FOSL2 deficiency delays nonalcoholic steatohepatitis progression by regulating LY6D-mediated NLRP3 activation.

Lymphocyte antigen 6 family member D (LY6D) was enhanced specifically in senescent cells, while its effects on pyroptosis, a programmed cell death, remains unknown. The goal of this study was to assess the role of LY6D in the mediation of pyroptosis during nonalcoholic steatohepatitis (NASH). After screening out LY6D as a specific liver fibrosis-associated gene using the GSE55747 dataset from the GEO database, we established a NASH mouse model using methionine and choline deficient-diet feeding and an in vitro model using lipopolysaccharide (LPS)-treated hepatocytes. LY6D was overexpressed in NASH livers as well as in LPS-treated hepatocytes. Silencing of LY6D inhibited NASH-associated hepatocyte pyroptosis. With the aid of bioinformatics analysis, promoter-luciferase reporter and ChIP-qPCR assays, we identified FOSL2 as an upstream transcription factor of LY6D. FOSL2, which was highly expressed in NASH, promoted LY6D transcription by binding to the promoter of LY6D. Depletion of FOSL2 significantly inhibited NASH-associated hepatocyte pyroptosis, which was significantly reversed after overexpression of LY6D. Moreover, the promotion of hepatocyte pyroptosis by the FOSL2/LY6D axis was significantly attenuated by specific inhibition of NLRP3. These findings suggesting that FOSL2/LY6D axis may be a key molecular axis and a potential target for NASH therapeutics.

[Combined application of ciprofloxacin and tobramycin on mutant selective windows of ciprofloxacin against Pseudomonas aeruginosa].

OBJECTIVE: To observe whether the mutant selective windows (MSW) of ciprofloxacin would be reduced after its combination against Pseudomonas aeruginosa in rabbits. METHODS: Firstly the minimal inhibitory concentration (MIC), mutant prevention concentration (MPC), mutant selective windows (MSW, MPC-MIC) and selective indices (SI, MPC/MIC) of ciprofloxacin and tobramycin were measured in vitro respectively with standard strain ATCC27853. And the MIC was detected for the combination of ciprofloxacin and tobramycin. The rabbit tissue cage model was constructed to determine the pharmacokinetic parameters of ciprofloxacin by HPLC (high performance liquid chromatography). Fifty-five rabbits were randomly divided by a random number table into 11 groups: physiological saline in 1 group, ciprofloxacin alone in 5 groups and ciprofloxacin plus tobramycin in another 5 groups. The rabbits received ciprofloxacin 10 times a day at a 2-hour dosing interval. In 2 dosing groups, the steady state concentrations of ciprofloxacin reached to 0.25, 0.5, 1.0, 2.0 and 4.0 mg/L respectively. The dose of tobramycin was 2.0 mg×kg(-1)×d(-1) and its peak concentration reached around 2.0 mg/L. At Day 3, the tissue juice was extracted, diluted and coated on agar plates with ciprofloxacin at a concentration of 0.25 mg/L so as to observe the growing condition of mutants. RESULTS: Against Pseudomonas aeruginosa, the values of MIC, MPC and SI of ciprofloxacin were 0.25 mg/L, 4.0 mg/L and 16 while 0.25 mg/L, 8.0 mg/L and 32 for tobramycin respectively. Single groups: the mutants were found in 0.25, 0.5, 1.0 and 2.0 mg/L groups, but none in 4.0 mg/L group. The MPC of ciprofloxacin was the same for in vivo and in vitro. Both were at 16. Combination groups: the mutants were only found in the group with a concentration of ciprofloxacin at 0.25 mg/L while no mutants in the other groups. And MPC was 0.5 mg/L and MIC 0.125 mg/L for ciprofloxacin plus tobramycin. And the value of SI was 4. CONCLUSION: The combined use of ciprofloxacin and tobramycin may reduce the mutant selective windows of ciprofloxacin against P. aeruginosa in rabbits so as to reduce the occurrence of mutants to control its drug resistance.