Tizoxanide mitigates inflammatory response in LPS-induced neuroinflammation in microglia via restraining p38/MAPK pathway.

OBJECTIVE: Traumatic brain injury (TBI) induced neuroinflammation is featured as excessive glial inflammatory activation and violent neurologic destruction and dysfunction. Massive microglia activation in situ and disrupt of blood-brain barrier contribute to severely collapsed nervous system. Tizoxanide (TIZ), a synthetic thiazolide derivative agent possessing a broad-spectrum anti-infective effect, currently shows a potential resistance against pathogens like bacteria, virus and parasites, while its underlying role in neuroinflammation is elusive. The study aimed to explore the effect of TIZ on neuroinflammation in vitro microglia. MATERIALS AND METHODS: Primary microglia were accepted to neuroinflammatory activation via lipopolysaccharide (LPS) administration. TIZ was conducted to pretreatment of microglia. Cell viability, inflammatory cytokines, chemotaxis, nitric oxide release, inflammation-related enzymes, and mitogen-activated protein kinase (MAPK) pathway activation in microglia were investigated respectively. RESULTS: We demonstrated that TIZ administration attenuates inflammatory cytokines and chemokines through quantitative real-time polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA) of medium supernatant. In addition, TIZ reduces pro-inflammatory mediators and nitric oxide release in microglia. Furtherly, TIZ inhibits the level of p38/MAPK pathway in LPS stimuli, indicating that TIZ negatively regulates neuroinflammation via inhibiting p38/MAPK pathway. CONCLUSIONS: TIZ is verified to be an anti-inflammation effect on neuroinflammation in microglia via downregulation of p38/MAPK pathway, which restrains inflammation by reduced inflammatory cytokines, chemokines and mediators and decreased nitric oxide release. To summarize, TIZ is considered to be a promising reagent to alleviate neuroinflammation targeting microglia in nervous system injury.

20-Hydroxyecdysone-upregulated proteases involved in Bombyx larval fat body destruction.

During insect larval-pupal metamorphosis, the obsolete larval organs and tissues undergo histolysis and programmed cell death to recycle cellular materials. It has been demonstrated that some cathepsins are essential for histolysis in larval tissues, but the process of tissue destruction is not well documented. Fat body, the homologous organ to mammalian liver and adipose tissue, goes through a distinct destruction process during larval-pupal transition. Herein, we found that most of the Bombyx proteases - including Bombyx cathepsin B (BmCatB) (BmCatLL-2), Bombyx cathepsin D (BmCatD), Bombyx cathepsin L like-1 (BmCatLL-1) and -2(BmCatLL-2), Bombyx fibroinase (BmBcp), Bombyx matrix metalloprotease (BmMmp), Bombyx A disintegrin and metalloproteinase with thrombospondin motifs 1 (BmAdamTS-1), Bombyx A disintegrin and metalloproteinase with thrombospondin motifs like (BmAdamTS L) and Bombyx cysteine protease inhibitor (Bmbcpi)- were expressed highly in fat body during feeding and metamorphosis, with a peak occurring during the nonfeeding moulting or prepupal stage, as well as being responsive to 20-hydroxyecdysone (20E). The aforementioned protease genes expression was upregulated by injection of 20E into the feeding larvae, while blocking 20E signalling transduction led to downregulation. Western blotting and immunofluorescent staining of BmCatB and BmBcp confirmed the coincident variation of their messenger RNA (mRNA) and protein level during the development and after the treatments. Moreover, BmCatB, BmBcp, BmMmp and BmAdamTS-1 RNA interference all led to blockage of larval fat body destruction. Taken together, we conclude that 20E regulates larval fat body destruction by upregulating related protease gene expression and protein levels during larval-pupal transition.