Caffeine-Stimulated Intestinal Epithelial Cells Suppress Lipid Accumulation in Adipocytes.

Caffeine is a methylxanthine derived from plant foods such as coffee beans and tea leaves, and has multiple biological activities against physiological response and several diseases. Although there are some reports about the direct effect of caffeine against anti-lipid accumulation in vitro, the effect of caffeine on lipid accumulation in adipocytes through stimulating intestinal epithelial cells is unknown. Since direct treatment with caffeine to 3T3-L1 cells did not affect lipid accumulation, we determined whether caffeine-stimulated intestinal epithelial Caco-2 cells influence the lipid accumulation in 3T3-L1 adipocytes. Caco-2 cells were cultured on a transwell insert with or without caffeine for 24 h. Subsequently, the basolateral component of the Caco-2 cell culture on the transwell was collected and termed caffeine-conditioning medium (CCM). When 3T3-L1 adipocytes were incubated with CCM, CCM decreased lipid accumulation and suppressed gene expression of proliferator activated receptor (PPAR) γ and CCAAT/enhancer binding protein (C/EBP) α in 3T3-L1 adipocytes. Furthermore, CCM decreased the expression of C/EBPß and C/EBPδ at the protein level, but not at the mRNA level. We observed that a proteasome inhibitor, MG132, inhibited CCM-caused down-expression of C/EBPß and C/EBPδ proteins, and that CCM promoted the ubiquitination level of C/EBPß and C/EBPδ proteins. Protein microarray analysis showed caffeine suppresses the secretion of inflammatory cytokines, interleukin-8 and plasminogen activator inhibitor-1 from Caco-2 cells. These results suggest that caffeine indirectly suppresses lipid accumulation in 3T3-L1 adipocytes through decreasing secretion of inflammatory cytokines from Caco-2 cells.

Luteolin suppresses TCDD-induced wasting syndrome in a cultured adipocyte model.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) causes various toxic effects, including wasting syndrome, through activation of an aryl hydrocarbon receptor (AhR). Our previous report demonstrated that certain flavonoids inhibit the activation of AhR and suppress its DNA binding activity. In this study, we searched for an active compound among 13 flavonoids that suppressed TCDD-induced loss of lipid accumulation using 3T3-L1 adipocytes as a cell culture model for wasting syndrome. Two flavonoids, luteolin and epigallocatechin gallate, suppressed TCDD-induced loss of lipid accumulation in this model. We further investigated luteolin to clarify the underlying molecular mechanism and confirmed that luteolin inhibited nuclear translocation of AhR caused by TCDD. Luteolin also inhibited the TCDD-driven decrease in protein expression of peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer-binding protein α (C/EBPα). Although TCDD alone did not change protein expression of C/EBPß and C/EBPδ, luteolin and TCDD up-regulated C/EBPδ expression in a dose-dependent manner. On the other hand, TCDD significantly decreased DNA binding of C/EBPß and C/EBPδ, and luteolin completely canceled TCDD-decreased DNA binding of them. We conclude that luteolin suppresses the TCDD-induced loss of lipid accumulation in 3T3-L1 adipocytes by preventing a decrease in protein expression of PPARγ and C/EBPα, the master regulators of adipocyte differentiation and in DNA binding of C/EBPß and C/EBPδ. Moreover, luteolin was rapidly incorporated and accumulated in 3T3-L1 adipocytes. Thus, luteolin is an attractive compound for the prevention of TCDD-induced wasting syndrome.

Luteolin modulates expression of drug-metabolizing enzymes through the AhR and Nrf2 pathways in hepatic cells.

Drugs, xenobiotics including environmental pollutants, and certain food components modulate expression of drug-metabolizing enzymes. An aryl hydrocarbon receptor (AhR) possesses possible expression of phase I and phase II enzymes directly by binding of its ligands such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and indirectly by regulating expression of nuclear factor-erythroid-2-related factor 2 (Nrf2). Previous our result demonstrated that luteolin, a natural flavonoid existing in vegetables and herbs, competed the binding of TCDD to AhR. In the present study, we investigated the effect of luteolin on the expression of drug-metabolizing enzymes through the AhR and Nrf2 pathways. Luteolin inhibited TCDD-induced protein expression of phase I enzyme cytochrome P450 1A1 (CYP1A1), phase II enzymes NAD(P)H: quinone oxidoreductase-1 (NQO1) and glutathione-S-transferase P1 (GSTP1) in HepG2, Hepa1c1c7 and RL-34 cells in a dose-dependent manner. Luteolin suppressed TCDD- and tert-butylhydroquinone-induced Nrf2 protein by decreasing its stability in HepG2 cells. In tert-butylhydroquinone treated cells, luteolin dose-dependently inhibited NQO1, GSTP1 and aldo-keto reductases (AKRs). Of these, protein expression of CYP1A1 and GSTP1 was mainly dominated by the AhR pathway, while that of NQO1 and AKRs was by the Nrf2 pathway. In conclusion, luteolin inhibits expression of phase I and phase II drug-metabolizing enzymes by modulating the AhR and Nrf2 pathways.