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This paper reviews in vivo studies on the interaction between porcine reproductive and respiratory syndrome virus (PRRSV) and LPS performed in the authors’ laboratory. The main aim was to develop a reproducible model to study the pathogenesis of PRRSV-induced multifactorial respiratory disease. The central hypothesis was that respiratory disease results from an overproduction of proinflammatory cytokines in the lungs. In a first series of studies, PRRSV was shown to be a poor inducer of TNF-α and IFN-α in the lungs, whereas IL-1 and the anti-inflammatory cytokine IL-10 were produced consistently during infection. We then set up a dual inoculation model in which pigs were inoculated intratracheally with PRRSV and 3–14 days later with LPS. PRRSV-infected pigs developed acute respiratory signs for 12–24h upon intratracheal LPS inoculation, in contrast to pigs inoculated with PRRSV or LPS only. Moreover, peak TNF-α, IL-1 and IL-6 titers were 10–100 times higher in PRRSV–LPS inoculated pigs than in the singly inoculated pigs and the cytokine overproduction was associated with disease. To further prove the role of proinflammatory cytokines, we studied the effect of pentoxifylline, a known inhibitor of TNF-α and IL-1, on PRRSV–LPS induced cytokine production and disease. The clinical effects of two non-steroidal anti-inflammatory drugs (NSAIDs), meloxicam and flunixin meglumine, were also examined. Pentoxifylline, but not the NSAIDs, significantly reduced fever and respiratory signs from 2 to 6h after LPS. The levels of TNF-α and IL-1 in the lungs of pentoxifylline-treated pigs were moderately reduced, but were still 26 and 3.5-fold higher than in pigs inoculated with PRRSV or LPS only. This indicates that pathways other than inhibition of cytokine production contributed to the clinical improvement. Finally, we studied a mechanism by which PRRSV may sensitize the lungs for LPS. We hypothesized that PRRSV would increase the amount of LPS receptor complex in the lungs leading to LPS sensitisation. Both CD14 and LPS-binding protein, two components of this complex, increased significantly during infection and the amount of CD14 in particular was correlated with LPS sensitisation. The increase of CD14 was mainly due to infiltration of strongly CD14-positive monocytes in the lungs. The PRRSV–LPS combination proved to be a simple and reproducible experimental model for multifactorial respiratory disease in pigs. To what extent the interaction between PRRSV and LPS contributes to the development of complex respiratory disease is still a matter of debate. [Copyright &y& Elsevier]

The expression of IL-1β and inducible nitric oxide synthase (iNOS) from iNOS hypo (GB2, B6B6) and hyper (K-strain, B15B15) responder chickens was examined. Compared to GB2, macrophages from K-strain expressed higher iNOS mRNA as quantitated by reverse transcriptase polymerase (RT-PCR) chain reaction after stimulation with 1 μg/ml of Escherichia coli (E. coli) lipopolysaccharide (LPS). On the contrary, IL-1β mRNA expression was comparable between K and GB2 macrophages at 3 h post-LPS stimulation but persisted up to 9 h only in GB2 macrophages. The LPS-inducible interleukin-1 (IL-1) surface receptor expression, measured by flow cytometry, was higher in GB2 than on K-strain macrophages. Blocking of IL-1 receptor by the anti-IL-1 receptor antibody reduced the LPS-mediated iNOS expression by 50% as quantified by competitive RT-PCR. Furthermore, iNOS activity (nitrite) was also reduced to 50%. However, this magnitude of inhibition was similar in both K and GB2 macrophages. While these observations suggest that IL-1β is involved in mediating LPS-induced iNOS expression and activity, the differential response of GB1 and K-strain macrophages in terms of LPS-induced iNOS expression and activity is unlikely to be modulated by IL-1β. [Copyright &y& Elsevier]

Microglial responses to endotoxin, including the synthesis of inflammatory factors, contribute to gliosis and neuron degeneration in cultured brain tissue. We have previously shown that Go¨6976, a protein kinase C (PKC) inhibitor, suppressed the lipopolysaccharide (LPS)-induced production of inflammatory factors in microglia and afforded marked protection of neurons from glia-mediated cytotoxicity. The purpose of this study was to identify the signal transduction pathway underlying the neuroprotective effect of Go¨6976. Go¨6976 suppressed the LPS-induced release of tumor necrosis factor α (TNFα) in the microglial cell line, BV2. We show in this study the inhibitory effect of Go¨6976 on TNFα release occurring through suppression of p38 mitogen-activated protein kinase (MAPK) phosphorylation and not through a PKC mechanism. While Go¨6976 did not inhibit the activity of p38 MAPK directly, it did suppress its activation by phosphorylation, indicating the target of action of Go¨6976 is a signaling event upstream of p38 MAPK. Although Go¨6976 is considered a selective inhibitor of certain PKC isozymes, suppression of TNFα production was not mediated through inhibition of PKC activity. Go¨6976 appears to play a novel role in neuroprotection by suppressing the release of pro-inflammatory factors by inhibiting the activation of p38 MAPK in microglia, rather than a PKC isoform. [Copyright &y& Elsevier]