The myeloperoxidase (MPO) system of activated phagocytes is central to normal host defense mechanisms, and dysregulated MPO contributes to the pathogenesis of inflammatory disease states ranging from atherosclerosis to cancer. Here we show that upon systemic administration, the small molecule luminol enables noninvasive bioluminescence imaging (BLI) of MPO activity in vivo. Luminol-BLI allowed quantitative longitudinal monitoring of MPO activity in animal models of acute dermatitis, mixed allergic contact hypersensitivity, focal arthritis and spontaneous large granular lymphocytic tumors. Bioluminescence colocalized with histological sites of inflammation and was totally abolished in gene-deleted Mpo −/− mice, despite massive tissue infiltration of neutrophils and activated eosinophils, indicating that eosinophil peroxidase did not contribute to luminol-BLI in vivo. Thus, luminol-BLI provides a noninvasive, specific and highly sensitive optical readout of phagocyte-mediated MPO activity in vivo and may enable new diagnostic applications in a wide range of acute and chronic inflammatory conditions. The heme-containing enzyme MPO is a key component of the cytotoxic armamentarium of phagocytic white blood cells 1,2 . MPO is by far the most abundant protein product in azurophilic granules of neutrophils (5%), constitutes approximately 1% of monocyte protein and is found in the lysosomes of other polymorphonuclear leukocytes and macrophages. The phagosomal oxidative burst is initiated by a stimulus-dependent assembly of the phagocytic NADPH oxidase (Phox), a multimeric protein complex located on the phagosomal membrane. Phox then reduces molecular oxygen to produce superoxide anion (O 2•− ), which further dismutates to yield the relatively unreactive hydrogen peroxide (H 2 O 2 ) 1 . Upon phagocytic activation, large quantities of active MPO are secreted into phagosomes, catalyzing the production of highly bactericidal hypochlorous acid (HOCl) with H 2 O 2 and chloride ions (Cl − ) as substrates (Fig. 1a) 1 .

The investigators present their analysis of primary cells from patients with human T-cell leukemia virus 1–associated adult T-cell leukemia/lymphoma treated in a phase 2 clinical trial with nivolumab to elucidate mechanisms of hyperprogression that halted the trial after just 3 patients received a single treatment.

The rapid spread of herpes simplex virus type 1 (HSV-1) in mucosal epithelia and neuronal tissue depends primarily on the ability of the virus to navigate within polarized cells and the tissues they constitute. To understand HSV entry and the spread of virus across cell junctions, we have previously characterized a human keratinocyte cell line, HaCaT. These cells appear to reflect cells infected in vivo more accurately than many of the cultured cells used to propagate HSV. HSV mutants lacking gE/gI are highly compromised in spread within epithelial and neuronal tissues and also show defects in cell-to-cell spread in HaCaT cells, but not in other, nonpolarized cells. HSV gD is normally considered absolutely essential for entry and cell-to-cell spread, both in cultured cells and in vivo. Here, an HSV-1 gD mutant virus, F-US6kan, was found to efficiently enter HaCaT cells and normal human keratinocytes and could spread from cell to cell without gD provided by complementing cells. By contrast, entry and spread into other cells, especially highly transformed cells commonly used to propagate HSV, were extremely inefficient. Further analyses of F-US6kan indicated that this mutant expressed extraordinarily low (1/500 wild-type) levels of gD. Neutralizing anti-gD monoclonal antibodies inhibited entry of F-US6kan, suggesting F-US6kan utilized this small amount of gD to enter cells. HaCaT cells expressed high levels of an HSV gD receptor, HveC, and entry of F-US6kan into HaCaT cells could also be inhibited with antibodies specific for HveC. Interestingly, anti-HveC antibodies were not fully able to inhibit entry of wild-type HSV-1 into HaCaT cells. These results help to uncover important properties of HSV and human keratinocytes. HSV, with exceedingly low levels of a crucial receptor-binding glycoprotein, can enter cells expressing high levels of receptor. In this case, surplus gD may be useful to avoid neutralization by anti-gD antibodies.

Herpes simplex virus type 1 (HSV-1) glycoprotein D (gD) is an essential component of the entry apparatus that is responsible for viral penetration and subsequent cell-cell spread. To test the hypothesis that gD may serve distinguishable functions in entry of free virus and cell-cell spread, mutants were selected for growth on U S 11cl19.3 cells, which are resistant to both processes due to the lack of a functional gD receptor, and then tested for their ability to enter as free virus and to spread from cell to cell. Unlike their wild-type parent, HSV-1(F), the variants that emerged from this selection, which were named SP mutants, are all capable of forming macroscopic plaques on the resistant cells. This ability is caused by a marked increase in cell-cell spread without a concomitant increase in efficiency of entry of free virus. gD substitutions that arose within these mutants are sufficient to mediate cell-cell spread in U S 11cl19.3 cells but are insufficient to overcome the restriction to entry of free virions. These results suggest that mutations in gD (i) are sufficient but not necessary to overcome the block to cell-cell spread exhibited by U S 11cl19.3 cells and (ii) are insufficient to mediate entry of free virus in the same cells.Herpes simplex virus (HSV) can enter a naive host cell by either of two distinct methods. Extracellular virions present during primary infection can enter cells in exposed tissue by entry of free virus. Once a cell is initially infected, subsequent infections can begin by lateral spread from the initially infected cell to adjacent uninfected neighbors by cell-cell spread. Both types of entry are important for sustained viral infection. The ability of a virus to spread from cell to cell provides a powerful advantage in vivo since it is able to avoid the strong humoral immune response elicited to extracellular HSV virions. The essential roles of HSV glycoproteins in entry of free virus and cell-cell spread remain poorly characterized.HSV entry requires a complicated cascade of virus-cell interactions. Initially, an interaction between cellular heparan sulfate proteoglycan and viral glycoprotein C (gC) and/or gB results in attachment of the virion to the cell surface (15,16,20,47,56). Fusion between the viral membrane and the cellular plasma membrane is not triggered until a secondary interaction occurs between gD and a cellular receptor (12,17,21,54). The fusion event itself requires the coordinated function of the viral glycoproteins gB, gD, gH, and gL. These four proteins play essential roles in both entry of free virus and cell-cell spread (3,11,43). These two processes are thus closely related. However, entry of free virus and cell-cell spread can be distinguished in HSV and in related alphaherpesviruses by their differing dependence on specific viral genes and by their differing sensitivities to mutations in the viral genome. Deletion of the viral glycoproteins gE and gI provides a clear distinction between entry of free virus and cell-cell spread since it significantly red...