CD4+ T Cells Are Not Required for the Induction of Dengue Virus-Specific CD8+ T Cell or Antibody Responses but Contribute to Protection after Vaccination

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CD4 T cells provide protection against cytomegalovirus (CMV) and other persistent viruses, and the ability to quantify and characterize epitope-specific responses is essential to gain a more precise understanding of their effector roles in this regard. Here, we report the first two I-A d -restricted CD4 T cell responses specific for mouse CMV (MCMV) epitopes and use a major histocompatibility complex class II (MHC-II) tetramer to characterize their phenotypes and functions. We demonstrate that MCMV-specific CD4 T cells can express high levels of granzyme B and kill target cells in an epitope-and organ-specific manner. In addition, CD4 T cell epitope vaccination of immunocompetent mice reduced MCMV replication in the same organs where CD4 cytotoxic T lymphocyte (CTL) activity was observed. Together, our studies show that MCMV epitope-specific CD4 T cells have the potential to mediate antiviral defense by multiple effector mechanisms in vivo. C ytomegalovirus (CMV)/human herpesvirus 5 (HHV-5) (the prototypic betaherpesvirus) infection is endemic in humans and wild mice and establishes a lifelong infection in the absence of acute disease in healthy hosts. Adaptive immunity is a critical component of CMV defenses, restricting primary infection and dampening reactivation, helping to maintain the largely benign host-virus equilibrium. However, if immunity is naive or compromised (e.g., in transplant recipients or congenital infection), CMV can cause serious disease (1, 2). In immunocompetent mice, CD8 T cells help to control acute CMV infection and establish latency (3, 4), and their adoptive transfer prevents disease in mice and humans with weakened immunity (5-8). Although much less well studied, CD4 T cells also contribute to defense against CMV. Their rapid expansion and numbers correlate with reduced disease in transplant and HIV patients (9-11), and correspondingly, delayed induction of CMV-specific CD4 T cells is associated with increased congenital infection (12) and prolonged viral shedding in infants (13). Notably, no protective correlates were seen with CMV-specific CD8 T cell responses in several of these studies. In mice, CD4 T cells are absolutely required to control mouse cytomegalovirus (MCMV) replication in the salivary glands-the key site of viral dissemination, where CD8 T cells can exert no control (14, 15)-and also contribute to immune control in several other organs. Adoptive transfer of MCMV-specific transgenic CD4 T cells provides some protection in immunocompromised mice (16), and cotransferring CMV-specific CD4 T cells reduces viral load and promotes "help" for CMV-specific CD8 T cell responses in patients receiving cellular immunotherapy (17).In addition to traditional helper functions, CD4 T cells can also mediate direct antiviral activity in some cases. In chronic human CMV (HCMV), Epstein-Barr virus (EBV), and HIV infections, CD4 T cells displaying a terminally differentiated effector phenotype and/or expressing canonical cytolytic molecules (e.g., granzymes and perforin) are pres...

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confidence: 99%

Cytomegalovirus-Specific CD4 T Cells Are Cytolytic and Mediate Vaccine Protection

Verma

Weiskopf

Gupta

et al. 2016

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“…In both the A129 and AG129 mice, DENV was cleared between days 4 and 6 postinfection at approximately the same rate in all tissues examined, with the exception of the CNS. This suggests that extraneural clearance is independent of IFN-␥R, although it is dependent on CD8 ϩ T cells (48,50). However, A129 mice, but not AG129 mice, clear DENV from the brain in an IFN-␥R-dependent manner.…”

Section: Discussionmentioning

confidence: 91%

“…Perhaps DENV's extreme susceptibility to the murine type I IFN-signaling pathway (33,34), the molecules of which differ sufficiently from human analogues to prevent DENV-mediated inhibition (3), limits extraneural antigen availability and results in a blunted cellular immune response. Accordingly, we have observed that the breadth and magnitude of DENV-specific T cell responses in wild-type mice are lower than in IFN-␣/␤ receptor-deficient mice (45,48,50). Combined with the immune-privileged status of the CNS and the capacity of DENV for infection of the CNS, a blunted T cell response may favor efficient DENV replication in the CNS.…”

Section: Discussionmentioning

confidence: 96%

Gamma Interferon (IFN-γ) Receptor Restricts Systemic Dengue Virus Replication and Prevents Paralysis in IFN-α/β Receptor-Deficient Mice

Prestwood

Morar

Zellweger

et al. 2012

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101

108

T he four serotypes of dengue virus (DENV1 to -4) belong to the Flaviviridae family, which also includes West Nile virus (WNV), St. Louis encephalitis virus (SLEV), Japanese encephalitis virus (JEV), and yellow fever virus (YFV). More than 2.5 billion people live in areas where DENV is endemic, and approximately 50 million people are infected each year (17). DENV generally manifests as an acute systemic infection that is cleared within 14 days (46). In most cases, infected individuals develop a febrile illness (dengue fever [DF]) characterized by flu-like symptoms, often accompanied by retro-orbital pain, ostealgia, and maculopapular rash. In nearly 500,000 cases per year, patients develop severe vascular leakage, which may result in hypovolemic shock (dengue hemorrhagic fever/dengue shock syndrome [DHF/ DSS]), ultimately leading to fatality rates as high as 10 to 15% in some countries or more than 25,000 deaths worldwide each year (15,17). In a subset of both DF and DHF/DSS cases (16), DENV infection was linked to encephalitis and signs of encephalopathy, including lethargy, confusion, seizure, and coma, as well as delayed neurological symptoms, such as paralysis of extremities, loss of sensation, and psychosis (2,21,23,30,36,38,43). DENVinduced neurological disease is being increasingly recognized as an important component of dengue disease in humans independent of DF and DHF/DSS (24). The prevalence of DENV-induced neurological disease has been estimated to be 4.2% of DENV cases (44) and as high as 18% of undiagnosed suspected viral central nervous system (CNS) infections in regions where DENV is endemic (21, 23).In the last decade, DENV has increasingly been linked to neurological disease in both the presence and absence of DF. Viruses of the Flaviviridae family generally cluster in a phylogenetic pattern that mirrors the typical disease manifestations they cause (13); for example, encephalitic flaviviruses cluster together. DENV, however, usually causes hemorrhagic illness but is genetically more homologous to the encephalitic viruses, including WNV, SLEV, and JEV, than it is to other hemorrhagic flaviviruses, such as YFV, based on several analyses of the nonstructural (NS) proteins NS3 and NS5 (11, 13). The close relationship of DENV to encephalitic flaviviruses may help explain the capacity of DENV to cause neurological disease and, possibly, infection of the CNS in humans (6, 37). Additionally, DENV-induced paralysis has been reported in numerous mouse models (4, 49), but only a few studies have investigated this phenomenon (1,18,20). Presently, whether DENV infects the CNS in natural human infections continues to be a subject of debate (25,30,32).Mice lacking the alpha/beta interferon and gamma interferon receptors (IFN-␣/␤R and IFN-␥R) in the 129/Sv genetic background (AG129) are highly susceptible to disease resembling human DHF/DSS, as well as paralysis, following infection with mouse-passaged variants of DENV2 clinical isolate PL046 (5,35,40,51). In contrast, congenic mice lacking only IFN-␣/␤R (A12...

“…It has also been shown that interferon is one of three major barriers that limit poliovirus trafficking from the periphery to the central nervous system (CNS), and knocking out part of the interferon response allowed a greater portion of the viral population to enter the brain (30). Interferon deficiency of AG129 mice aids in the systemic spread of viral infections (31,32) while maintaining humoral and cell-mediated immunity (33)(34)(35). In our model, eliminating interferon allowed the virus to spread to more organs, including the brain.…”

Section: Figmentioning

confidence: 99%

A Single Mutation in the VP1 of Enterovirus 71 Is Responsible for Increased Virulence and Neurotropism in Adult Interferon-Deficient Mice

Caine

Moncla

Ronderos

et al. 2016

Hand, foot, and mouth disease (HFMD) has spread throughout the Asia-Pacific region, affecting millions of young children, who develop symptoms ranging from painful blisters around their mouths and hands to neurological complications. Many members of the genus Enterovirus (family Picornaviridae) cause HFMD. Enterovirus 71 (EV71) is one of the primary causative agents and has been linked to severe disease. Vaccine efficacy and pathogenesis studies for EV71 have been limited because there is a lack of suitable animal models. Previously, we generated a mouse-adapted EV71 (mEV71) capable of infecting 12-week-old interferon receptor-deficient AG129 mice and used the model to evaluate the efficacy of candidate HFMD vaccines. Here, we present data investigating the genetic correlates of EV71 adaptation and characterize the virus's tissue tropism in mice. Using reverse genetics, a VP1 mutation (K244E) was shown to be necessary for mEV71 virulence in adult mice. Another VP1 mutation (H37R) was required for mEV71 recovery on rhabdomyosarcoma (RD) cells. Viral loads determined by real-time reverse transcription (RT)-PCR confirmed the presence of mEV71 in the sera and multiple organs of mice. Histological analysis revealed signs of meningitis and encephalitis, characteristic of severe human disease. The further description of this model has provided insight into EV71 pathogenesis and demonstrates the importance of the VP1 region in facilitating mEV71 adaptation. IMPORTANCEEV71 is a reemerging pathogen, and little is known about the genetic determinants involved in its pathogenesis. The absence of animal models has contributed to this lack of knowledge. The data presented here improve upon the existing animal models by characterizing a mouse-adapted strain of EV71. We determined that a VP1 mutation (K244E) was needed for EV71 virulence in adult AG129 mice. While this mutation was found previously for EV71 adaptation in 5-day-old BALB/c mice, neurotropic disease did not develop. Using interferon-deficient mice, we raised the age of susceptibility beyond 6 weeks and provided clear evidence that our model mimics severe human infections. The model can be exploited to identify determinants of EV71 virulence and to reveal molecular mechanisms that control the virus-host interaction, especially those associated with neurotropic disease. Furthermore, these data provide useful information regarding the importance of VP1, specifically position 244, in host adaptation and tissue dissemination. O ver the past decade, enterovirus 71 (EV71) (family Picornaviridae) was identified as one of the main causative agents responsible for large outbreaks of hand, foot, and mouth disease (HFMD) across the Asia-Pacific region. Other members of the genus Enterovirus cause HFMD, including coxsackieviruses A16, A6, A5, A7, A9, A10, B2, and B5, but EV71 has been linked to severe complications, including brainstem encephalitis, aseptic meningitis, and pulmonary edema (1-3). Picornaviruses, including the enteroviruses, have positive-sense, single-s...