Adenovirus Infection Triggers a Rapid, MyD88-Regulated Transcriptome Response Critical to Acute-Phase and Adaptive Immune Responses In Vivo

Abstract

Nearly 50 years ago, the discovery of interferon prompted the notion that host cells innately respond to viral invasion. Since that time, technological advances have allowed this response to be extensively characterized and dissected in vitro. However, these advances have only recently been applied to highly complex, in vivo biological systems. To this end, we exploited high-titer adenovirus (Ad) vectors to globally investigate the innate immune response to nonenveloped viral infection in vivo. Our results indicated a potent cellular transcriptome response shortly after infection, with global assessments revealing significant dysregulation in ϳ15% of the measured transcripts derived from Ad vector-transduced tissue. Bioinformatics-based transcriptome analysis revealed a complex innate response to Ad infection, with induction of proinflammatory responses (and suppression of metabolism and mitochondrial genes) akin to those observed when mice are challenged with lipopolysaccharide. Despite this commonality, there were many unique aspects of the Ad-dependent transcriptome response, including the upregulation of several RNA regulatory mechanisms and apoptosisrelated pathways, accompanied by the suppression of lysosomal and endocytic genes. Our results also implicated the Toll-like receptors (TLRs) in these responses, prompting specific investigations into this pathway. By using MyD88KO mice, our results confirmed that Ad-induced dysregulation of five functionally related gene clusters are significantly dependent on this TLR adaptor gene. MyD88 deficiency also resulted in significantly diminished, although not abolished, adaptive and acute-phase immune responses to Ad, confirming the transcriptome data, as well as specifically identifying MyD88 as a significant Ad immunity amplifier and regulator in vivo.For the past 100 years, a significant body of research has sought to understand the mammalian innate immune system, with recent efforts focusing on understanding its molecular workings (5). The discovery of a new family of "pathogen-sensing" genes, the Toll-like receptors (TLRs) and the deciphering of their role in mammalian innate immunity has been an especially robust area of recent investigation (2, 28). However, critical issues remain unanswered, chief among which is the intracellular innate response to viral infections in vivo. While recent studies have sought to investigate intracellular viral immunity and TLR involvement, almost all have relied on tissue culture systems, with little or no attention paid to in vivo models.

Wild-type adenoviruses from groups A–F evoke unique innate immune responses, of which HAd3 and SAd23 are partially complement dependent

“…From these analyses, it became clear that many known innate immunity genes, such as NOD-like receptors and TLRs were induced following Ad injection, and that TLR adaptor proteins, such as MyD88 and TRIF were required in Ad-specific innate immune responses. 9,36 Furthermore, Ad-induced type I IFNs as well as IL-6 and IL-12 have been found to be partially dependent on -and HAd41-injected animals, n ¼ 3 for all tissues at both 6 and 24 h.p.i. n ¼ 4 at both time points for HAd5(P)-injected animals.…”

“…Our recent publications describe the specific transcriptome profile of Ad-infected mouse embryonic fibroblasts in vitro, and the general transcriptome responses in the livers of Ad-injected C57BL/6 mice in vivo. 9,10,36 The results of these array-based analyses indicated a global induction of host immune response genes, inclusive of several immune response gene networks, such as TLR pathways among others. From these analyses, it became clear that many known innate immunity genes, such as NOD-like receptors and TLRs were induced following Ad injection, and that TLR adaptor proteins, such as MyD88 and TRIF were required in Ad-specific innate immune responses.…”

“…[37][38][39] In light of these data, we analyzed the expression of a selected subset of these Ad-induced genes to determine whether alternative Ad serotypes influenced the expression of these genes in a manner similar to that previously noted for HAd5 ( Our previous studies showed that maximal changes in liver transcriptome responses occur 6 h after Ad injection. 9 The expression of most of these genes (except for TLR9, NAÀN=1 for data generated from this primer pair. ANOVA followed by a Dunnet's post hoc test was completed to determine a statistical difference versus mock injected animals.…”

“…9,11,36 This innate inflammatory response 'profile' has implications in the use of Ad as both a vaccine as well as a vector for gene therapy applications. For this reason, we characterized the acute inflammatory response induced by each of the various Ad serotypes and compared those responses to those typically induced by HAd5 (Figure 2).…”

“…injection of high-titer HAd5-based vectors can cause several innate immune responses and hepatotoxicities, some associated with activation of complement and Toll-like receptor (TLR) pathways. [9][10][11] These responses can include induction of thrombocytopenia, and expression of a spectrum of cytokines and chemokines including interleukin (IL)-1a, IL-6, IL-12, keratinocyte chemoattractant (KC), monocyte chemotactic protein-1 (MCP-1), granulocyte-colony stimulating factor (G-CSF), interferon (IFN)-g, macrophage inflammatory protein-1 (MIP-1)b and RANTES (regulated on activation normal T-cell-expressed and -secreted).…”