Mollugin inhibits IL-1β production by reducing zinc finger protein 91-regulated Pro-IL-1β ubiquitination and inflammasome activity

Background: Rubia cordifolia L. has been formally included in the Chinese Pharmacopoeia and utilized for centuries as a traditional Chinese medicine. Mollugin, a quinone compound, is a major active compound extracted from Rubia cordifolia L. Mollugin was reported has multiple pharmacological activity, including anti-inflammatory, anti-tumor effects. However, the anti-inflammatory mechanism is not yet clear. In this study, we explored the anti-inflammatory activity and potential mechanism of mollugin in vitro and in vivo.

Materials and methods: We explored the mechanisms that mollugin suppressed IL-1β expression through ZFP91 using various assays, including western blot, immunofluorescence, immunoprecipitation, MTT, RT-PCR, and ELISA assays in vitro. In vivo, oral administration of DSS induced colitis in mice and intraperitoneal injection of alum induced peritonitis in mice.

Results: First, the results demonstrated that mollugin dramatically suppressed IL-1β secretion through reducing ZFP91 in macrophages. Crucially, we proved that mollugin inhibited K63-linked Pro-IL-1β ubiquitination through ZFP91 and limitated Pro-IL-1β cleavage efficacy. In addition, ZFP91-mediated Caspase-8 inflammasome component expression was inhibited by mollugin. Furthermore, mollugin inhibited the assembly of the Caspase-8 inflammasome complex by downregulating ZFP91. In vivo studies further revealed that mollugin improved DSS-induced colitis and alum-induced peritonitis in mice by reducing ZFP91. Notely, mollugin significantly altered the abundance of gut flora in DSS-induced colitis mice, which in turn ameliorated the colitis.

Conclusion: We present a novel finding that mollugin inhibition of ZFP91 is a crucial regulatory step, preventing undue inflammatory responses and thereby maintaining immune homeostasis. The current study offers new insight into the development of anti-inflammatory therapeutics targeting ZFP91.