Ocular Infection of HCMV: Immunology, Pathogenesis, and Interventions

Abstract

Human cytomegalovirus (HCMV) retinitis accounts for 70% of herpesvirus-infected ocular diseases. Recent advances in knowledge of innate immune responses to viral infections have elucidated a complex network of the interplay between the invading virus, the target cells, and the host immune responses. Ocular cytomegalovirus latency exacerbates the development of choroidal neovascularization. Viruses have various strategies to evade or delay the cytokine response, and buy time to replicate in the host. Some signaling proteins impact the virologic, immunologic, and pathological processes of herpesvirus infection with particular emphasis on retinitis caused by HCMV. The accumulated data suggest that signaling proteins can differentially affect the severity of viral diseases in a highly cell-type-specific manner, reflecting the diversity and complexity of herpesvirus infection and the ocular compartment. By summarizing the immunological characteristics and pathogenesis of HCMV ocular infection, it will provide important information on the development of antiviral therapy, immunotherapy, and antidrug resistance.

1. Introduction

Human cytomegalovirus (HCMV) is a member of the beta-herpesvirus family, which tends to establish asymptomatic and lifelong latent infection [1]. Opportunistic HCMV reactivation is a common cause of increasing morbidity and mortality in newborns, the aged population, solid organ transplant patients, hematologic malignancy, or immunodeficient patients [2, 3].

HCMV was first reported to induce HCMV retinitis in 1957 [4], which is known to predominantly target retinal vascular endothelial cells, glial cells, and retinal pigment epithelial cells in the eye [5]. HCMV keratitis or retinitis is the most common opportunistic complication of infection in immunocompromised patients [2, 6, 7], including HIV-1-infected individuals. In general, HIV-1-infected individuals who have viral retinitis tend to be severe, long-lasting, and resistant to conventional treatment with a high rate of complications and significant visual morbidity [7]. Despite the widespread use of highly active antiretroviral therapy (HAART), up to 50–85% of AIDS patients develop ocular manifestations [8, 9, 10]. It has been well known that the HCMV can infect the immune-privileged retina site, lead to severe visual loss, and affect the quality of life in HIV-1-positive individuals [5]. Opportunistic infections develop when there is a deterioration of the immune status of the individual, which can be measured with the help of CD4⁺ T-cell counts [5]. The proportion of HCMV retinitis manifestations was also correlated with the CD4⁺ T-cell counts in patients [8]. Retinitis symptom has been classified into two categories, namely, infectious and noninfectious with the vast majority of manifestations occurring in the former. The infectious group mainly consists of the herpetic group of viral infections. Bacterial causes may be due to Staphylococcus epidermidis, Staphylococcus aureus, Pseudomonas aeruginosa, alpha-hemolytic Streptococcus, Micrococcus, and Bacillus. Fungal keratitis in HIV-1-infected individuals depend on the geographic locations from which the patient comes. Microsporidia and Acanthamoeba are common protozoal pathogens. Noninfective inflammatory causes include peripheral ulcerative keratitis, keratoconjunctivitis sicca, and squamous cell carcinoma of the conjunctiva. Posterior segment lesions caused by HCMV show severe visual disorders [7]. A severity that is abnormally severe or minimally reactive makes clinicians suspect immunosuppression. In the HAART era, the incidence, visual morbidity, and mortality of HCMV-related retinitis, and other HIV-1-related retinopathies showed a decline [8, 10]. In this chapter, we will focus on the immunological mechanisms and pathological processes of HCMV infection and strive to highlight those clinical manifestations that should alert the clinicians to suspect underlining HIV-1 infection and provide a basis for intervention.

2. Immunology of HCMV ocular diseases

2.1 Immunology

HCMV has evolved a variety of mechanisms to evade host immune surveillance and to establish latent infection with the ability to reactivate when the immune surveillance is compromised [11]. The host immune responses to HCMV involve both innate and adaptive immune systems, which play an important role in resolving both primaries, reactivating, and superinfections [12]. Under the innate and adaptive immune responses, a low viral load and latent state are established in the host after HCMV infection, but under the stimulation of the irregular and intermittent viral antigen reactivation, the functions of T-cells are exhausted [13]. At the same time, the HCMV virus can also encode a large number of gene products that interfere with the immune clearance responses to evade immune surveillance [11, 14, 15]. Considering that immunological clearance and evasion are associated with clinical outcomes, we summarized the immunological mechanisms of HCMV infection.

2.1.1 Innate immune responses

The natural killer (NK) cell is an important member of the innate lymphoid cell family for defense against HCMV during the early stages of infection and before the development of adaptive immune response due to its strong ability to kill infected or transformed cells [15]. The activities of NK-cells or NKT-cells (a subset of T-cells that co-express T- and NK-cell receptors) depend on the balance between activating and inhibitory signals transduced by its receptors [15]. They are also protected by releasing anti-viral cytokine interferon (IFN)-γ or by direct lysis, or autophagy of infected cells [15]. This will determine the disease progression of HCMV infection with ocular target cells.

2.1.2 Adaptive immune responses

Accumulating studies have shown that NK-cells take part in adaptive immune responses, such as clonal expansion and immune memory, during cytomegalovirus infection [16]. Clonal expansion not only serves to amplify the number of specific lymphocytes and mount robust protective responses against the pathogen but also results in the selection and differentiation of the responding lymphocytes [16]. In both innate and adaptive lymphocytes, clonal expansion is a critical process for host defenses. It has been shown that antigen (Ag)-specific T-cell expansion was estimated up to 400,000-fold [17]. The intensity of the adaptive immune responses suppresses the acute inflammatory responses caused by HCMV, causing less ocular tissue damage and sequelae.

In primary HCMV infections, CD4⁺ T-cells play a vital role in controlling symptomatic disease in healthy and immunocompromised patients. It is important to note that HCMV-infected cells can induce impairment of HCMV-specific effector CD4⁺ T-cell responses [18]. A subpopulation of HCMV-specific CD4⁺ T-cells has been shown to express Foxp3 and to perform functions similar to regulatory T-cells, such as the production of IL-10 [18, 19]. Latent infection is associated with secretory expression of CCL8, IL-10, and TGF-β [13, 18]. The presence of viral genes and viral IL-10 lead to down-regulate human leukocyte antigen (HLA) class II molecules and limit antigen presentation to CD4⁺ T-cells in antiviral immunity [12, 18]. At the peak of HCMV infection, HCMV-specific CD4⁺ T-cells are CD45RA⁺ CD45RO⁺ and express CD27⁺, CD28⁺, CD38⁺, and CD40L⁺. During the latent infection period, the HCMV-specific CD4⁺ T-cells are rich in CD27⁻ CD28⁻ CD4⁺ T-cells (5–10%) [13]. It has been known that HCMV-specific CD4⁺ T-cells are required for the maintenance of HCMV-specific CD8⁺ T- and B-cell responses in adoptive T-cell immunotherapy in transplant patients [13]. CD8⁺ T-cells undergo extensive expansion before differentiating into cytotoxic T-cells capable of producing high levels of cytokines, including IL-2, IFN-γ, TNF-α, perforin, and granzyme B [13]. For therapeutics, CD8⁺ T-cells with long-term survival rates and the potential to respond to challenges are very useful in adoptive transfer strategies for treating HCMV infection. Therefore, HCMV-specific CD8⁺ T-cell responses, including the maintenance, distribution, effector function, and metabolic requirements of these cells, have been highly interesting from a vaccine perspective.

Activated HCMV is typically controlled by CD4⁺ and CD8⁺ T-cell responses, while the virus replicates under the immunosuppressive condition and spreads rapidly to nearby tissues, resulting in worsening of retinitis, such as the patients accompanied with HIV-1 infection and chemotherapy for cancers. In addition to eliminating or perturbing surface immune recognition molecules (HLA I or HLA II molecules) from the antigen-presenting cells (B lymphocytes, dendritic cells, monocytes, or macrophages), HCMV immune evasion mechanisms have evolved to escape recognition and immune clearance of infected cells by effector cells through innate immunity, mimicked the inhibitory ligands or downregulate the activating ligands of NK-cells [11].

2.2 Pathogenesis of HCMV ocular diseases

Although HCMV infection can occur in healthy individuals, it is uncommon to observe symptomatic infection in individuals without immune suppression [3, 20]. HCMV retinitis is a clinical syndrome characterized by full-thickness necrotizing retinitis, which can result in profound vision loss, retinal detachment, and permanent vision loss [21]. The possible transmission paths of HCMV include blood, prenatal intrauterine infection, perinatal infection through breast milk or genital secretions, saliva, and sperm [22, 23].

In immunocompromised patients, primary HCMV infection causes severe complications, including pyrexia, viremic-septicemia, pneumonitis, and immunosuppression [24, 25]. In total, 60% of patients have been infected with HCMV prior to the onset of critical illness, and are commonly infected before adulthood [26]. It has been shown that patients who suffered HCMV reactivation during critical illness have ~2-flod the mortality rate of those who have not reactivated [27]. The clinical data suggest that the clinical outcome is not only the cause of HCMV replication but also the degree of virus-associated immune responses [3, 25]. In addition to HCMV and HIV-1 co-infected patients with lower CD4⁺ T-cell counts have a higher risk of mortality, HCMV has sophisticated strategies to circumvent immunocyte recognition, such as changing the signals of immunomodulatory molecules and subverting T-cell and NK-cell function, and allowing it to establish lifelong infection in blood and bone marrow [16]. In these disorders, HCMV-infected individuals could be accompanied by hemophagocytic lymphohistiocytosis-associated genetic defects. The active and latent infection induce sustained systemic inflammatory responses and predisposes patients to produce autoantibodies, which increased the autoimmune disease progression [28]. It has been demonstrated that individuals infected with one HCMV strain may not necessarily be able to resist other HCMV strains [29]. HCMV also causes immunosuppression associated with T-cell exhaustion, which contributes to the persistence of infection [13].

HCMV retinitis is caused by lytic infection, the conclusion supported by clinical resolution with antiviral therapy. Healing is through fibrosis, which predisposes patients to future retinal detachment and is also the cause of severe vision loss [30]. When antiretroviral therapy is introduced, some individuals with HIV-1 develop immune-restorative uveitis, which is an inflammatory response to the presence of HCMV antigens in the eye by activating viral immediate-early gent product-2 and increasing FasL secretion [31]. This condition may cause more visual impairment in patients than potential retinitis [30]. One possible explanation is that the damage of HIV-1 infection to the blood-retinal barrier may contribute to the preferential entry of HCMV into the oculus [30].

2.3 Interventions of HCMV ocular diseases

Acute retinal necrosis was first described in Japan as acute unilateral panuveitis, retinal periarteritis, and necrotizing retinitis progressing to retinal detachment [32]. The symptoms of acute retinal necrosis include redness, ocular pain, photophobia, floaters, and blurred vision [32]. These studies suggest that the challenges in diagnosis and therapeutic challenges primarily in the absence of guidelines or evidence-based literature to follow [21].

Ganciclovir was licensed in 1989 and remains the only licensed drug sufficient to treat active HCMV infection. Although the oral prodrug valganciclovir was licensed in 2001, it delivers the same active ingredient. Ganciclovir-resistant HCMV disease has become a serious clinical problem in transplanted populations. Mutations in viral kinase (UL97) or polymerase (UL54) have been shown to mediate resistance to ganciclovir and valganciclovir [33]. For strains of HCMV-resistant to ganciclovir, foscarnet is used off-label. Thus, this field would benefit from more licensed drugs that are both safe and effective anti-HCMV [30]. This becomes particularly important for clinical trials seeking to test the anti-HCMV activity of novel compounds.

In large randomized controlled trials of HIV-1-associated HCMV retinitis in the era before combination anti-HIV treatment, in which the primary endpoint was an objective progression of CMV retinitis, an intra-ocular ganciclovir implant (15% of patients progressed after 100 days of treatment) was superior to intravenous ganciclovir (65% of patients progressed). The limitation of the intra-ocular ganciclovir implant was its failure to prevent CMV disease in the contralateral eye. In a subsequent randomized controlled trial of HIV-associated CMV retinitis, treatment with oral valganciclovir (38% of patients progressed after 100 days of treatment) was similarly effective to initial intravenous ganciclovir for 4 weeks followed by oral valganciclovir (45% of patients progressed). During the latter trial, most patients were also taking a combination anti-HIV treatment. As the ocular penetration of systemically administered anti-CMV drugs is limited, current clinical guidelines include consideration of intraocular injection of anti-HCMV drugs for patients who have sight-threatening HCMV retinitis (Tables 1 and 2). In addition to ganciclovir, given that the retina shows acute necrosis of one eye, corticosteroid or methylprednisolone is very important because of its effects in relieving intense inflammatory responses [21].

Agent	Route	Dosage	Side effects	Monitoring
Ganciclovir	IV	5 mg/kg 12 h for 2 wk	Myelosuppression, renal impairment, hepatic impairment, gastrointestinal symptoms, CNS disturbances; less well-tolerated	FBC, UEC, LFT
Valganciclovir	PO	900 mg BD for 3 wk		FBC, UEC, LFT
Foscarnet	IV	60 mg/kg 8 h for 2–3 wk; 90 mg/kg 12 h for 2–3 wk	Nephrotoxicity, hypocalcemia, hypomagnesemia (can lead to seizures), anemia, genital ulceration	FBC, UEC, calcium, and magnesium level
Cidofovir	IV	5 mg/kg once weekly for 2 wk; probenecid pre- and post-infusion	Renal impairment, neutropenia, ocular hypotony, iritis	FBC, UEC
Ganciclovir	Intravitreal	2–4 mg/0.05–0.1 mL	—	—
Foscarnet	Intravitreal	2.4 mg/0.1 mL	—	—
Cidofovir	Intravitreal	0.02 mg/0.1 mL	Narrow therapeutic window, retinotoxic	—

Table 1.

Antiviral treatment for HCMV retinitis [32].

Abbreviations: IV, intravenous; PO, oral; BD, twice daily; FBC, full blood count; UEC, urea electrolytes creatinine; LFT, liver function test

Agent	Bilaterality	Outcomes
Rituximab	Unilateral or bilaterality	Resolved
Basiliximab	—	Not reported
Anti-thymocyte globulin	Unilateral or bilaterality	Resolved
Alemtuzumab	Unilateral or bilaterality	Resolved and marked improvement
Natalizumab	Unilateral or bilaterality	Retinal detachment or blind
Ruxolitinib	Unilateral	Not reported
Tofacitinib	Bilaterality	Not reported

Table 2.

Biologic immunosuppression and HCMV retinitis [32].

The HCMV persistent or progressive retinitis may be resolved by systemic administration of virus-specific cytotoxic T-cells (CTLs) [34]. HCMV-specific CTL therapy may become a novel monotherapy or adjunctive therapy, or both, for retinitis, especially in eyes that are resistant, refractory, or intolerant of antiviral therapies [34]. In addition, it has been demonstrated that HCMV strain-specific antibodies play an important role in preventing viral recrudescence after transplantation [29]. Antibodies, natural killer cells, and macrophages theoretically contribute to protective immune responses and are expected to interact and cooperate with T-cells to control HCMV replication. It was also recommended that studies of active immunization should proceed concurrently with passive immunotherapy using monoclonal antibodies with defined reactivity against specific proteins of HCMV against the resistance [34]. Recently, letermovir has been used in ganciclovir-resistant patients at doses of 720–960 mg, while intravitreal therapy with formic acid or ganciclovir was also used, and by monitoring continuous hematologic, renal, and hepatic function, some patients experienced an improvement in symptoms [32].

3. Conclusions

The interaction between HCMV and the host immune system is complex. NK-cells play an important role in the virus infecting the ocular target cells and take part in the processes of innate immune response and adaptive immune response. After the acute infection in adolescent accompanied by acute symptoms, the virus easily establishes latent infection and reactivate in immunodeficient HIV-1-infected and transplanted individuals. T-cell function is important in controlling HCMV recurrence. Immunodeficient individuals are susceptible to developing HCMV retinitis, which can be treated with systemic and intraocular topical medications, but are also prone to developing drug resistance. Therefore, understanding the immunology and pathogenic mechanisms of HCMV will help us further develop effective antiviral drugs for the treatment or mitigation of HCMV ocular disease.

Acknowledgments

This work was supported by the Wuxi Key Medical Talents Program (ZDRC024) and the Top Talent Support Program for Young and Middle-Aged People of the Wuxi Health Committee (BJ2020094).

Conflict of interest

The authors declare no conflict of interest.