2023 Impact Factor
Hepatic insulin resistance has been shown to be strongly associated with excess lipid accumulation in the liver and non-alcoholic fatty liver disease (NAFLD) (Loomba
Cluster of differentiation 38 (CD38), known as cyclic ADP ribose hydrolase (Morandi
Various studies have evaluated the preventive efficacy of safe food-derived bioactive compounds (Kessoku
AML-12 hepatocytes originating from mice were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA). The cells were cultivated at 37°C in a humidified atmosphere containing 5% CO2. AML-12 cells were cultured in Dulbecco’s modified Eagle’s medium/F12 medium (1:1) with 10% fetal bovine serum, antibiotics (Welgene, Daegu, Korea), 40 ng/mL dexamethasone (Sigma-Aldrich, St. Louis, MO, USA), and Insulin-Transferrin-Selenium-Sodium Pyruvate (ITS) solution (Gibco, Waltham, MA, USA). To establish a model of nonalcoholic fatty liver disease, AML-12 cells were treated with a non-fat BSA-conjugated combination of oleic acid and palmitic acid (OPA, a mixture of 800 μM oleic acid and 150 μM palmitic acid) (Sigma-Aldrich) in the presence or absence of RBN (ChemFaces, Wuhan, China) for 18 h.
Five-week-old male C57/BL6 mice were purchased from Nara Biotech (Seoul, Korea) and housed under standardized conditions and a 12-h light/dark cycle. Animal experiments were conducted in accordance with the Institutional Ethics and Animal Committee of the Korea Food Research Institute (KFRI-M-19005). After 1 week of acclimation, the mice were fed a Western diet (WD, Research Diet Inc., New Brunswick, NJ, USA) containing 0.025% RBN or 0.5% RBN (w/w) for developing a diet-induced obesity (DIO) model. The control group was fed a control diet (CD; Research Diet Inc.). Forty male C57/BL6 mice were randomly divided into four groups as follows: 1) CD group (n=10), 2) WD group (n=10), 3) WD with 0.025% RBN group, and 4) WD with 0.05% RBN group (Supplementary Table 3). Body weight and food intake of the mice were measured weekly. After 12 weeks, the mice were sacrificed following isoflurane exposure under anesthesia, and blood samples were obtained for serum isolation. The liver tissues were immediately harvested and weighed.
The structure of p300 (PDB ID: 3BIY) (Liu
Data were analyzed using either Student’s
In an ongoing screening study for identifying phytochemicals that exhibit an anti-lipogenic effect, we found that RBN effectively inhibited TG accumulation in mouse hepatocytes, AML-12 cells. Furthermore, the effect was similar to that of EGCG, apigenin, and piceatannol, which are already known to have anti-lipogenic effects (Supplementary Fig. 1). For confirming this result, AML-12 cells were treated with oleic and palmitic acid (OPA) and different doses of RBN. RBN blocked OPA-induced TG accumulation in AML-12 cells without causing cytotoxicity (Fig. 1A, 1B). The mRNA expression levels of peroxisome proliferator-activated receptor gamma 2 (
The evidence has been accumulated to show relevance between histone acetyltransferases (HAT) and NAFLD (Chung
To identify the mechanism underlying RBN action in hepatocytes, we identified the protein targets of RBN using the publicly available BioAssay database (Wang
Generally, p300-mediated histone acetylation in the promoter region increases the transcriptional activity of genes (Tolsma and Hansen, 2019). To evaluate the involvement of p300 in CD38 regulation, we first determined its conserved promoter sequence. Comparing the sequence motif with a prediction based on a position weight matrix model, we determined a plausible region for the transcription of CD38 containing the TATA box and binding for a transcription factor, E2A (Fig. 4A). To elucidate whether RBN attenuated
In the present study, our data clearly showed that RBN prevented TG accumulation without causing cytotoxicity in AML-12 cells. NAFLD is characterized by fat accumulation in the liver. It is also used to describe a range of states of hepatic TG accumulation (Moon, 2017). Thus, the inhibition of OPA-induced TG by RBN reflected the attenuation of a significant NAFLD pathological feature. In AML-12 cells, this phenomenon appeared to be due to the obstructive effect of RBN on mRNA expression of key transcription factors such as
Funally, to determine whether the anti-p300 activity of RBN negatively regulated
In the present study, we first demonstrated that RBN is a potent p300 inhibitor. Furthermore, our findings indicate the possibility that, through its p300 inhibitory activity, RBN can prevent or improve the significant characteristics of metabolic failure in the liver caused by CD38 induction (Fig. 5). Although this study revealed the beneficial effects of RBN, it failed to demonstrate the dose-dependent effects of RBN
The authors declare that they have no conflicts of interest.
This work was supported by the Main Research Program (E-0210400 and E-0210601) of the Korea Food Research Institute (KFRI) and funded by the Ministry of Science, ICT & Future Planning. Part of this paper was presented as a poster at FEBS2023.
Conceptualization: H.-K.C. and J.-T.H.; Data curation: J.-T. H.; Funding acquisition: H.-K.C. and J.-T.H.; Investigation: J.-H.S., H.-J.K., and J.L.; Methodology: S.-P.H., J.L., J.-H.S., M.-Y.C., and Y.-G.L.; Project administration: H.-K.C. and J.-T.H.; Supervision: H.-K.C. and J.-H.P.; Writing – original draft: H.-K.C.; Validation: H.-K.C and J.-H.S.; Writing – review & editing: H.-K.C. and J.-H.P.