Interferon Control of Neurotropic Viral Infections.

Interferons (IFNs) comprise a pleiotropic family of signaling molecules that are often the first line of defense against viral infection. Inflammatory responses induced by IFN are generally well tolerated during peripheral infections; yet, the same degree of inflammation during infection of the central nervous system (CNS) could be catastrophic. Thus, IFN responses must be modified within the CNS to ensure host survival. In this review, we discuss emerging principles highlighting unique aspects of antiviral effects of IFN protection following neurotropic viral infection, chiefly using new techniques in rodent models. Evaluation of these unique responses provides insights into how the immune system eradicates or controls pathogens, while avoiding host damage. Defining these principles may have direct impact on the development of novel clinical approaches.

Immune-Mediated Control of a Dormant Neurotropic RNA Virus Infection.

Genomic material from many neurotropic RNA viruses (e.g., measles virus [MV], West Nile virus [WNV], Sindbis virus [SV], rabies virus [RV], and influenza A virus [IAV]) remains detectable in the mouse brain parenchyma long after resolution of the acute infection. The presence of these RNAs in the absence of overt central nervous system (CNS) disease has led to the suggestion that they are viral remnants, with little or no potential to reactivate. Here we show that MV RNA remains detectable in permissive mouse neurons long after challenge with MV and, moreover, that immunosuppression can cause RNA and protein synthesis to rebound, triggering neuropathogenesis months after acute viral control. Robust recrudescence of viral transcription and protein synthesis occurs after experimental depletion of cells of the adaptive immune response and is associated with a loss of T resident memory (Trm) lymphocytes within the brain. The disease associated with loss of immune control is distinct from that seen during the acute infection: immune cell-depleted, long-term-infected mice display severe gait and motor problems, in contrast to the wasting and lethal disease that occur during acute infection of immunodeficient hosts. These results illuminate the potential consequences of noncytolytic, immune-mediated viral control in the CNS and demonstrate that what were once considered "resolved" RNA viral infections may, in fact, induce diseases later in life that are distinct from those caused by acute infection.IMPORTANCE Viral infections of neurons are often not cytopathic; thus, once-infected neurons survive, and viral RNAs can be detected long after apparent viral control. These RNAs are generally considered viral fossils, unlikely to contribute to central nervous system (CNS) disease. Using a mouse model of measles virus (MV) neuronal infection, we show that MV RNA is maintained in the CNS of infected mice long after acute control and in the absence of overt disease. Viral replication is suppressed by the adaptive immune response; when these immune cells are depleted, viral protein synthesis recurs, inducing a CNS disease that is distinct from that observed during acute infection. The studies presented here provide the basis for understanding how persistent RNA infections in the CNS are controlled by the host immune response, as well as the pathogenic consequences of noncytolytic viral control.

Interleukin-36ß provides protection against HSV-1 infection, but does not modulate initiation of adaptive immune responses.

Interleukin-36 (IL-36) represents three cytokines, IL-36α, IL-36ß and IL-36γ, which bind to the same receptor, IL-1RL2; however, their physiological function(s) remain poorly understood. Here, the role of IL-36 in immunity against HSV-1 was examined using the flank skin infection mouse model. Expression analyses revealed increased levels of IL-36α and IL-36ß mRNA in infected skin, while constitutive IL-36γ levels remained largely unchanged. In human keratinocytes, IL-36α mRNA was induced by HSV-1, while IL-1ß and TNFα increased all three IL-36 mRNAs. The dominant alternative splice variant of human IL-36ß mRNA was isoform 2, which is the ortholog of the known mouse IL-36ß mRNA. Mice deficient in IL-36ß, but not IL-36α or IL-36γ, succumbed more frequently to HSV-1 infection than wild type mice. Furthermore, IL-36ß-/- mice developed larger zosteriform skin lesions along infected neurons. Levels of HSV-1 specific antibodies, CD8+ cells and IFNγ-producing CD4+ cells were statistically equal in wild type and IL-36ß-/- mice, suggesting similar initiation of adaptive immunity in the two strains. This correlated with the time at which HSV-1 genome and mRNA levels in primary skin lesions started to decline in both wild type and IL-36ß-/- mice. Our data indicate that IL-36ß has previously unrecognized functions protective against HSV-1 infection.

Unprocessed Interleukin-36α Regulates Psoriasis-Like Skin Inflammation in Cooperation With Interleukin-1.

Generalized pustular psoriasis is a severe skin disease characterized by epidermal hyperplasia, neutrophil-rich abscesses within the epidermis, and a mixed inflammatory infiltrate in the dermis. The disease may be caused by missense mutations in the IL-36 receptor antagonist, IL-36Ra. Curiously, the related IL-1Ra has therapeutic effects in some of these latter patients. Here, using an experimental mouse model of psoriasiform skin inflammation, we demonstrate in vivo connections between IL-36 and IL-1 expression. After disease initiation, IL-36α-deficient mice exhibited dramatically diminished skin pathology, including absence of epidermal neutrophils, reduced keratinocyte acanthosis, and less dermal edema. In contrast, IL-36ß and IL-36γ knockout mice developed disease indistinguishable from that of wild-type mice. The endogenous IL-36α was not processed through proteolysis. Although IL-36α expression was strongly induced in an IL-1 signaling-dependent manner during disease, expression of IL-1α was also dependent upon IL-36α. Hence, after being upregulated by IL-1α, IL-36α acts through a feedback mechanism to boost IL-1α levels. Analyses of double knockout mice further revealed that IL-36α and IL-1α cooperate to promote psoriasis-like disease. In conclusion, IL-1α and IL-36α form a self-amplifying inflammatory loop in vivo that in patients with insufficient counter regulatory mechanisms may become hyper-engaged and/or chronic.

Interleukin-1α released from HSV-1-infected keratinocytes acts as a functional alarmin in the skin.

Herpes simplex virus-1 (HSV-1) is a human pathogen that utilizes several strategies to circumvent the host immune response. An immune evasion mechanism employed by HSV-1 is retention of interleukin-1ß (IL-1ß) in the intracellular space, which blocks the pro-inflammatory activity of IL-1ß. Here we report that HSV-1-infected keratinocytes actively release the also pro-inflammatory IL-1α, preserving the ability of infected cells to signal danger to the surrounding tissue. The extracellular release of IL-1α is independent of inflammatory caspases. In vivo recruitment of leukocytes to early HSV-1 microinfection sites within the epidermis is dependent upon IL-1 signalling. Following cutaneous HSV-1 infection, mice unable to signal via extracellular IL-1α exhibit an increased mortality rate associated with viral dissemination. We conclude that IL-1α acts as an alarmin essential for leukocyte recruitment and protective immunity against HSV-1. This function may have evolved to counteract an immune evasion mechanism deployed by HSV-1.

IL-1R1 signaling facilitates Munro's microabscess formation in psoriasiform imiquimod-induced skin inflammation.

Munro's microabscesses contain polymorphonuclear leukocytes and form specifically in the epidermis of psoriasis patients. The mechanism whereby the neutrophils are recruited into the epidermis is poorly understood. Using a combination of human and mouse primary keratinocyte cell cultures and the imiquimod-induced psoriasis-like mouse model of skin inflammation, we explored the role of IL-1 signaling in microabscess formation. In vitro imiquimod stimulated production of IL-1α and neutrophil recruiting chemokines. Imiquimod-activated chemokine expression was dependent upon adenosine signaling and independent of IL-1α and IL-1 receptor type 1 (IL-1R1); nevertheless, IL-1α could enhance chemokine expression initiated by imiquimod. Topical application of imiquimod in vivo led to epidermal microabscess formation, acanthosis, and increased IL-1α and chemokine expression in the skin of wild-type mice. However, in IL-1R1-deficient mice these responses were either absent or dramatically reduced. These results demonstrate that IL-1α and IL-1R1 signaling is essential for microabscess formation, neutrophil recruiting chemokine expression, and acanthosis in psoriasis-like skin inflammation induced by imiquimod.

The double-stranded RNA analogue polyinosinic-polycytidylic acid induces keratinocyte pyroptosis and release of IL-36γ.

IL-36 is the common name for the three IL-1 family members IL-36α, IL-36ß, and IL-36γ, formerly known as IL-1F6, IL-1F8, and IL-1F9, respectively. IL-36 appears to have pro-inflammatory activities; however, the physiological function of these cytokines remains unknown. Expression of IL-36 by keratinocytes implies its possible involvement in innate immune responses in the skin. We observed that, of the three IL-36 isoforms, human keratinocytes express high levels of IL-36γ. IL-36γ mRNA expression was dramatically induced by the Toll-like receptor ligands polyinosinic-polycytidylic acid (poly(I:C)) and flagellin. Surprisingly, the IL-36γ protein was released by cells treated with poly(I:C), but remained intracellular in cells treated with flagellin only. poly(I:C), but not flagellin, induced cell death and caspase-3/7 activation. Inhibition of caspase-3/7 and caspase-1 blocked extracellular release of IL-36γ from poly(I:C)-treated cells. Furthermore, caspase-1 inhibition prevented poly(I:C)-induced caspase-3/7 activation. Interestingly, transcription of the gene IL36G was dependent on caspase-1, but not caspase-3/7, activation. This demonstrates that the pathways leading to IL36G transcription and caspase-3/7 activation branch after caspase-1. This divergence of the pathways allows the cells to enter a state of de novo protein synthesis before committing to pyroptosis. Overall, our observations suggest that IL-36γ may be an alarmin that signals the cause, e.g., viral infection, of cell death.