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Little is known about the role of viral genes in modulating host cytokine responses. Here we report a new functional role of the viral encoded IE1 protein of the murine cytomegalovirus in sculpting the inflammatory response in an acute infection. In time course experiments of infected primary macrophages (MΦs) measuring cytokine production levels, genetic ablation of the immediate-early 1 (ie1) gene results in a significant increase in TNFα production. Intracellular staining for cytokine production and viral early gene expression shows that TNFα production is highly associated with the productively infected MΦ population of cells. The ie1- dependent phenotype of enhanced MΦ TNFα production occurs at both protein and RNA levels. Noticeably, we show in a series of in vivo infection experiments that in multiple organs the presence of ie1 potently inhibits the pro-inflammatory cytokine response. From these experiments, levels of TNFα, and to a lesser extent IFNβ, but not the anti-inflammatory cytokine IL10, are moderated in the presence of ie1. The ie1- mediated inhibition of TNFα production has a similar quantitative phenotype profile in infection of susceptible (BALB/c) and resistant (C57BL/6) mouse strains as well as in a severe immuno-ablative model of infection. In vitro experiments with infected macrophages reveal that deletion of ie1 results in increased sensitivity of viral replication to TNFα inhibition. However, in vivo infection studies show that genetic ablation of TNFα or TNFRp55 receptor is not sufficient to rescue the restricted replication phenotype of the ie1 mutant virus. These results provide, for the first time, evidence for a role of IE1 as a regulator of the pro-inflammatory response and demonstrate a specific pathogen gene capable of moderating the host production of TNFα in vivo., Author Summary The suppression of the production rather than the blockage of action of the potent inflammatory mediator TNFα is a particular hallmark of anti-TNFα mechanisms associated with microbial and parasitic infections. Whether this mode of counter-regulation is an important feature of infection by viruses is not clear. Also, it remains to be determined whether a specific pathogen gene in the context of an infection in vivo is capable of modulating levels of TNFα production. In this study we disclose a virus-mediated moderation of TNFα production, dependent on the ie1 gene of murine cytomegalovirus (MCMV). The ie1 gene product IE1 is a well-characterized nuclear protein capable of altering levels of host and viral gene expression although its biological role in the context of a natural infection is to date unknown. We provide evidence showing that ie1 is associated with a moderated pro-inflammatory cytokine response, in particular with TNFα production. Further, we show that the viral moderation of this cytokine is not only readily apparent in vitro but also in the natural host. The identification of a viral gene responsible for this mode of regulation in vivo may have therapeutic potential in the future in both anti-viral and anti-inflammatory strategies.

One of the most famous cytokines, tumor necrosis factor (TNF), shows a vast spectrum of bioactivities influencing majority of cell types. Produced mostly by macrophages, it is considered to be a major mediator of inflammation and cellular immune response. Although infection with several viruses has been shown to induce production of TNF, indicating its importance in successful viral clearance, it seems that not many viruses have learned how to control this potent cytokine. However, considering their reputation as masters of disguise, it does not come as a surprise that both human and murine cytomegaloviruses (MCMV) have developed means to avoid TNF-induced response (caspase dependent and independent apoptosis and expression of TNF-induced genes). Here it is shown that the MCMV ie1 gene product acts as a negative regulator of TNF expression in vitro and in vivo at both transcriptional and translational level, thus revealing yet another immunoevasive strategy employed by MCMV. Mice infected with MCMVdie1 show lower viral titers, but higher levels of TNF when compared to mice infected with MCMVdie1-rev or wt virus and this effect is not strain dependant. NK cell depletion didn’t rescue the virus from its attenuated phenotype, indicating that this immunoevasion strategy is directed mostly at macrophages, which in addition of being the main producers of TNF, are also a major site of viral replication, dissemination and establishment of latency. Interestingly, despite the attenuated phenotype, growth kinetics of MCMVdie1 showed reactivation around 30th day PI in spleen, liver, kidney and, most prominently, in salivary gland.

A study was carried out to investigate the potential for an oil-absorber placed upstream to a bioscrubber configuration to act as buffer for sequentially alternating loads of 1,2-dichloroethane (DCE) and fluorobenzene (FB) in waste gas streams. A bioscrubber system (without the oil-absorber present) was inoculated with strain F11 and Xanthobacter autotrophicus strain GJ10 and was submitted to sequential alternations of the pollutant fed (DCE and FB). The re-introduction of the substrates after each cycle caused system instability. An oil-absorber, utilizing a cost effective and environmentally friendly absorbent (Sunflower Oil), was introduced prior to the bioscrubber to control the negative effects caused by the sequential switches in the feed composition between DCE and FB. The performance of the combined Oil-Absorber-Bioscubber system was stable after the substrates re-introduction following each switch in the feed. The combined Oil-Absorber-Bioscrubber system is a robust technology, offering an effective solution to the biological treatment of waste-gas streams. This is an abstract of a paper presented at the 2006 AIChE Annual Meeting (San Francisco, CA 11/12-17/2006).