding in tears that trigger the recurrent corneal herpetic disease. In this report, we determined how the dual blockade of PD-1 and LAG-3 immune checkpoints, combined with vaccination, improved the function of CD8+ TRM cells associated with a significant reduction in recurrent ocular herpes in HLA transgenic (Tg) rabbit model. The combined blockade of PD-1 and LAG-3 appeared to have a synergistic effect in generating frequent polyfunctional CD8+ TRM cells that infiltrated both the cornea and the TG. The preclinical findings using the established HLA Tg rabbit model of recurrent herpes highlight that blocking immune checkpoints combined with a T cell-based vaccine would provide an important strategy to combat recurrent ocular herpes in the clinic.

Blockade of LAG-3 Immune Checkpoint Combined With Therapeutic Vaccination Restore the Function of Tissue-Resident Anti-viral CD8+ T Cells and Protect Against Recurrent Ocular Herpes Simplex Infection and Disease.

Recurrent viral diseases often occur after the viruses evade the hosts' immune system, by inducing exhaustion of antiviral T cells. In the present study, we found that functionally exhausted herpes simplex virus type 1 (HSV-1) -specific CD8+ T cells, with elevated expression of lymphocyte activation gene-3 (LAG-3), an immune checkpoint receptor that promotes T cell exhaustion, were frequent in symptomatic (SYMP) patients with a history of numerous episodes of recurrent corneal herpetic disease. Similarly, following UV-B induced virus reactivation from latency the symptomatic wild-type (WT) B6 mice that developed increase virus shedding and severe recurrent corneal herpetic disease had more exhausted HSV-specific LAG-3+CD8+ T cells in both trigeminal ganglia (TG) and cornea. Moreover, a therapeutic blockade of LAG-3 immune checkpoint with antagonist antibodies combined with a therapeutic immunization with gB498−505 peptide immunodominant epitope of latently infected B6 mice significantly restored the quality and quantity of functional HSV-1 gB498−505 specific CD8+ T cells in both TG and cornea and protected against UV-B induced recurrent corneal herpes infection and disease. In contrast to dysfunctional HSV-specific CD8+ T cells from WT B6 mice, more functional HSV-specific CD8+ T cells were detected in LAG-3−/− deficient mice and were associated with less UV-B induced recurrent corneal herpetic disease. Thus, the LAG-3 pathway plays a fundamental role in ocular herpes T cell immunopathology and provides an important immune checkpoint target that can synergizes with T cell-based therapeutic vaccines against symptomatic recurrent ocular herpes.

A Therapeutic Antiviral Antibody Inhibits the Anterograde Directed Neuron-to-Cell Spread of Herpes Simplex Virus and Protects against Ocular Disease

Herpes simplex virus (HSV) is a leading cause of blindness and viral encephalitis in the developed world. Upon reactivation from sensory neurons, HSV returns via axonal transport to peripheral tissues where it causes, e.g., severe, potentially blinding ocular diseases. In the present study we investigated whether the HSV-1/2 glycoprotein B-specific antibody mAb 2c or its humanized counterpart mAb hu2c can protect from ocular disease in a mouse model of HSV-1-induced acute retinal necrosis (ARN). In this model the viral spread from the initially infected to the contralateral eye resembles the routes taken in humans upon HSV reactivation. Systemic antibody treatment prior or early after infection effectively protected the mice from the development of ARN. These observations suggest that the antibody potently neutralized the infection and inhibited the viral transmission, since there was almost no virus detectable in the contralateral eyes and trigeminal ganglia of antibody treated mice. Besides of neutralizing free virus or limiting the infection via activating the complement or cellular effector functions, blocking of the anterograde directed neuron-to-cell spread of HSV represents a viable mode of action how mAb 2c protected the mice from ARN. We proved this hypothesis using a microfluidic chamber system. Neurons and epithelial cells were cultured in two separate compartments where the neurons sent axons via connecting microgrooves to the epithelial cells. Neurons were infected with a reporter HSV-1 strain expressing mCherry, and the co-culture was treated with neutralizing antibodies. In contrast to commercial polyclonal human HSV-neutralizing immunoglobulins, mAb 2c effectively blocked the anterograde directed neuron-to-cell transmission of the virus. Our data suggest that the humanized HSV-1/2-gB antibody protects mice from ocular disease by blocking the neuronal spread of HSV. Therefore, mAb hu2c may become a potent novel therapeutic option for severe ocular HSV infections.

Antibody-based immunotherapy of aciclovir resistant ocular herpes simplex virus infections

The increasing incidence of aciclovir- (ACV) resistant strains in patients with ocular herpes simplex virus (HSV) infections is a major health problem in industrialized countries. In the present study, the humanized monoclonal antibody (mAb) hu2c targeting the HSV-1/2 glycoprotein B was examined for its efficacy towards ACV-resistant infections of the eye in the mouse model of acute retinal necrosis (ARN). BALB/c mice were infected by microinjection of an ACV-resistant clinical isolate into the anterior eye chamber to induce ARN and systemically treated with mAb hu2c at 24h prior (pre-exposure prophylaxis) or at 24, 40, and 56h after infection (post-exposure immunotherapy). Mock treated controls and ACV-treated mice showed pronounced retinal damage. Mice treated with mAb hu2c were almost completely protected from developing ARN. In conclusion, mAb hu2c may become a reliable therapeutic option for drug/ACV-resistant ocular HSV infections in humans in order to prevent blindness.

The role of pro‐resolution lipid mediators in infectious disease

Inflammation is an essential host defence against infection, but can be damaging when excessive. Resolution of inflammation is an active process, and the pro-resolution effects of lipoxins, resolvins and protectins have received significant interest. Here, we review emerging data on the role of these lipid mediators in infectious disease. Lipoxins influence host control of Mycobacterium tuberculosis, Toxoplasma gondii, Trypanosoma cruzi and Plasmodium berghei cerebral malaria in mice. Their effects are protective in toxoplasmosis, T. cruzi infection and cerebral malaria but detrimental in tuberculosis; related to the balance between pathogen-control and excessive immune response. Topical lipoxin abrogates the tissue damage seen in a rabbit model of Porphyromonas gingivalis periodontitis. The increased virulence of H5N1 influenza A virus in mice correlates with reduced expression of SOCS2, required to mediate the effects of lipoxin. Mice unable to synthesize lipoxin suffer increased lung pathology during respiratory syncytial virus infection. Protectin suppresses influenza A virus replication in vitro and increases survival in a mouse model of severe influenza infection. Resolvins were investigated in a number of animal models of systemic bacterial infection, and were found to enhance phagocytic clearance of bacteria, reduce inflammation severity, promote neutrophil apoptosis, modulate neutrophil chemotaxis and importantly, reduce mortality. Interestingly, resolvin also enhances the antibacterial effect of ciprofloxacin and vancomycin. Topical resolvin application reduces the severity of herpes simplex virus ocular infection in mice. If the effects of these mediators translate from pre-clinical studies into successful clinical trials, they represent promising new strategies in managing infectious disease.

Design of xanthone propionate photolabile protecting group releasing acyclovir for the treatment of ocular herpes simplex virus

We have attached the antiviral drug acyclovir (ACV) to a xanthone photolabile protecting group (or photocage) through the O6 position of acyclovir, a procedure designed for the treatment of ocular herpes simplex virus infections. Acyclovir is photoreleased from the photocage, under physiological conditions, with a quantum yield (Φ(ACVrelease)) of 0.1-0.3 and an uncaging cross section (Φ·ε) of 450-1350 M cm(-1). We demonstrate that this photorelease method outcompetes alternative reaction pathways, such as protonation. Furthermore, complete release of the drug is theoretically possible given a sufficient dose of light . Surprisingly the acyclovir photocage, also showed some antiviral activity towards HSV-1.

Ganciclovir Gel—A New Topical Treatment for Herpetic Keratitis

Herpes simplex virus (HSV) infection of the eye is a major cause of corneal opacity in the US 1 and other developed countries. 2 Infection with either HSV-1 or HSV-2 is common. Data from the US National Health and Nutrition Examination Survey (NHANES) suggest an overall 58% seroprevalence rate for HSV-1—the predominant cause of herpes simplex keratitis 2 (HSK)—with rates in individual ethnic groups as high as 89%. 3 A three-month prospective epidemiological study conducted in France put the incidence of HSK at 31.5 cases per 100,000 per person-year. 4 Of these, 13.2 cases represented primary infections and 18.3 were recurrences. HSV infection is lifelong and recurrent, with the virus becoming latent in sensory nerves after the original outbreak. Estimates in the US indicate approximately 20,000 new cases of HSK each year with another 28,000 cases of recurrent disease. 5 The only currently available topical treatment for HSK in the US is trifluridine (TFT), a drug whose toxic effects highlight the need for more effective and less harmful treatments. This article will summarize HSV ocular infection and review current treatments approved both in the US and worldwide. It will also focus on efficacy and safety findings for ganciclovir 0.15% gel, an antiviral ophthalmic preparation available outside the US to treat HSK that is a promising new drug for the US market.

Controlling Viral Immuno-Inflammatory Lesions by Modulating Aryl Hydrocarbon Receptor Signaling

Ocular herpes simplex virus infection can cause a blinding CD4+ T cell orchestrated immuno-inflammatory lesion in the cornea called Stromal Keratitis (SK). A key to controlling the severity of SK lesions is to suppress the activity of T cells that orchestrate lesions and enhance the representation of regulatory cells that inhibit effector cell function. In this report we show that a single administration of TCDD (2, 3, 7, 8- Tetrachlorodibenzo-p-dioxin), a non-physiological ligand for the AhR receptor, was an effective means of reducing the severity of SK lesions. It acted by causing apoptosis of Foxp3- CD4+ T cells but had no effect on Foxp3+ CD4+ Tregs. TCDD also decreased the proliferation of Foxp3- CD4+ T cells. The consequence was an increase in the ratio of Tregs to T effectors which likely accounted for the reduced inflammatory responses. In addition, in vitro studies revealed that TCDD addition to anti-CD3/CD28 stimulated naïve CD4+ T cells caused a significant induction of Tregs, but inhibited the differentiation of Th1 and Th17 cells. Since a single TCDD administration given after the disease process had been initiated generated long lasting anti-inflammatory effects, the approach holds promise as a therapeutic means of controlling virus induced inflammatory lesions.

Primary ocular herpes simplex virus infection with zosteriform spreading, accompanied with meningism

Primary ocular herpes simplex virus infection with zosteriform spreading, accompanied with meningism

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A 3,545-g, 35–3/7 weeks', large for gestational age (LGA) male is delivered to an insulin-dependent diabetic mother who became pregnant with the help of a fertility specialist. After conception, a general obstetrician managed her pregnancy. The antenatal course was uneventful except for a history of herpes simplex virus (HSV) infection. A cesarean section was planned at 38 weeks' gestation because of a relatively large fetus and a small pelvic outlet. The mother developed premature rupture of membranes (PROM) at 35 weeks' gestation, and 48 hours later, labor began. The obstetrician was out of town, and the mother was triaged to a midwife for the delivery. The mother's last HSV outbreak was 2 weeks before delivery, with lesions healing 4 days before delivery. The mother claimed in her deposition that she told the midwife about her history of herpes. The medical records from the fertility specialist and the obstetrician documented the history of HSV. The medical records reflecting the herpes were computerized and available to the midwife and the nurses who managed labor and delivery. The infant was delivered by a spontaneous vaginal vertex delivery. The Apgar scores were 8 at 1 minute and 9 at 5 minutes. The infant was asymptomatic and observed for 6 hours before rooming-in with his mother. Multiple bedside glucose evaluations were read as normal. A complete blood count (CBC) yielded unremarkable results. A blood culture was reported as negative on the final reading. Vital signs were stable. The infant was sleepy at the breast, but the mother fed him formula. The discharge examination was performed at 41 hours, and the only notable finding besides the LGA and late preterm gestation was that an ophthalmic examination could not be performed because the eyes were closed. The grandmother stated in her deposition that the baby's eyes were …

Cell culture isolation can miss the laboratory diagnosis of HSV ocular infection.

We compared polymerase chain reaction (PCR) to cell culture isolation for the laboratory diagnosis of ocular herpes simplex virus (HSV) disease. Laboratory and medical records of consecutive patients were reviewed for results of 1) HSV PCR testing, 2) HSV cell culture isolation, and 3) clinical diagnosis. PCR results were statistically compared to cell culture isolation and patients initially diagnosed for ocular HSV infection. Of 581 cases submitted for laboratory testing, 520 were PCR negative, cell culture negative (89.6%); 0 were PCR negative, cell culture positive (0%); 27 were PCR positive, cell culture negative (4.6%); and 34 were PCR positive, cell culture positive (5.8%). PCR tested more positive than cell culture isolation (McNemar's, P=0.0001). Of 47 HSV PCR positive cases with complete medical records, 19 were cell culture negative for HSV and 28 were cell culture positive for HSV. Fourteen of 19 cell culture negative cases (74%) (Without PCR, 5 cases of HSV would be missed) and 25 of the 28 cell culture positive cases (89%) (Laboratory testing was necessary for diagnosing 3 cases) were clinically diagnosed with HSV at the initial examination. PCR was a more definitive test for diagnosing HSV ocular infection than cell culture isolation. Cell culture isolation alone can miss an atypical presentation of HSV ocular infection.

Recognition and treatment of herpes simplex keratitis

AbstractAlthough the incidence of ocular herpes simplex virus infection has not diminished in recent years, we are encountering fewer severe problems such as necrotizing stromal keratitis and diffuse disciform keratitis which may necessitate penetrating keratoplasty. The earlier recognition of herpes simplex viral keratitis and the better management utilizing oral antivirals and topical corticosteroids is probably responsible for this improved outlook of herpes simplex keratitis. Early recognition and management of acute and chronic herpetic keratitis, as well as the management of keratoplasty for herpes will be reviewed.

Homeostatic expansion of CD4 + T cells upregulates VLA-4 and exacerbates HSV-induced corneal immunopathology

Using a viral-induced immunopathology model, we showed that when CD4 + T cells were allowed to undergo homeostatic expansion prior to ocular herpes simplex virus infection, mice developed more severe inflammatory lesions with the increased severity associated with enhanced effector function of ocular CD4 + T cells, and blocking their functional activity reduced the lesion severity. Additionally, homeostatically expanded CD4 + T cells upregulated VLA-4, and in vivo administration of anti-VLA-4 mAb significantly decreased the homeostatic proliferation. Furthermore, blocking of VLA-4 interaction also diminished the infiltration of CD4 + T cells into the cornea and decreased lesion severity. Our results imply that homeostatic expansion of T cells, as could occur in a virus-induced lymphopenia, may generate cells with enhanced effector function that can contribute to tissue damage.

OCULAR HERPES SIMPLEX INFECTIONS IN CHILDREN

A survey of "hospitalised" Herpes simplex infections in otherwise normal healthy children under the age of 13 years over a 15 year period between 1964 and 1979 at Princess Margaret Hospital for Children. This survey highlighted the unnecessarily high transference rate of Herpes simplex virus (HSV) infection to the under 3 year old within the home environment. There were three hundred and forty (340) cases of non-fatal "hospitalized" HSV infections in the under 3 year old of which sixty three (63) were ocular infections. Over all age groups one hundred and forty five (145) children were hospitalised on one or several occasions with ocular HSV infection. Of these children before reaching the age of 13 years, fifteen (15) developed intraocular HSV infection and twenty seven (27) had a permanent visual acuity of less than 6/12. As a result of this survey the Princess Margaret Hospital motto for the I.Y.D.P. is: "Cuddle but do not kiss your child when you have an acute coldsore"

Primary Conjunctival Herpetic Geographic Ulcer in an Immunocompetent Patient

Herpes simplex virus ocular infections have been recognized as a major cause of corneal blindness in the developed world. The major signs of a primary infection are limited to the lids, conjunctiva, and cornea. Presence of a conjunctival dendrite without corneal involvement has been reported. Although conjunctival ulceration has been associated with recurrent attacks, it has not been documented as a manifestation of primary herpetic infection. We report a case of primary herpetic infection with geographic conjunctival ulcer with multiple corneal dendrites.

IL-10 and natural regulatory T cells: two independent anti inflammatory mechanisms in HSV induced ocular immonopathology (B63)

Abstract Our past studies have shown a protective role for both CD4+CD25+Foxp3+ natural regulatory T cells (nTregs) as well as IL-10 in Herpes Simplex Virus (HSV) induced Stromal Keratitis (SK). Among several other mechanisms, IL-10 has been shown to be involved in the nTreg mediated suppression. We have shown in vitro that blocking of IL-10 could partially block the Treg mediated suppression of activated effectors isolated from SK lesions. But it is not known whether in the in vivo conditions, IL-10 and nTreg influence the SK pathology independently or in concert. Interestingly, IL-10−/− animals depleted of nTregs prior to ocular HSV infection had more severe SK lesion as compared to undepleted IL-10−/− animals. These results show the presence of two independent anti inflammatory mechanisms in SK. Along this line, by using mice expressing GFP-Foxp3 we could not show CD4+GFP-Foxp3+IL-10+ cells in the DLNs and spleens of infected wild-type (WT) mice although animals depleted of nTregs had higher levels of IL-10 in the DLNs post infection. Furthermore, by using WT, IL-10−/− and nTreg depleted WT and IL-10−/− animals we have compared the relative importance of IL-10 and nTregs in reducing SK pathology. Although disease severity was significantly higher in IL-10−/− and both the nTreg depleted groups as compared to WT, highest severity was found in the IL-10−/− mice depleted of nTregs. Intracellular cytokine staining with UV-HSV and polyclonal stimulation of the cells isolated from infected corneas, DLN and spleens revealed that IL-10 has more profound anti inflammatory effects in SK as compared to nTregs and that the later could act in an IL-10 independent manner.