초록/요약

Background: Mer tyrosine kinase (MerTK) activity necessary for amyloid-stimulated phagocytosis strongly implicates that MerTK dysregulation might contribute to chronic inflammation implicated in Alzheimer's disease (AD) pathology. However, the precise mechanism involved in the regulation of MerTK expression by amyloid-beta (A beta) in proinflammatory environment has not yet been ascertained. Methods: The objective of this study was to determine the underlying mechanism involved in A beta-mediated decrease in MerTK expression through A beta-mediated regulation of MerTK expression and its modulation by sulforaphane in human THP-1 macrophages challenged with A beta 1-42. We used protein preparation, Ca2+ influx fluorescence imaging, nuclear fractionation, Western blotting techniques, and small interfering RNA (siRNA) knockdown to perform our study. Results: A beta 1-42 elicited a marked decrease in MerTK expression along with increased intracellular Ca2+ level and induction of proinflammatory cytokines such as IL-1 beta and TNF-alpha. Ionomycin A and thapsigargin also increased intracellular Ca2+ levels and production of IL-1 beta and TNF-alpha, mimicking the effect of A beta 1-42. In contrast, the A beta 1-42-evoked responses were attenuated by depletion of Ca2+ with ethylene glycol tetraacetic acid. Furthermore, recombinant IL-1 beta or TNF-alpha elicited a decrease in MerTK expression. However, immunodepletion of IL-1 beta or TNF-alpha with neutralizing antibodies significantly inhibited A beta 1-42-mediated downregulation of MerTK expression. Notably, sulforaphane treatment potently inhibited A beta 1-42-induced intracellular Ca2+ level and rescued the decrease in MerTK expression by blocking nuclear factor-kappa B (NF-kappa B) nuclear translocation, thereby decreasing IL-1 beta and TNF-alpha production upon A beta 1-42 stimulation. Such adverse effects of sulforaphane were replicated by BAY 11-7082, a NF-kappa B inhibitor. Moreover, sulforaphane's antiinflammatory effects on A beta 1-42-induced production of IL-1 beta and TNF-alpha were significantly diminished by siRNA-mediated knockdown of MerTK, confirming a critical role of MerTK in suppressing A beta 1-42-induced innate immune response. Conclusion: These findings implicate that targeting of MerTK with phytochemical sulforaphane as a mechanism for preventing A beta 1-42-induced neuroinflammation has potential to be applied in AD therapeutics.