The anti-obesity effect of Lethariella cladonioides in 3T3-L1 cells and obese mice

Abstract

The aim of this study was to investigate whether a water extract of L. cladonioides (LC) has an anti-obesity effect in 3T3-L1 cells and obese mice. Treatment of differentiated 3T3-L1 adipocytes with LC caused a significant increase in glycerol release and reduced the protein expression of the adipogenic transcription factors, PPARγ and C/EBPα. In an animal model, obese mice were artificially induced by a high fat diet for 10 weeks. Experimental groups were treated with LC (100 mg/kg/day) by gavage for the next 10 weeks. At the end of experiment, the body weight of the LC group mice was reduced by 14.2% compared to the high fat diet (HFD) group. The treatment also decreased liver (31.0%), epididymal (18.0%) and retroperitoneal (19.3%) adipose tissue, and kidney (6.7%) weights, respectively, compared with those of the HFD group. LC prevented diet-induced increases in the serum level of TC (22.6%), TG (11.6%), and glucose (35.0%), respectively, compared with the HFD group. However, the HDL-C level was higher in the LC group (26.1%) than the HFD group. The results of this study thus suggest that LC suppressed lipid accumulation and expression of adipogenic transcription factors, and increased the amount of glycerol release. LC also indicated an anti-obese and anti-hyperlipidemic effect.

Keywords: 3T3-L1 cells; Lethariella cladonioides; obese mice; obesity.

Fig. 1

The effect of L. cladonioides on cell viability in 3T3-L1 cells. Differentiated 3T3-L1 cells were incubated with L. cladonioides at the indicated various concentrations for 24, 48, and 72 hr. Data are expressed as percent growth rate of cells cultured in the presence of L. cladonioides, compared with untreated control cells, taken as 100% (mean ± SD). Statistical analysis was performed using a one-way ANOVA with repeated measures followed by Duncan's multiple range tests. Letters with different superscripts are significantly different (P < 0.05) among groups by one-way ANOVA.

Fig. 2

The effect of L. cladonioides on apoptosis/necrosis in 3T3-L1 cells. Differentiated 3T3-L1 cells were incubated with L. cladonioides at the indicated various concentrations for 72 hr by Annexin V/PI double staining. Annexin V FITC^-/PI^- (Normal cells), Annexin V FITC⁺/PI^- (Early apoptotic cells), Annexin V FITC⁺/PI⁺ (Late apoptotic cells), and Annexin V FITC^-/PI⁺ (Necrotic cells). Statistical analysis was performed using one-way ANOVA with repeated measures followed by Duncan's multiple range tests. Letters with different superscripts are significantly different (P < 0.05) among groups by one-way ANOVA.

Fig. 3

Effect of L. cladonioides on lipid accumulation in 3T3-L1 cells. Differentiated 3T3-L1 cells were treated with various concentrations of L. cladonioides. As a positive control, TNF-α (1 ng/mL) was used. After 72 hr, lipid accumulation was evaluated using Oil Red O staining. Each experiment was performed in triplicate. Values are means ± SD. Statistical analysis was performed using one-way ANOVA with repeated measures followed by Duncan's multiple range tests. Letters with different superscripts are significantly different (P < 0.05) among groups by one-way ANOVA.

Fig. 4

The effect of L. cladonioides on glycerol release in 3T3-L1 cells. Differentiated 3T3-L1 cells were treated with various concentrations of L. cladonioides. Cell supernatants were collected after 24, 48, and 72 hr and free glycerol release was assayed. Each experiment was performed in triplicate. Values are mean ± SD. Statistical analysis was performed using a one-way ANOVA with repeated measures followed by Duncan's multiple range tests. Letters with different superscripts are significantly different (P < 0.05) among groups by one-way ANOVA.

Fig. 5

The effect of L. cladonioides on the expression of the adipogenic enzyme in 3T3-L1. Differentiated 3T3-L1 cells were treated with various concentrations of L. cladonioides. After 72 hr, cells were collected and they were washed in PBS and lysed in RIPA buffer. Each experiment was performed in triplicate. Values are mean ± SD. Statistical analysis was performed using one-way ANOVA with repeated measures followed by Duncan's multiple range tests. Letters with different superscripts are significantly different (P < 0.05) among groups by one-way ANOVA.

Fig. 6

Comparisons of body weight in DIO mice treated with L. cladonioides. ND, Normal diet; HFD, high fat diet; CA, caffeine; LC, L. cladonioides. Values for the normal diet group were measured, but excluded from statistics. Data were presented as the mean ± SD. Statistical analysis was performed using a one-way ANOVA with repeated measures followed by Duncan's multiple range tests. Letters with different superscripts are significantly different (P < 0.05) among groups by one-way ANOVA.