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J. Microbiol. Biotechnol. 2019; 29(5): 739-748

Published online May 28, 2019 https://doi.org/10.4014/jmb.1902.02034

Copyright © The Korean Society for Microbiology and Biotechnology.

Secondary Fermented Extract of Chaga-Cheonggukjang Attenuates the Effects of Obesity and Suppresses Inflammatory Response in the Liver and Spleen of High-Fat Diet-Induced Obese Mice

Ha Gyoon Na 1, Yuna Park 2, Min-Ah Kim 1, Jin Woo Lee 2, Gyeongseop So 2, Sung Hyeok Kim 2, Ki-Hyo Jang 1, Mi-Ja Kim 1, Seung Namkoong 3, Hyun Jung Koo 4, Sung Ryul Lee 5 and Eun-Hwa Sohn 2*

1Department of Food & Nutrition, Kangwon National University, Republic of Korea, 2Department of Herbal Medicine Resource, Kangwon National University, Republic of Korea, 3Department of Physical Therapy, Kangwon National University, Republic of Korea, 4Department of Medicinal and Industrial Crops, Korea National College of Agriculture and Fisheries, Republic of Korea, 5Department of Convergence Biomedical Science, Cardiovascular and Metabolic Disease Center, College of Medicine, Inje University, Republic of Korea

Correspondence to:Eun-Hwa  Sohn
ehson@kangwon.ac.kr

Received: February 22, 2019; Accepted: April 26, 2019

Abstract

Cheonggukjang and chaga mushrooms have numerous health benefits, and have been used in alternative medicine. Therefore, a powder mixture of 98: Cheonggukjang and 2: Chaga extracts was fermented with Lactobacillus acidophilus KCTC3925 (FCC) and its anti-obesity effects in high-fat diet (HFD)-induced obese mice were determined. Five-week-old male ICR mice were fed a normal diet or HFD in the presence or absence of 3% and 5% FCC by weight (n = 10 per group). After 12 weeks, the mice were sacrificed, and the serum and tissue samples were collected for analysis. Body weight and epididymal fat pad weight were significantly lowered in the 3% and 5% FCC groups compared with those in the HFD control group (p < 0.01). FCC supplementation suppressed serum triglyceride and increased serum HDL-C levels (p < 0.01). Serum GOT, GPT, and leptin levels, hepatic COX-2 mRNA expression, and splenic COX-2 and IL-4 mRNA expression were significantly higher in the HFD groups than in the control group (p > 0.05); however, except for splenic IL-4 levels, the increases were significantly attenuated by FCC supplementation. Expression of ICAM-1, an aortic inflammatory marker, was significantly increased in the HFD group; this effect was suppressed in the 3% FCC group (p < 0.01) but not in the 5% FCC group. FCC suppressed the body weight and epididymal fat pad weight gain, as well as inflammatory responses in the liver and spleen of HFD-fed mice. Thus, FCC supplementation will be beneficial for the treatment of obesity-related effects.

Keywords: Cheonggukjang, chaga mushroom, Lactobacillus acidophilus KCTC 3925, high-fat diet, anti-obesity, inflammatory molecules

Fig 1.

Figure 1.Effects of FCC on body weight change, rate of food uptake and serum leptin levels. Male ICR mice were fed ND and HFD in the presence or absence of SV, 3% and 5% FCC for 12 weeks (n = 10 per group). Changes in body weight (A) and rate of food uptake (B) were measured weekly. After 12 weeks, serum leptin levels were determined using a commercial assay kit (C). Values represent the mean ± SD. #p < 0.05 and ##p < 0.01 compared with the ND group; *p < 0.05 and **p < 0.01 compared with the HFD group. ND, normal diet; HFD, high-fat diet; FCC; secondary extract of the mixture of Cheonggukjang and Chaga mushroom extract fermented by L. acidophilus KCTC 3925; SV, simvastatin (10 mg/kg body weight).
Journal of Microbiology and Biotechnology 2019; 29: 739-748https://doi.org/10.4014/jmb.1902.02034

Fig 2.

Figure 2.Histology of epididymal adipose tissue and liver. Representative microphotographs of mouse epididymal fat pad (A) and liver (B). Male ICR mice were fed ND and HFD in the presence or absence of SV, 3% and 5% FCC for 12 weeks (n = 10 per group). Fresh epididymal adipose tissue was fixed in 10% formalin, embedded in paraffin, sectioned at a thickness of 5 μm and stained with hematoxylin and eosin (H&E). All images are shown at 200× magnification. ND, normal diet; HFD, high-fat diet; FCC; secondary extract of mixture of Cheonggukjang and Chaga mushroom extract fermented by L. acidophilus KCTC 3925; SV, simvastatin (10 mg/kg body weight).
Journal of Microbiology and Biotechnology 2019; 29: 739-748https://doi.org/10.4014/jmb.1902.02034

Fig 3.

Figure 3.Effect of FCC on the blood lipid profiles, and GOT and GPT levels. After 12 weeks, triglyceride (A), total cholesterol (B), LDL-C (C), HDL-C (D), GOT (E), and GPT (F) levels were determined by enzymatic methods. ##p < 0.01 compared with the ND group; *p < 0.05 or **p < 0.01 compared with the HFD group. Values represent mean ± SD (n = 10 per group). ND, normal diet; HFD, high-fat diet; FCC; secondary extract of the mixture of Cheonggukjang and Chaga mushroom extract fermented by L. acidophilus KCTC 3925; SV, simvastatin (10 mg/kg body weight); LDL-C, low-density lipoprotein-cholesterol; HDL-C, high-density lipoprotein-cholesterol; GOT, aspartate aminotransferase; GPT, alanine aminotransferase.
Journal of Microbiology and Biotechnology 2019; 29: 739-748https://doi.org/10.4014/jmb.1902.02034

Fig 4.

Figure 4.Changes in COX-2 mRNA level in the liver, COX-2 and IL-4 mRNA levels in the spleen, and ICAM-1 protein level in the aorta. After 12 weeks, the liver, spleen, and aorta tissues were removed from the mice. The mRNA expression levels of: hepatic COX-2 (A), splenic COX-2 (B), and splenic IL-4 (C) were quantified by qRT-PCR. Values represent the relative ratio to β-actin as a mean ± SD (n = 10 per group). The protein expression levels of ICAM-1 in the aorta were quantified by immunoblotting. ##p < 0.01 compared with the ND group; **p < 0.01 compared with the HFD group. ND; normal chow diet, HFD; high-fat diet, FCC; secondary extract of the mixture of Cheonggukjang and Chaga extract fermented by L. acidophilus KCTC 3925; SV, simvastatin (10 mg/kg body weight).
Journal of Microbiology and Biotechnology 2019; 29: 739-748https://doi.org/10.4014/jmb.1902.02034

Table 1 . Composition of experimental diets..

Ingredients(g/kg)Experimental diets

NDHFDFCC3FCC5
Casein200200200200
L-Cystine3333
Corn starch315000
Maltodextrin 1035125125125
Sucrose35068.868.868.8
Cellulose, BW 20050505050
Soybean oil25252525
Lard20245245245
Mineral mix S1002610101010
D-calcium phosphate13131313
Calcium carbonate5.55.55.55.5
Potassium citrate16.516.516.516.5
Vitamin mix V1000110101010
Choline bitartrate2222
FD&C yellow dye #50.05---
FD&C blue dye #1-0.050.050.05
FCC3050
Total (g)1055.05773.85803.85823.85
Calories per gram4.0575.2435.2205.203

ND: normal diet, HFD: high-fat diet, FCC; secondary fermented extract of Chaga-cheonggukjang. FCC3; HFD containing 3% FCC (weight/weight); FCC5; HFD containing 5% FCC (weight/weight)..


Table 2 . Changes in body weight and tissue weight of the spleen, liver, and epididymal fat..

Parameters(n = 10 per group)HFD

ND-FCC 3%FCC 5%SV 10mg/kg
Body weight (g)
Baseline32.8 ± 0.5331.1 ± 0.2833.2 ± 1.2931.7 ± 1.6832.5 ± 1.90
Final44.2 ± 3.1964.4 ± 4.44##45.4 ± 5.41**44.4 ± 3.24**55.2 ± 3.82**
Weight gain (g)11.42 ± 2.8533.23 ± 4.05##12.20 ± 4.82**12.72 ± 2.98**22.66 ± 2.27**
Tissue weight (g/kg body weight)
Spleen3.45 ± 0.092.68 ± 0.08#2.61 ± 0.133.04 ± 0.09*2.52 ± 0.16
Liver36.20 ± 0.5534.70 ± 3.6131.38 ± 3.1133.30 ± 3.8830.76 ± 3.10
Epididymal fat pad10.72 ± 1.2743.02 ± 1.34##37.10 ± 1.79*11.25 ± 0.63**49.43±0.78

ND; normal chow diet, HFD; high fat diet, FCC; fermented mixture of Chaga and Cheonggukjang with L. acidophilus KCTC 3925. SV; simvastatin (10 mg/kg body weight) treatment on HFD mice. # and ## indicate p < 0.05 and p < 0.01 compared with the ND group, respectively; * and ** indicate p < 0.05 and p < 0.01compared with the HFD group, respectively..