Transcriptome analysis is shown to be an effective strategy to understand the potential function of natural products. Here, it is reported that 11 previously undescribed hydroanthraquinones [nigroquinones A−K (1−11)], along with eight known congeners, were isolated from Nigrospora sphaerica. Their structures were elucidated by interpreting spectroscopic and spectrometric data including high-resolution mass spectra and nuclear magnetic resonance. The absolute configurations of 1−11 were confirmed by electronic circular dichroism calculations. Transcriptome analysis revealed that 3 (isolated in the largest amount) might be antiinflammatory. Assays based on LPS-induced RAW264.7 macrophages and zebrafish embryos confirmed that some of the isolated hydroanthraquinones attenuated the secretion of pro-inflammatory mediators in vitro and in vivo. Further Western blotting and immunofluorescence experiments indicated that 4 (which showed the most obvious nitric oxide inhibition) could suppress the expression of nuclear factor-kappa-B (NF-κB), phosphorylation of the inhibitor of NF-κB kinase and inhibit the transportation of NF-κB to the nucleus. Hence, the suppression of the NF-κB signaling pathway may be responsible for the anti-inflammatory effect. These results show that bioactivity evaluation on the basis of transcriptome analysis may be effective in the functional exploration of natural products.

Neuroinflammation mediated by microglia is an important pathophysiological mechanism in neurodegenerative diseases. However, there is a lack of effective drugs to treat neuroinflammation. N-acetyldopamine dimer (NADD) is a natural compound from the traditional Chinese medicine Isaria cicada . In our previous study, we found that NADD can attenuate DSS-induced ulcerative colitis by suppressing the NF-κB and MAPK pathways. Does NADD inhibit neuroinflammation, and what is the target of NADD? To answer this question, lipopolysaccharide (LPS)-stimulated BV-2 microglia was used as a cell model to investigate the effect of NADD on neuroinflammation. Nitric oxide (NO) detection, reactive oxygen species (ROS) detection and enzyme-linked immunosorbent assay (ELISA) results show that NADD attenuates inflammatory signals and proinflammatory cytokines in LPS-stimulated BV-2 microglia, including NO, ROS, tumor necrosis factor (TNF)-α, interleukin (IL)-1β and interleukin-6 (IL-6). Western blot analysis show that NADD inhibits the protein levels of Toll-like receptor 4 (TLR4), nuclear factor kappa-B (NF-κB), NOD-like receptor thermal protein domain associated protein 3 (NLRP3), ASC and cysteinyl aspartate specific proteinase (Caspase)-1, indicating that NADD may inhibit neuroinflammation through the TLR4/NF-κB and NLRP3/Caspase-1 signaling pathways. In addition, surface plasmon resonance assays and molecular docking demonstrate that NADD binds with TLR4 directly. Our study reveals a new role of NADD in inhibiting the TLR4/NF-κB and NLRP3/Caspase-1 pathways, and shows that TLR4-MD2 is the direct target of NADD, which may provide a potential therapeutic candidate for the treatment of neuroinflammation.

Excessive inflammation causes chronic diseases and tissue damage. Although there has been drug treatment, its side effects are relatively large. Searching for effective anti-inflammatory drugs from natural products has become the focus of attention. First isolated from Trichoderma longibraciatum, trichodimerol is a natural product with TNF inhibition. In this study, lipopolysaccharide (LPS)-induced RAW264.7 macrophages were used as a model to investigate the anti-inflammatory activity of trichodimerol. The results of nitric oxide (NO) detection, enzyme-linked immunosorbent assay (ELISA), and reactive oxygen species (ROS) showed that trichodimerol could reduce the production of NO, ROS, and the proinflammatory cytokines interleukin (IL)-6 and tumor necrosis factor (TNF)-α. Western blotting results showed that trichodimerol could inhibit the production of inflammatory mediators such as cyclooxygenase (COX)-2 and inducible nitric oxide synthase (iNOS) and the protein expression of nuclear transcription factor-kappaB (NF-κB), p-IKK, p-IκB, Toll-like receptor 4 (TLR4), NOD-like receptor thermal protein domain associated protein 3 (NLRP3), cysteinyl aspartate specific proteinase (Caspase)-1, and ASC, which indicated that trichodimerol may inhibit inflammation through the NF-κB and NLRP3 pathways. At the same time, molecular docking showed that trichodimerol can directly combine with the TLR4-MD2 complex. Hence, trichodimerol inhibits inflammation by obstructing the interaction between LPS and the TLR4-MD2 heterodimer and suppressing the downstream NF-κB and NLRP3 pathways.