Chylomicron-bound LPS selectively inhibits the TNF-alpha signaling pathway in rat hepatocytes

Abstract

We have previously shown that pretreating primary rat hepatocytes with chylomicron-bound endotoxin (CM-LPS) renders the cells unresponsive to subsequent stimulation by a combination of pro-inflammatory cytokines (TNFα, IL-1β, and IFN-γ), a process we term cytokine tolerance. Thus, we questioned whether the hepatocellular responses elicited by each cytokine were equally inhibited by CM-LPS. Given the prominent role of TNFα as an early circulating mediator of inflammation, we hypothesized that CM-LPS selectively inhibits TNFα signaling rather than inhibiting the effects of all three cytokines. To test this hypothesis, primary rat hepatocytes were pretreated with CM-LPS, recovered, and then stimulated with TNFα, IL-1β, and IFN-γ, individually and in combination. NO production was measured as an indicator of hepatocellular activation and compared to that of non-pretreated, cytokine-stimulated cells. To enhance our ability to study TNFα-induced activation of NF-kB in hepatocytes, McA-RH7777 cells (hepatoma) were stably transfected with a NF-kB-LUC plasmid. Pretreated hepatocytes failed to show any attenuation in response to stimulation with either IL-1β or IFN-γ alone or in combination (data not shown). However, as expected, the cells demonstrated cytokine tolerance when TNFα was added to the cytokine mixture. Stimulation of transfected liver cells showed that pretreatment with CM-LPS significantly inhibited the hepatocellular response to TNFα (Fig. 1) . In summary, CM-LPS complexes inhibit the TNFα signal transduction pathway in hepatocytes. Selective inhibition of this pleiotrophic cytokine leads to a loss of synergy between pro-inflammatory cytokines, and may illustrate a role for lipoproteins in regulating the host innate immune response to infection.