METHODS: The ATGL-predicted protein structure, verified by PROCHECK, was determined using AlphaFold. Molecular docking, molecular dynamics simulation, and prime molecular mechanic-generalized born surface area were performed using LigPrep, Desmond, and prime MM-GBSA modules of Schrödinger software release 2021-2, respectively. For pharmacological validation, immunoblotting was performed to assess ATGL protein expression. The fluorescence intensity and glycerol concentration were quantified to evaluate the efficiency of phillyrin in inhibiting ATGL.
OBJECTIVE: We previously identified AtWRKY50 as a transcriptional activator of SAR gene PR1. Although PR1 accumulates to high levels in plants after attack by pathogens, its function is still elusive. Here we investigated the effects of overexpression of several WRKY proteins, including AtWRKY50, on the metabolome of Arabidopsis thaliana.
METHODS: The influence of overexpression of WRKY proteins on the metabolites of Arabidopsis was investigated by using an NMR spectroscopy-based metabolomic approach. The 1H NMR data was analysed using the multivariate data analysis methods, such as principal component analysis, hierarchical cluster analysis and partial least square-discriminant analysis.
RESULTS: The results showed that the metabolome of transgenic Arabidopsis seedlings overexpressing AtWRKY50 was different from wild type Arabidopsis and transgenic Arabidopsis overexpressing other WRKY genes. Amongst other metabolites, sinapic acid and 1-O-sinapoyl-β-D-glucose especially appeared to be the most prominent discriminating metabolites, accumulating to levels 2 to 3 times higher in the AtWRKY50 overexpressor lines.
CONCLUSION: Our results indicate a possible involvement of AtWRKY50 in secondary metabolite production in Arabidopsis, in particular of hydroxycinnamates such as sinapic acid and 1-O-sinapoyl-β-D-glucose.
BACKGROUND: CKD is associated with fluid excess that can be estimated by bioimpedance spectroscopy. We aimed to assess effects of sodium glucose co-transporter 2 inhibition on bioimpedance-derived "Fluid Overload" and adiposity in a CKD population.
METHODS: EMPA-KIDNEY was a double-blind placebo-controlled trial of empagliflozin 10 mg once daily in patients with CKD at risk of progression. In a substudy, bioimpedance measurements were added to the main trial procedures at randomization and at 2- and 18-month follow-up visits. The substudy's primary outcome was the study-average difference in absolute "Fluid Overload" (an estimate of excess extracellular water) analyzed using a mixed model repeated measures approach.
RESULTS: The 660 substudy participants were broadly representative of the 6609-participant trial population. Substudy mean baseline absolute "Fluid Overload" was 0.4±1.7 L. Compared with placebo, the overall mean absolute "Fluid Overload" difference among those allocated empagliflozin was -0.24 L (95% confidence interval [CI], -0.38 to -0.11), with similar sized differences at 2 and 18 months, and in prespecified subgroups. Total body water differences comprised between-group differences in extracellular water of -0.49 L (95% CI, -0.69 to -0.30, including the -0.24 L "Fluid Overload" difference) and a -0.30 L (95% CI, -0.57 to -0.03) difference in intracellular water. There was no significant effect of empagliflozin on bioimpedance-derived adipose tissue mass (-0.28 kg [95% CI, -1.41 to 0.85]). The between-group difference in weight was -0.7 kg (95% CI, -1.3 to -0.1).
CONCLUSIONS: In a broad range of patients with CKD, empagliflozin resulted in a sustained reduction in a bioimpedance-derived estimate of fluid overload, with no statistically significant effect on fat mass.
TRIAL REGISTRATION: Clinicaltrials.gov: NCT03594110 ; EuDRACT: 2017-002971-24 ( https://eudract.ema.europa.eu/ ).