Vinyl chloride (VC) is a ubiquitous environmental contaminant for which human risk is incompletely understood. We have previously reported that high occupational exposure to VC directly caused liver damage in humans. However, whether VC may also potentiate liver injury from other causes is not known. C57Bl/6J mice were administered chloroethanol (CE), a major metabolite of VC, and lipopolysaccharide (LPS) 24 h after chloroethanol. Samples were harvested for determination of liver damage, inflammation and changes in carbohydrate and lipid metabolism. In mice, CE exposure alone caused no detectable liver damage. LPS exposure caused inflammatory liver damage, oxidative stress, lipid accumulation, and glycogen depletion; the effect of all of these variables was potentiated by CE pre-exposure. In vitro experiments suggest that VC metabolite chloroacetaldehyde (CAA) directly damages mitochondria, which may explain the sensitization effect observed in vivo. Moreover, co-exposure of cells to CAA and TNFα caused increased cell death, supporting the hypothesis of sensitization by VC metabolites. Taken together, these data demonstrate that exposure to VC/metabolites at levels that are not overtly hepatotoxic can potentiate liver injury caused by another hepatotoxicant. This serves as proof-of-concept that VC hepatotoxicity may be modified by an additional metabolic stress such as endotoxemia, which commonly occurs in acute (e.g., sepsis) and chronic (e.g., NAFLD) diseases.

The ongoing circulation of influenza A H5N1 in the United States has raised concerns of a pandemic caused by highly pathogenic avian influenza. Although the United States has stockpiled and is prepared to produce millions of vaccine doses to address an H5N1 pandemic, currently circulating H5N1 viruses contain multiple mutations within the immunodominant head domain of hemagglutinin (HA) compared to the antigens used in stockpiled vaccines. It is unclear if these stockpiled vaccines will need to be updated to match the contemporary H5N1 strains. Here we show that a replicating RNA vaccine expressing the HA of an H5N1 isolated from a US dairy cow confers complete protection against homologous lethal challenge in mice. A repRNA encoding the HA of a clade 1 H5 from 2004 (A/Vietnam/1203/2004) as utilized by some stockpiled vaccines, confers only partial protection. Our data highlight the utility of nucleic acid vaccines to be rapidly updated to match emergent viruses of concern while demonstrating that contemporary bovine H5N1 viruses can evade immunity elicited by historical HA antigens.

The recent emergence and rapid response to severe acute respiratory syndrome coronavirus 2 was enabled by prototype pathogen and vaccine platform approaches, driven by the preemptive application of RNA vaccine technology to the related Middle East respiratory syndrome coronavirus. Recently, the National Institutes of Allergy and Infectious Diseases identified nine virus families of concern, eight enveloped virus families and one nonenveloped virus family, for which vaccine generation is a priority. Although RNA vaccines have been described for a variety of enveloped viruses, a roadmap for their use against nonenveloped viruses is lacking. Enterovirus D68 was recently designated a prototype pathogen within the family Picornaviridae of nonenveloped viruses because of its rapid evolution and respiratory route of transmission, coupled with a lack of diverse anti-enterovirus vaccine approaches in development. Here, we describe a proof-of-concept approach using a clinical stage RNA vaccine platform that induced robust enterovirus D68–neutralizing antibody responses in mice and nonhuman primates and prevented upper and lower respiratory tract infections and neurological disease in mice. In addition, we used our platform to rapidly characterize the antigenic diversity within the six genotypes of enterovirus D68, providing the necessary data to inform multivalent vaccine compositions that can elicit optimal breadth of neutralizing responses. These results demonstrate that RNA vaccines can be used as tools in our pandemic-preparedness toolbox for nonenveloped viruses.

Chronic alcohol exposure is a clinically important risk factor for the development of acute respiratory distress syndrome, the most severe form of acute lung injury (ALI). However, the mechanisms by which alcohol sensitizes the lung to development of this disease are poorly understood. We determined the role of the antifibrinolytic protein plasminogen activator inhibitor-1 (PAI-1) in alcohol enhancement of experimental endotoxin-induced ALI. Wild-type, PAI-1−/−, and integrin β3−/− mice were fed ethanol-containing Lieber-DeCarli liquid or a control diet for 6 weeks, followed by systemic LPS challenge. LPS administration triggered coagulation cascade activation as evidenced by increased plasma thrombin-antithrombin levels and pulmonary fibrin deposition. Ethanol-exposed animals showed enhanced PAI-1 expression and pulmonary fibrin deposition with coincident exaggeration of pulmonary inflammatory edematous injury. PAI-1 deficiency markedly reduced pulmonary fibrin deposition and greatly reduced inflammation and injury without impacting upstream coagulation. Interestingly, pulmonary platelet accumulation was effectively abolished by PAI-1 deficiency in ethanol/LPS-challenged mice. Moreover, mice lacking integrin αIIBβ3, the primary platelet receptor for fibrinogen, displayed a dramatic reduction in early inflammatory changes after ethanol/LPS challenge. These results indicate that the mechanism whereby alcohol exaggerates LPS-induced lung injury requires PAI-1–mediated pulmonary fibrin accumulation, and suggest a novel mechanism whereby alcohol contributes to inflammatory ALI by enhancing fibrinogen-platelet engagement.

Viral hemorrhagic fevers (VHFs) encompass a group of diseases with cardinal symptoms of fever, hemorrhage, and shock. The liver is a critical mediator of VHF disease pathogenesis and high levels of ALT/AST transaminases in plasma correlate with poor prognosis. In fact, Lassa Fever (LF), the most prevalent VHF in Africa, was initially clinically described as hepatitis. Previous studies in non-human primate (NHP) models also correlated LF pathogenesis with a robust proliferative response in the liver. The purpose of the current study was to gain insight into the mechanism of liver injury and to determine the potential role of proliferation in LF pathogenesis. C57Bl/6J mice were infected with either the pathogenic (for NHPs) strain of lymphocytic choriomeningitis virus (LCMV, the prototypic arenavirus), LCMV-WE, or with the non-pathogenic strain, LCMV-ARM. As expected, LCMV-WE, but not ARM, caused a hepatitis-like infection. LCMV-WE also induced a robust increase in the number of actively cycling hepatocytes. Despite this increase in proliferation, there was no significant difference in liver size between LCMV-WE and LCMV-ARM, suggesting that cell cycle was incomplete. Indeed, cells appeared arrested in the G₁ phase and LCMV-WE infection increased the number of hepatocytes that were simultaneously stained for proliferation and apoptosis. LCMV-WE infection also induced expression of a non-conventional virus receptor, AXL-1, from the TAM (TYRO3/AXL/MERTK) family of receptor tyrosine kinases and this expression correlated with proliferation. Taken together, these results shed new light on the mechanism of liver involvement in VHF pathogenesis. Specifically, it is hypothesized that the induction of hepatocyte proliferation contributes to expansion of the infection to parenchymal cells. Elevated levels of plasma transaminases are likely explained, at least in part, by abortive cell cycle arrest induced by the infection. These results may lead to the development of new therapies to prevent VHF progression.