Sufu limits sepsis-induced lung inflammation via regulating phase separation of TRAF6.

Rationale: Sepsis is a potentially life-threatening condition caused by the body's response to a severe infection. Although the identification of multiple pathways involved in inflammation, tissue damage and aberrant healing during sepsis, there remain unmet needs for the development of new therapeutic strategies essential to prevent the reoccurrence of infection and organ injuries. Methods: Expression of Suppressor of Fused (Sufu) was evaluated by qRT-PCR, western blotting, and immunofluorescence in murine lung and peritoneal macrophages. The significance of Sufu expression in prognosis was assessed by Kaplan-Meier survival analysis. The GFP-TRAF6-expressing stable cell line (GFP-TRAF6 Blue cells) were constructed to evaluate phase separation of TRAF6. Phase separation of TRAF6 and the roles of Sufu in repressing TRAF6 droplet aggregation were analyzed by co-immunoprecipitation, immunofluorescence, Native-PAGE, FRAP and in vitro assays using purified proteins. The effects of Sufu on sepsis-induced lung inflammation were evaluated by cell function assays, LPS-induced septic shock model and polymicrobial sepsis-CLP mice model. Results: We found that Sufu expression is reduced in early response to lipopolysaccharide (LPS)-induced acute inflammation in murine lung and peritoneal macrophages. Deletion of Sufu aggravated LPS-induced and CLP (cecal ligation puncture)-induced lung injury and lethality in mice, and augmented LPS-induced proinflammatory gene expression in cultured macrophages. In addition, we identified the role of Sufu as a negative regulator of the Toll-Like Receptor (TLR)-triggered inflammatory response. We further demonstrated that Sufu directly interacts with TRAF6, thereby preventing oligomerization and autoubiquitination of TRAF6. Importantly, TRAF6 underwent phase separation during LPS-induced inflammation, which is essential for subsequent ubiquitination activation and NF-κB activity. Sufu inhibits the phase-separated TRAF6 droplet formation, preventing NF-κB activation upon LPS stimulation. In a septic shock model, TRAF6 depletion rescued the augmented inflammatory phenotype in mice with myeloid cell-specific deletion of Sufu. Conclusions: These findings implicated Sufu as an important inhibitor of TRAF6 in sepsis and suggest that therapeutics targeting Sufu-TRAF6 may greatly benefit the treatment of sepsis.

Polydatin Inhibits Cell Viability, Migration, and Invasion Through Suppressing the c-Myc Expression in Human Cervical Cancer.

Polydatin, an active ingredient from the roots of Polygonum cuspidatum, is considered to have protective effects on the cardiovascular system and liver. In this study, we demonstrated that polydatin has antitumor activity against human cervical cancer. Polydatin efficiently inhibited cervical cancer cell proliferation by regulating cell cycle-related proteins including p21, p27, CDK2, CDK4, Cyclin D1, and Cyclin E1. Furthermore, polydatin suppressed cell invasion and migration by regulating epithelial-mesenchymal transition (EMT) markers, including E-cadherin, N-cadherin, Snail and Slug. The c-Myc, as a proto-oncogene, is considered to be closely associated with the proliferation and metastasis of tumor cells. After polydatin treatment, the protein expression of c-Myc showed a significant decrease. Based on these data, we overexpressed c-Myc in cervical cancer cells and observed that the overexpression of c-Myc rescued the inhibitory effect of polydatin on cell proliferation and metastasis. These results indicated that polydatin can inhibit cell proliferation and metastasis through suppressing the c-Myc expression in human cervical cancer.