RESUMEN
The role of CD8+ T cells in SARS-CoV-2 pathogenesis or mRNA-LNP vaccine-induced protection from lethal COVID-19 is unclear. Using mouse-adapted SARS-CoV-2 virus (MA30) in C57BL/6 mice, we show that CD8+ T cells are unnecessary for the intrinsic resistance of female or the susceptibility of male mice to lethal SARS-CoV-2 infection. Also, mice immunized with a di-proline prefusion-stabilized full-length SARS-CoV-2 Spike (S-2P) mRNA-LNP vaccine, which induces Spike-specific antibodies and CD8+ T cells specific for the Spike-derived VNFNFNGL peptide, are protected from SARS-CoV-2 infection-induced lethality and weight loss, while mice vaccinated with mRNA-LNPs encoding only VNFNFNGL are protected from lethality but not weight loss. CD8+ T cell depletion ablates protection in VNFNFNGL but not in S-2P mRNA-LNP-vaccinated mice. Therefore, mRNA-LNP vaccine-induced CD8+ T cells are dispensable when protective antibodies are present but essential for survival in their absence. Hence, vaccine-induced CD8+ T cells may be critical to protect against SARS-CoV-2 variants that mutate epitopes targeted by protective antibodies.
Asunto(s)
Anticuerpos Antivirales , Linfocitos T CD8-positivos , Vacunas contra la COVID-19 , COVID-19 , SARS-CoV-2 , Glicoproteína de la Espiga del Coronavirus , Animales , Linfocitos T CD8-positivos/inmunología , Ratones , SARS-CoV-2/inmunología , COVID-19/inmunología , COVID-19/prevención & control , Femenino , Glicoproteína de la Espiga del Coronavirus/inmunología , Glicoproteína de la Espiga del Coronavirus/genética , Vacunas contra la COVID-19/inmunología , Masculino , Anticuerpos Antivirales/inmunología , Ratones Endogámicos C57BL , Humanos , Modelos Animales de EnfermedadRESUMEN
Receptor-interacting protein kinase 3 (RIPK3) and mixed lineage kinase domain-like pseudokinase (MLKL) are proteins that are critical for necroptosis, a mechanism of programmed cell death that is both activated when apoptosis is inhibited and thought to be antiviral. Here, we investigated the role of RIPK3 and MLKL in controlling the Orthopoxvirus ectromelia virus (ECTV), a natural pathogen of the mouse. We found that C57BL/6 (B6) mice deficient in RIPK3 (Ripk3-/-) or MLKL (Mlkl-/-) were as susceptible as wild-type (WT) B6 mice to ECTV lethality after low-dose intraperitoneal infection and were as resistant as WT B6 mice after ECTV infection through the natural footpad route. Additionally, after footpad infection, Mlkl-/- mice, but not Ripk3-/- mice, endured lower viral titers than WT mice in the draining lymph node (dLN) at three days postinfection and in the spleen or in the liver at seven days postinfection. Despite the improved viral control, Mlkl-/- mice did not differ from WT mice in the expression of interferons or interferon-stimulated genes or in the recruitment of natural killer (NK) cells and inflammatory monocytes (iMOs) to the dLN. Additionally, the CD8 T-cell responses in Mlkl-/- and WT mice were similar, even though in the dLNs of Mlkl-/- mice, professional antigen-presenting cells were more heavily infected. Finally, the histopathology in the livers of Mlkl-/- and WT mice at 7 dpi did not differ. Thus, the mechanism of the increased virus control by Mlkl-/- mice remains to be defined. IMPORTANCE The molecules RIPK3 and MLKL are required for necroptotic cell death, which is widely thought of as an antiviral mechanism. Here we show that C57BL/6 (B6) mice deficient in RIPK3 or MLKL are as susceptible as WT B6 mice to ECTV lethality after a low-dose intraperitoneal infection and are as resistant as WT B6 mice after ECTV infection through the natural footpad route. Mice deficient in MLKL are more efficient than WT mice at controlling virus loads in various organs. This improved viral control is not due to enhanced interferon, natural killer cell, or CD8 T-cell responses. Overall, the data indicate that deficiencies in the molecules that are critical to necroptosis do not necessarily result in worse outcomes following viral infection and may improve virus control.
Asunto(s)
Ectromelia Infecciosa , Animales , Ratones , Virus de la Ectromelia , Ectromelia Infecciosa/inmunología , Interferones/metabolismo , Ratones Endogámicos C57BL , Necroptosis/inmunología , Proteínas Quinasas/genética , Proteínas Quinasas/inmunología , Proteína Serina-Treonina Quinasas de Interacción con Receptores/genética , Proteína Serina-Treonina Quinasas de Interacción con Receptores/inmunologíaRESUMEN
Type I interferons (IFN-I) are antiviral cytokines that signal through the ubiquitous IFN-I receptor (IFNAR). Following footpad infection with ectromelia virus (ECTV), a mouse-specific pathogen, C57BL/6 (B6) mice survive without disease, while B6 mice broadly deficient in IFNAR succumb rapidly. We now show that for survival to ECTV, only hematopoietic cells require IFNAR expression. Survival to ECTV specifically requires IFNAR in both natural killer (NK) cells and monocytes. However, intrinsic IFNAR signaling is not essential for adaptive immune cell responses or to directly protect non-hematopoietic cells such as hepatocytes, which are principal ECTV targets. Mechanistically, IFNAR-deficient NK cells have reduced cytolytic function, while lack of IFNAR in monocytes dampens IFN-I production and hastens virus dissemination. Thus, during a pathogenic viral infection, IFN-I coordinates innate immunity by stimulating monocytes in a positive feedback loop and by inducing NK cell cytolytic function.
Asunto(s)
Virus de la Ectromelia/inmunología , Ectromelia Infecciosa/inmunología , Receptor de Interferón alfa y beta/metabolismo , Transducción de Señal , Animales , Citocinas/inmunología , Resistencia a la Enfermedad , Ectromelia Infecciosa/virología , Femenino , Hepatocitos/inmunología , Hepatocitos/virología , Inmunidad Innata , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/virología , Masculino , Ratones , Ratones Endogámicos C57BL , Monocitos/inmunología , Monocitos/virología , Receptor de Interferón alfa y beta/genéticaRESUMEN
CMV has been proposed to play a role in cancer progression and invasiveness. However, CMV has been increasingly studied as a cancer vaccine vector, and multiple groups, including ours, have reported that the virus can drive antitumor immunity in certain models. Our previous work revealed that intratumoral injections of wild-type murine CMV (MCMV) into B16-F0 melanomas caused tumor growth delay in part by using a viral chemokine to recruit macrophages that were subsequently infected. We now show that MCMV acts as a STING agonist in the tumor. MCMV infection of tumors in STING-deficient mice resulted in normal recruitment of macrophages to the tumor, but poor recruitment of CD8+ T cells, reduced production of inflammatory cytokines and chemokines, and no delay in tumor growth. In vitro, expression of type I IFN was dependent on both STING and the type I IFNR. Moreover, type I IFN alone was sufficient to induce cytokine and chemokine production by macrophages and B16 tumor cells, suggesting that the major role for STING activation was to produce type I IFN. Critically, viral infection of wild-type macrophages alone was sufficient to restore tumor growth delay in STING-deficient animals. Overall, these data show that MCMV infection and sensing in tumor-associated macrophages through STING signaling is sufficient to promote antitumor immune responses in the B16-F0 melanoma model.
Asunto(s)
Infecciones por Herpesviridae/inmunología , Melanoma/inmunología , Proteínas de la Membrana/metabolismo , Muromegalovirus/fisiología , Neoplasias Cutáneas/inmunología , Macrófagos Asociados a Tumores/inmunología , Animales , Movimiento Celular , Modelos Animales de Enfermedad , Humanos , Inmunidad/genética , Interferón Tipo I/metabolismo , Melanoma/virología , Melanoma Experimental , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neoplasias Cutáneas/virología , Carga Tumoral , Microambiente TumoralRESUMEN
NK cells play an important role in antiviral resistance. The integrin α2, which dimerizes with integrin ß1, distinguishes NK cells from innate lymphoid cells 1 and other leukocytes. Despite its use as an NK cell marker, little is known about the role of α2ß1 in NK cell biology. In this study, we show that in mice α2ß1 deficiency does not alter the balance of NK cell/ innate lymphoid cell 1 generation and slightly decreases the number of NK cells in the bone marrow and spleen without affecting NK cell maturation. NK cells deficient in α2ß1 had no impairment at entering or distributing within the draining lymph node of ectromelia virus (ECTV)-infected mice or at becoming effectors but proliferated poorly in response to ECTV and did not increase in numbers following infection with mouse CMV (MCMV). Still, α2ß1-deficient NK cells efficiently protected from lethal mousepox and controlled MCMV titers in the spleen. Thus, α2ß1 is required for optimal NK cell proliferation but is dispensable for protection against ECTV and MCMV, two well-established models of viral infection in which NK cells are known to be important.
Asunto(s)
Ectromelia Infecciosa/inmunología , Infecciones por Herpesviridae/inmunología , Integrina alfa2beta1/metabolismo , Células Asesinas Naturales/inmunología , Animales , Recuento de Células , Proliferación Celular , Modelos Animales de Enfermedad , Virus de la Ectromelia/inmunología , Ectromelia Infecciosa/sangre , Ectromelia Infecciosa/virología , Femenino , Infecciones por Herpesviridae/sangre , Infecciones por Herpesviridae/virología , Humanos , Inmunidad Innata , Integrina alfa2beta1/inmunología , Células Asesinas Naturales/metabolismo , Masculino , Ratones , Muromegalovirus/inmunología , Replicación Viral/inmunologíaRESUMEN
Cells sensing infection produce Type I interferons (IFN-I) to stimulate Interferon Stimulated Genes (ISGs) that confer resistance to viruses. During lympho-hematogenous spread of the mouse pathogen ectromelia virus (ECTV), the adaptor STING and the transcription factor IRF7 are required for IFN-I and ISG induction and resistance to ECTV. However, it is unknown which cells sense ECTV and which pathogen recognition receptor (PRR) upstream of STING is required for IFN-I and ISG induction. We found that cyclic-GMP-AMP (cGAMP) synthase (cGAS), a DNA-sensing PRR, is required in bone marrow-derived (BMD) but not in other cells for IFN-I and ISG induction and for resistance to lethal mousepox. Also, local administration of cGAMP, the product of cGAS that activates STING, rescues cGAS but not IRF7 or IFN-I receptor deficient mice from mousepox. Thus, sensing of infection by BMD cells via cGAS and IRF7 is critical for resistance to a lethal viral disease in a natural host.
Asunto(s)
Médula Ósea/virología , Virus de la Ectromelia/patogenicidad , Ectromelia Infecciosa/virología , Nucleótidos Cíclicos/metabolismo , Animales , Médula Ósea/metabolismo , Interacciones Huésped-Patógeno/inmunología , Inmunidad Innata/inmunología , Interferón Tipo I/metabolismo , Ratones Transgénicos , Nucleotidiltransferasas/genética , Proteínas Serina-Treonina Quinasas/metabolismoRESUMEN
Because of the rapid mutations of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), an effective vaccine against SARS-CoV-2 variants is needed to prevent coronavirus disease 2019 (COVID-19). T cells, in addition to neutralizing antibodies, are an important component of naturally acquired protective immunity, and a number of studies have shown that T cells induced by natural infection or vaccination contribute significantly to protection against several viral infections including SARS-CoV-2. However, it has never been tested whether a T cell-inducing vaccine can provide significant protection against SARS-CoV-2 infection in the absence of preexisting antibodies. In this study, we designed and evaluated lipid nanoparticle (LNP) formulated mRNA vaccines that induce only T cell responses or both T cell and neutralizing antibody responses by using two mRNAs. One mRNA encodes SARS-CoV-2 Omicron Spike protein in prefusion conformation for induction of neutralizing antibodies. The other mRNA encodes over one hundred T cell epitopes (multi-T cell epitope or MTE) derived from non-Spike but conserved regions of the SARS-CoV-2. We show immunization with MTE mRNA alone protected mice from lethal challenge with the SARS-CoV-2 Delta variant or a mouse-adapted virus MA30. Immunization with both mRNAs induced the best protection with the lowest viral titer in the lung. These results demonstrate that induction of T cell responses, in the absence of preexisting antibodies, is sufficient to confer protection against severe disease, and that a vaccine containing mRNAs encoding both the Spike and MTE could be further developed as a universal SARS-CoV-2 vaccine.
Asunto(s)
Vacunas contra la COVID-19 , COVID-19 , Animales , Humanos , Ratones , COVID-19/prevención & control , SARS-CoV-2 , Anticuerpos Neutralizantes , Epítopos de Linfocito T , ARN Mensajero/genéticaRESUMEN
It is known that aging decreases natural resistance to viral diseases due to dysfunctional innate and adaptive immune responses, but the nature of these dysfunctions, particularly in regard to innate immunity, is not well understood. We have previously shown that C57BL/6J (B6) mice lose their natural resistance to footpad infection with ectromelia virus (ECTV) due to impaired maturation and recruitment of natural killer (NK) cells to the draining popliteal lymph node (dLN). More recently, we have also shown that in young B6 mice infected with ECTV, the recruitment of NK cells is dependent on a complex cascade whereby migratory dendritic cells (mDCs) traffic from the skin to the dLN, where they produce CCL2 and CCL7 to recruit inflammatory monocytes (iMOs). In the dLN, mDCs also upregulate NKG2D ligands to induce interferon gamma (IFN-γ) expression by group 1 innate lymphoid cells (G1-ILCs), mostly NK in cells but also some ILC1. In response to the IFN-γ, the incoming uninfected iMOs secret CXCL9 to recruit the critical NK cells. Here, we show that in aged B6 mice, the trafficking of mDCs to the dLN in response to ECTV is decreased, resulting in impaired IFN-γ expression by G1-ILCs, reduced accumulation of iMOs, and attenuated CXCL9 production by iMOs, which likely contributes to decrease in NK cell recruitment. Together, these data indicate that defects in the mDC response to viral infection during aging result in a reduced innate immune response in the dLN and contribute to increased susceptibility to viral disease in the aged.
Asunto(s)
Células Dendríticas/metabolismo , Virus de la Ectromelia/inmunología , Inmunidad Innata/inmunología , Ganglios Linfáticos/metabolismo , Envejecimiento , Animales , RatonesRESUMEN
Traditionally, baseball players have used a heavy bat for warm-up before competition. Because bat velocity is an essential component to hitting a baseball, and because players warm up differently, there is a need to investigate the best way to maximize post warm-up bat velocity. The purpose of this study was to determine the effects of warm-up with different weighted bats on normal baseball bat velocity. Nineteen recreational male baseball players (age, 24.5 +/- 3.9 years; height, 181.1 +/- 8.4 cm; body mass, 87.9 +/- 18.4 kg) participated in this study. Three different randomized warm-up conditions were completed and analyzed for velocity and for their effect on post warm-up normal baseball bat velocity. Subjects were instructed to perform 5 maximal swings with each of 3 different weighted bats-light (LB = 9.6 oz), normal (NB = 31.5 oz), and heavy (HB = 55.2 oz)-followed by 30-second rest and then 5 swings of the NB. Analysis of variance revealed that warm-up velocity of the LB (63.57 +/- 3.58 mph) was significantly (p < 0.05) faster than that of NB (51.25 +/- 3.01 mph) and HB (41.79 +/- 3.01 mph), whereas warm-up velocity of NB was also significantly faster than that of HB. For post warm-up, LB (52.29 +/- 2.68 mph) and NB (50.60 +/- 3.04 mph) produced significantly faster velocity of the normal bat than the HB (48.26 +/- 2.98 mph). Warming up with 5 swings of a light or normal bat appears to increase post warm-up velocity of the normal bat when compared with warming up with a heavy bat after a rest period of 30 seconds. Within the bat weight spectrum of this study, it is suggested that when preparing to hit, 5 warm-up swings with either a light or normal bat will allow a player to achieve the greatest velocity of their normal bat.
Asunto(s)
Aceleración , Béisbol , Equipo Deportivo , Percepción del Peso , Adulto , Humanos , MasculinoRESUMEN
During disseminating viral infections, a swift innate immune response (IIR) in the draining lymph node (dLN) that restricts systemic viral spread is critical for optimal resistance to disease. However, it is unclear how this IIR is orchestrated. We show that after footpad infection of mice with ectromelia virus, dendritic cells (DCs) highly expressing major histocompatibility complex class II (MHC class IIhi DCs), including CD207+ epidermal Langerhans cells (LCs), CD103+CD207+ double-positive dermal DCs (DP-DCs), and CD103-CD207- double-negative dermal DCs (DN-DCs) migrate to the dLN from the skin carrying virus. MHC class IIhi DCs, predominantly LCs and DP-DCs, are the first cells upregulating IIR cytokines in the dLN. Preventing MHC class IIhi DC migration or depletion of LCs, but not DP-DC deficiency, suppresses the IIR in the dLN and results in high viral lethality. Therefore, LCs are the architects of an early IIR in the dLN that is critical for optimal resistance to a disseminating viral infection.