IFN-γ-mediated control of SARS-CoV-2 infection through nitric oxide.

Introduction: The COVID-19 pandemic has highlighted the need to identify mechanisms of antiviral host defense against SARS-CoV-2. One such mediator is interferon-g (IFN-γ), which, when administered to infected patients, is reported to result in viral clearance and resolution of pulmonary symptoms. IFN-γ treatment of a human lung epithelial cell line triggered an antiviral activity against SARS-CoV-2, yet the mechanism for this antiviral response was not identified. Methods: Given that IFN-γ has been shown to trigger antiviral activity via the generation of nitric oxide (NO), we investigated whether IFN-γ induction of antiviral activity against SARS-CoV-2 infection is dependent upon the generation of NO in human pulmonary epithelial cells. We treated the simian epithelial cell line Vero E6 and human pulmonary epithelial cell lines, including A549-ACE2, and Calu-3, with IFN-γ and observed the resulting induction of NO and its effects on SARS-CoV-2 replication. Pharmacological inhibition of inducible nitric oxide synthase (iNOS) was employed to assess the dependency on NO production. Additionally, the study examined the effect of interleukin-1b (IL-1ß) on the IFN-g-induced NO production and its antiviral efficacy. Results: Treatment of Vero E6 cells with IFN-γ resulted in a dose-responsive induction of NO and an inhibitory effect on SARS-CoV-2 replication. This antiviral activity was blocked by pharmacologic inhibition of iNOS. IFN-γ also triggered a NO-mediated antiviral activity in SARS-CoV-2 infected human lung epithelial cell lines A549-ACE2 and Calu-3. IL-1ß enhanced IFN-γ induction of NO, but it had little effect on antiviral activity. Discussion: Given that IFN-g has been shown to be produced by CD8+ T cells in the early response to SARS-CoV-2, our findings in human lung epithelial cell lines, of an IFN-γ-triggered, NO-dependent, links the adaptive immune response to an innate antiviral pathway in host defense against SARS-CoV-2. These results underscore the importance of IFN-γ and NO in the antiviral response and provide insights into potential therapeutic strategies for COVID-19.

Myocarditis, disseminated infection, and early viral persistence following experimental coxsackievirus B infection of cynomolgus monkeys.

Coxsackievirus B (CVB) infection is a common cause of acute viral myocarditis. The clinical presentation of myocarditis caused by this enterovirus is highly variable, ranging from mildly symptoms to complete hemodynamic collapse. These variations in initial symptoms and in the immediate and long term outcomes of this disease have impeded development of effective treatment strategies. Nine cynomolgus monkeys were inoculated with myocarditic strains of CVB. Virological studies performed up to 28 days post-inoculation demonstrated the development of neutralizing antibody in all animals, and the presence of CVB in plasma. High dose intravenous inoculation (n = 2) resulted in severe disseminated disease, while low dose intravenous (n = 6) or oral infection (1 animal) resulted in clinically unapparent infection. Transient, minor, echocardiographic abnormalities were noted in several animals, but no animals displayed signs of significant acute cardiac failure. Although viremia rapidly resolved, signs of myocardial inflammation and injury were observed in all animals at the time of necropsy, and CVB was detected in postmortem myocardial specimens up to 28 days PI. This non-human primate system replicates many features of illness in acute coxsackievirus myocarditis and demonstrates that myocardial involvement may be common in enteroviral infection; it may provide a model system for testing of treatment strategies for enteroviral infections and acute coxsackievirus myocarditis.

Virological evidence supporting the use of raltegravir in HIV post-exposure prophylaxis regimens.

BACKGROUND: The goal of post-exposure prophylaxis (PEP) for HIV is to prevent the establishment of a persistent infection following exposure to the virus. Integrase inhibitors have several potential advantages in PEP regimens, including the capacity to inhibit integration of HIV genomes that have already proceeded through reverse transcription, thereby becoming refractory to reverse transcriptase inhibitors. We sought to determine if integrase inhibitors extend the window of time during which PEP intervention might be successful. METHODS: Primary costimulated CD4(+) T-cells or macrophages were infected with a luciferase-bearing HIV reporter virus, permitting sensitive detection of viral gene expression under different drug treatment conditions. Relevant antiretroviral agents were added at various pre- or post-infection time points. RESULTS: We showed that raltegravir effectively blocks HIV infection, even when cells are challenged with a large amount of virus. We also demonstrated that during infection of both primary costimulated CD4(+) T-cells and primary macrophages, raltegravir can inhibit infection when added at later post-infection time points than the reverse transcriptase inhibitor efavirenz. CONCLUSIONS: This longer post-infection efficacy window, coupled with favourable pharmacokinetic properties and low toxicity, suggest that raltegravir may prove useful in HIV PEP.

Thymic volume, T-cell populations, and parameters of thymopoiesis in adolescent and adult survivors of HIV infection acquired in infancy.

OBJECTIVES: Antiretroviral therapy has significantly prolonged the lifespan of children who acquire HIV infection in infancy, but the impact of HIV on thymus-mediated maintenance of T lymphocytes has not been studied. To examine the long-term effects of HIV infection in childhood on thymopoiesis, thymic volume and parameters of thymic function from clinically stable adolescents and young adults with HIV infection acquired in infancy were compared with those from uninfected controls. METHODS: Thymic volume was determined using three-dimensional reconstruction and volumetric analysis of non-contrast enhanced computed tomography images of the upper chest. The degree of fat involution was assessed using a semiquantitive scoring system. CD4 and CD8 T cell populations and T cell receptor recombination excision circles (TREC) concentrations in peripheral blood lymphocytes were measured in all subjects. RESULTS: Twenty youths (aged 17.6 +/- 2.5 years) with HIV infection acquired perinatally (n = 18) or by neonatal transfusion (n = 2) were enrolled whose HIV plasma viral load had been undetectable for a median of 3.1 years, along with 18 seronegative healthy young adults (aged 20.6 +/- 1.3 years). HIV infected subjects and controls had indistinguishable CD4 T cell counts, thymus volumes (20.5 versus 15.8 cm), thymic index scores, and TREC values. Thymic volume correlated with the number and percentage of CD4 T lymphocytes in the control group and with the number of TREC in CD4 lymphocytes in the HIV infected group. CONCLUSIONS: Long term survivors of pediatric HIV infection appear to have retained or recovered thymic volume and thymic activity approximating uninfected youths.

Tetanus immunity after diphtheria, tetanus toxoids, and acellular pertussis vaccination in children with clinically stable HIV infection.

BACKGROUND: HIV infection often impairs the immune response to childhood vaccines. OBJECTIVE: We sought to study the ability of HIV-infected children receiving highly active antiretroviral therapy (HAART) to generate a booster response to immunization with a recall antigen to which they had lost humoral immunity. METHODS: Diphtheria, tetanus toxoids, and acellular pertussis (DTaP) vaccination was given at either 16 or 36 weeks after initiation of HAART to 37 HIV-infected children 2 to 9 years of age with a history of DTaP or diphtheria-tetanus-pertussis receipt who had negative tetanus antibody titers (

Immunological response to highly active antiretroviral therapy in children with clinically stable HIV-1 infection.

We studied changes in 60 immunological parameters after the administration of highly active antiretroviral therapy (HAART) in 192 clinically stable antiretroviral drug-experienced HIV-1-infected children 4 months-17 years old. The studied immunological parameters included standard lymphocyte subsets and lymphocyte surface markers of maturation and activation. The most significant changes during the 48-week study period were seen for CD8(+), CD8(+)CD62L(+)CD45RA(+), CD8(+)CD38(+)HLA-DR(+), and CD4(+) T cell percentages (P < .0001 for all parameters). These changes suggest that significant decreases in the expression of activation markers and increases in the expression of naive markers in the CD8(+) T cell population may be related to better virologic control in these HIV-1-infected children, who had relatively stable immune function at the initiation of HAART. At week 44 of HAART, the major immunological parameters in these HIV-1-infected children moved from baseline values to about halfway to two-thirds of the way toward the values in healthy, uninfected children.

Growth of human immunodeficiency virus-infected children receiving highly active antiretroviral therapy.

BACKGROUND: Weight and height growth of HIV-infected children tends to lag behind that of uninfected children of similar age. Previous reports of the effect of highly active antiretroviral therapy (HAART) on the growth of HIV-infected children have been contradictory. METHODS: Age- and gender-adjusted height and weight z scores were studied for 192 HIV-infected children, 4 months to 17 years of age, who had been treated with antiretroviral therapy for at least 16 weeks. These children, in clinically and immunologically stable condition, were enrolled into one of 4 HAART regimens and evaluated for 96 weeks. RESULTS: At baseline, these HIV-infected children were significantly shorter than uninfected children (mean z score, -0.57; 95% confidence interval, -0.73 to -0.41; P < 0.001). Children with greater viral loads at baseline were significantly shorter and lighter than children with smaller viral loads (both P < 0.001). Administration of HAART led to an increase in mean weight z scores to normal values (mean z score increase, from -0.16 to >0) by week 48 and an increase in mean height z scores of 72% toward normal values (mean z score increase, from -0.57 to -0.16) by week 96. Younger children gained height more rapidly (P < 0.001), and children with greater baseline viral loads gained weight more rapidly (P < 0.001). There was no evidence of differential height or weight changes in 48 weeks between children with different degrees of virologic control. CONCLUSIONS: HAART improved the average weight gain of HIV-infected children from subnormal to normal after 1 year and improved average height growth to nearly normal after 2 years.

Reported adherence as a determinant of response to highly active antiretroviral therapy in children who have human immunodeficiency virus infection.

OBJECTIVE: The complexity of highly active antiretroviral therapy (HAART), with multiple medications, formulations, and dosing intervals, makes adherence challenging. Little is known about the adherence of children to HAART. The objective of this study was to identify correlates of adherence to HAART and the relationship between adherence and study outcomes in a pediatric clinical trial. METHODS: Pediatric AIDS Clinical Trials Group 377 is a phase I/II randomized trial of 4 HAART regimens in antiretroviral-experienced, clinically stable children aged 4 months to 17 years. The 4 treatment arms include various 3- or 4-drug combinations of d4T, 3TC, nevirapine, ritonavir, and nelfinavir. After informed consent was obtained, 193 children were enrolled between December 1997 and September 1998. Questionnaires were developed to collect subject- or caregiver-reported adherence to study medications and to identify problems associated with medication administration. Every 3 months, the number of doses of each medication missed during the 3 days preceding the study visit was recorded. Full adherence (FA) and non-full adherence were defined as missing no doses and missing at least 1 dose, respectively. RESULTS: Adherence data from study week 48 or the most recent study visit were available for 125 children (week 48 for 109 children). Overall, 70% of children reported FA and 30% reported non-full adherence. Adherence did not differ by treatment arm, age, or the child's knowledge of his or her human immunodeficiency virus infection status. There was a suggestion that adherence was less for white than nonwhite children (40% vs 73% FA) and did not differ between black and Hispanic children. Rates of FA were 82% for d4T, 79% for 3TC, 83% for nevirapine, 84% for ritonavir, and 68% for nelfinavir. Despite the similar rates of FA, difficulties with taking specific medications were reported most frequently for ritonavir and nelfinavir. These included poor taste, patient refusal, and scheduling problems. Adherence was associated with the virologic response: FA was seen in 92% of children with > or =2 log10 drop in viral load and in 64% with <2 log10 drop in viral load. CONCLUSION: In children, reported adherence predicts the virologic response to HAART therapy and is a useful measure of adherence. Interventions and regimens to increase adherence to HAART should result in an improved outcome.