Improvement Effect of Bifidobacterium lactis V9 on NAFLD Rats Induced by High-fat Diet

doi:10.13560/j.cnki.biotech.bull.1985.2021-0630

Abstract

Abstract:

This work aims to study the effect of probiotic Bifidobacterium lactis V9（V9）on lipid metabolism and intestinal injury in non-alcoholic fatty liver disease（NAFLD）rats induced by high-fat diet,and to explore its mechanism. Male Wistar rats were randomly divided into 5 groups（n=8）：blank control group（Control）,high-fat diet model group（HFD）,berberine-positive drug group（Berberine）,the probiotic B. lactis V9 treatment group（HFD/V9）,and the probiotic B. lactis V9 control group（V9）. Except Control and V9 were given ordinary maintenance feed,others were given a high-fat diet for 6 weeks to construct a NAFLD model. After 6 weeks,the high-fat diet was replaced with ordinary maintenance feed,and the rats were given probiotic B. lactis V9（1×10 ⁹CFU/mL）or berberine（50 mg/kg）by gavage. After 4 weeks,blood,liver and colon tissues were collected for follow-up testing. Compared with the Control group,the mRNA expressions of liver non-esterified fatty acids（NEFA）,triglycerides（TG）,liver fatty acid synthase（FAS）and liver fatty acid binding protein 1（Fabp1）in the HFD group significantly increased（P<0.01）. At the same time,mRNA expressions of the intestinal cytokine interleukin 1β（IL-1β）,tumor necrosis factor α（TNF-α）,Toll-like receptor 2（TLR2）and Toll-like receptor 9（TLR9）in the HFD model group also significantly increased（P<0.01）. On the contrary,the expression of tight junction protein ZO-1 and occludin mRNA in the HFD model group significantly decreased（P<0.01）. After treatment with probiotic B. lactis V9,the contents of NEFA and TG in the liver significantly reduced（P<0.01）,the mRNA expressions of FAS and Fabp1 in the liver also significantly reduced（P<0.01）. Both probiotic B. lactis V9 and berberine treatment reduced the transcription levels of intestinal IL-1β,TNF-α,TLR2 and TLR9（P<0.01）,which increased the expressions of intestinal tight junction proteins ZO-1 and occludin mRNA. HE staining showed that there was no significant difference in colonic pathology among the above groups. Probiotic Probiotics B. lactis V9 can improve NAFLD induced by high-fat diet by alleviating lipid metabolism disorders,reducing intestinal inflammation and improving mulosal barrier.

Key words: intestinal microflora, non-alcoholic fatty liver disease, probiotics, high fat diet

ZHONG Ming-yue, LIU Chun-yan, YAN Yan, ZHANG Xiao-hui, YUAN Hai-sheng, XU Guo-quan, ZHANG He-ping, WANG Yu-zhen. Improvement Effect of Bifidobacterium lactis V9 on NAFLD Rats Induced by High-fat Diet[J]. Biotechnology Bulletin, 2022, 38(3): 181-187.

Figures/Tables 7

References 22

[1]	Liu JY, Ayada I, Zhang XF, et al. Estimating global prevalence of metabolic dysfunction-associated fatty liver disease in overweight or obese adults[J]. Clin Gastroenterol Hepatol, 2021. DOI: 10.1016/j.cgh,2021.02.030. doi: 10.1016/j.cgh,2021.02.030
[2]	Diehl AM, Day C. Cause, pathogenesis, and treatment of nonalcoholic steatohepatitis[J]. N Engl J Med, 2017, 377(21):2063-2072. doi: 10.1056/NEJMra1503519 URL
[3]	M I, Singh C, Ganie MA, et al. NASH:The hepatic injury of metabolic syndrome:a brief update[J]. Int J Health Sci:Qassim, 2009, 3(2):265-270.
[4]	İlyas TUNCER, Hanefi ÖZBEK, Topal C, et al. The serum levels of IL-1b, IL-6, IL-8 and TNF-a in nonalcoholic fatty liver[J]. Turkish Journal of Medical Sciences, 2003, 33(24):381-386.
[5]	Day CP, James OF. Steatohepatitis:a tale of two “hits” ?[J]. Gastroenterology, 1998, 114(4):842-845. pmid: 9547102
[6]	Buzzetti E, Pinzani M, Tsochatzis EA. The multiple-hit pathogenesis of non-alcoholic fatty liver disease(NAFLD)[J]. Metabolism, 2016, 65(8):1038-1048. doi: 10.1016/j.metabol.2015.12.012 URL
[7]	Fang YL, Chen H, Wang CL, et al. Pathogenesis of non-alcoholic fatty liver disease in children and adolescence:From “two hit theory” to “multiple hit model”[J]. World J Gastroenterol, 2018, 24(27):2974-2983. doi: 10.3748/wjg.v24.i27.2974 URL
[8]	Tilg H, Moschen AR. Evolution of inflammation in nonalcoholic fatty liver disease:The multiple parallel hits hypojournal[J]. Hepatology, 2010, 52(5):1836-1846. doi: 10.1002/hep.v52:5 URL
[9]	Rinaldi L, Pafundi PC, Galiero R, et al. Mechanisms of non-alcoholic fatty liver disease in the metabolic syndrome. A narrative review[J]. Antioxidants, 2021, 10(2):270. doi: 10.3390/antiox10020270 URL
[10]	Li ZP, Yang SQ, Lin HZ, et al. Probiotics and antibodies to TNF inhibit inflammatory activity and improve nonalcoholic fatty liver disease[J]. Hepatology, 2003, 37(2):343-350. doi: 10.1053/jhep.2003.50048 URL
[11]	Sun ZH, Chen X, Wang JC, et al. Complete genome sequence of probiotic Bifidobacterium animalis subsp. lactis strain V9[J]. J Bacteriol, 2010, 192(15):4080-4081. doi: 10.1128/JB.00369-10 URL
[12]	陈霞. 具有益生功能的Bifidobacterium animalis subsp. lactis V9的安全性评估、生理功效及其全基因组学研究[D]. 呼和浩特:内蒙古农业大学, 2010.
	Chen X. Safety evaluation, physiological function & whole genome sequence of potential probiotic Bifidobacterium animalis subsp. lactis V9[D]. Hohhot:Inner Mongolia Agricultural University, 2010.
[13]	Wong WK, Chan WK. Nonalcoholic fatty liver disease:a global perspective[J]. Clin Ther, 2021, 43(3):473-499. doi: 10.1016/j.clinthera.2021.01.007 URL
[14]	Dongiovanni P, Paolini E, Corsini A, et al. NAFLD or MAFLD diagnoses and cardiovascular diseases:from epidemiology to drug approaches[J]. European Journal of Clinical Investigation, 2021, 14(5):e13519.
[15]	Cai GS, Su H, Zhang J. Protective effect of probiotics in patients with non-alcoholic fatty liver disease[J]. Medicine, 2020, 99(32):e21464. doi: 10.1097/MD.0000000000021464 URL
[16]	Kwong EK, Puri P. Gut microbiome changes in nonalcoholic fatty liver disease & alcoholic liver disease[J]. Transl Gastroenterol Hepatol, 2021, 6:3. doi: 10.21037/tgh.2020.02.18 pmid: 33409398
[17]	Bajaj JS, Heuman DM, Hylemon PB, et al. Randomised clinical trial:Lactobacillus GG modulates gut microbiome, metabolome and endotoxemia in patients with cirrhosis[J]. Aliment Pharmacol Ther, 2014, 39(10):1113-1125. doi: 10.1111/apt.2014.39.issue-10 URL
[18]	Yan Y, Liu C, Zhao S, et al. Probiotic Bifidobacterium lactis V9 attenuates hepatic steatosis and inflammation in rats with non-alcoholic fatty liver disease[J]. AMB Express, 2020, 10(1):101. doi: 10.1186/s13568-020-01038-y pmid: 32472368
[19]	Kumar R, Goh BG, Kam JW, et al. Comparisons between non-alcoholic steatohepatitis and alcohol-related hepatocellular carcinoma[J]. Clin Mol Hepatol, 2020, 26(2):196-208. doi: 10.3350/cmh.2019.0012 URL
[20]	Zhou HP, Urso CJ, Jadeja V. Saturated fatty acids in obesity-associated inflammation[J]. J Inflamm Res, 2020, 13:1-14. doi: 10.2147/JIR URL
[21]	蒋李妍, 肖新华. 肠道菌群与非酒精性脂肪肝病相关性研究进展[J]. 临床与病理杂志, 2016, 36(12):2060-2065.
	Jiang LY, Xiao XH. Research progress on correlation between gut microbiota and non-alcoholic fatty liver disease[J]. J Clin Pathol Res, 2016, 36(12):2060-2065.
[22]	Dai X, Wang BM. Role of gut barrier function in the pathogenesis of nonalcoholic fatty liver disease[J]. Gastroenterol Res Pract, 2015, 2015:1-6.

	脂肪 Fat/%	蛋白质 Protein/%	碳水化合物 Carbohydrate/%
普通维持饲料 Ordinaryl feed	11.4	27.5	65.8
高脂饲料High-fat diet （HFD）	33.1	19.6	47.1

	脂肪 Fat/%	蛋白质 Protein/%	碳水化合物 Carbohydrate/%
普通维持饲料 Ordinaryl feed	11.4	27.5	65.8
高脂饲料High-fat diet （HFD）	33.1	19.6	47.1

引物名称 Primer name	引物序列 Primer sequence
IL-1β antisense	5'-GAATGCCACCTTTTGACAGTG-3'
IL-1β sense	5'-TGGATGCTCTCATCAGGACAG-3'
TNF-α antisense	5'-TTGATGGTGGTGCATGAGAG-3'
TNF-α sense	5'-AAACACAAGATGCTGGGACA-3'
TLR2 antisense	5'-CTCCTGTGAACTCCTGTCCTT-3'
TLR2 sense	5'AGCTGTCTGGCCAGTCAAC-3'
TLR9 antisense	5'-CCGAAGACCTAGCCAACCT-3'
TLR9 sense	5'-TGATCACAGCGACGGCAATT-3'
Fas antisense	5'-GAGCGTTCGTGAAACCGACA-3'
Fas sense	5'-AGGTTGGTGCACCTCCACTTG-3'
Fabp1 antisense	5'-TTCCCCAGTCATGGTCTCCA-3'
Fabp1 sense	5'-TCATGAAGGCGATGGGTCTG-3'
ZO-1 antisense	5'-CCATCTTTGGACCGATTGCTG-3'
ZO-1 sense	5'-TAATGCCCGAGCTCCGATG-3'
Occludin antisense	5'-TTGGGAGCCTTGACATCTTGTTC-3'
Occludin sense	5'-GCCATACATGTCATTGCTTGGTG-3'
GAPDH antisense	5'-AGGTCGGTGTGTGAACGGATTTG-3'
GAPDH sense	5'-TGTAGACCATGTAGTTGAGGTCA-3'

引物名称 Primer name	引物序列 Primer sequence
IL-1β antisense	5'-GAATGCCACCTTTTGACAGTG-3'
IL-1β sense	5'-TGGATGCTCTCATCAGGACAG-3'
TNF-α antisense	5'-TTGATGGTGGTGCATGAGAG-3'
TNF-α sense	5'-AAACACAAGATGCTGGGACA-3'
TLR2 antisense	5'-CTCCTGTGAACTCCTGTCCTT-3'
TLR2 sense	5'AGCTGTCTGGCCAGTCAAC-3'
TLR9 antisense	5'-CCGAAGACCTAGCCAACCT-3'
TLR9 sense	5'-TGATCACAGCGACGGCAATT-3'
Fas antisense	5'-GAGCGTTCGTGAAACCGACA-3'
Fas sense	5'-AGGTTGGTGCACCTCCACTTG-3'
Fabp1 antisense	5'-TTCCCCAGTCATGGTCTCCA-3'
Fabp1 sense	5'-TCATGAAGGCGATGGGTCTG-3'
ZO-1 antisense	5'-CCATCTTTGGACCGATTGCTG-3'
ZO-1 sense	5'-TAATGCCCGAGCTCCGATG-3'
Occludin antisense	5'-TTGGGAGCCTTGACATCTTGTTC-3'
Occludin sense	5'-GCCATACATGTCATTGCTTGGTG-3'
GAPDH antisense	5'-AGGTCGGTGTGTGAACGGATTTG-3'
GAPDH sense	5'-TGTAGACCATGTAGTTGAGGTCA-3'