Single and Mixed Strains of Probiotics Reduced Hepatic Fat Accumulation and Inflammation and Altered Gut Microbiome in a Nonalcoholic Steatohepatitis Rat Model
Abstract
:1. Introduction
2. Materials and Methods
2.1. Animals and Experimental Design
2.2. Probiotics Preparation
2.3. Liver Histopathological Evaluation
2.4. Hepatic Lipid Accumulation Evaluation
2.5. Immunohistochemistry
2.6. Hepatic Free Fatty Acid Measurement
2.7. Serum TNF-α and IL-6 Measurement
2.8. Fecal Microbiota
2.9. Statistical Analysis
3. Results
3.1. Effects of Single- and Mixed-Strain Probiotics on Liver Histology
3.2. Effects of Single- and Mixed-Strain Probiotics on Hepatic Lipid Accumulation and Hepatic Free Fatty Acid Levels
3.3. Effects of Single- and Mixed-Strain Probiotics on Serum TNF-α and IL-6 Levels
3.4. Effects of Single- and Mixed-Strain Probiotics on CD14 and TLR4 Expressions
3.5. Effects of Single- and Mixed-Strain Probiotics on Gut Microbiota Change
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
References
- Buzzetti, E.; Pinzani, M.; Tsochatzis, E.A. The multiple-hit pathogenesis of non-alcoholic fatty liver disease (NAFLD). Metab. Clin. Exp. 2016, 65, 1038–1048. [Google Scholar] [CrossRef] [PubMed]
- Kirpich, I.A.; Marsano, L.S.; McClain, C.J. Gut-liver axis, nutrition, and non-alcoholic fatty liver disease. Clin. Biochem. 2015, 48, 923–930. [Google Scholar] [CrossRef]
- Do, M.H.; Lee, E.; Oh, M.-J.; Kim, Y.; Park, H.-Y. High-Glucose or -Fructose Diet Cause Changes of the Gut Microbiota and Metabolic Disorders in Mice without Body Weight Change. Nutrients 2018, 10, 761. [Google Scholar] [CrossRef] [PubMed]
- Ilan, Y. Leaky gut and the liver: A role for bacterial translocation in nonalcoholic steatohepatitis. World J. Gastroenterol. 2012, 18, 2609–2618. [Google Scholar] [CrossRef] [PubMed]
- Sharifnia, T.; Antoun, J.; Verriere, T.G.; Suarez, G.; Wattacheril, J.; Wilson, K.T.; Peek, R.M., Jr.; Abumrad, N.N.; Flynn, C.R. Hepatic TLR4 signaling in obese NAFLD. Am. J. Physiol. Gastrointest Liver Physiol. 2015, 309, G270–G278. [Google Scholar] [CrossRef] [PubMed]
- Ferreira, D.F.; Fiamoncini, J.; Prist, I.H.; Ariga, S.K.; de Souza, H.P.; de Lima, T.M. Novel role of TLR4 in NAFLD development: Modulation of metabolic enzymes expression. Biochim. Biophys. Acta. Mol. Cell Biol. Lipids. 2015, 1851, 1353–1359. [Google Scholar] [CrossRef] [PubMed]
- Arroyo-Espliguero, R.; Avanzas, P.; Jeffery, S.; Kaski, J.C. CD14 and toll-like receptor 4: A link between infection and acute coronary events? Heart 2004, 90, 983–988. [Google Scholar] [CrossRef] [PubMed]
- Zanoni, I.; Ostuni, R.; Marek, L.R.; Barresi, S.; Barbalat, R.; Barton, G.M.; Granucci, F.; Kagan, J.C. CD14 controls the LPS-induced endocytosis of Toll-like receptor 4. Cell 2011, 147, 868–880. [Google Scholar] [PubMed]
- Brun, P.; Castagliuolo, I.; Leo, V.D.; Buda, A.; Pinzani, M.; Palù, G.; Martines, D. Increased intestinal permeability in obese mice: New evidence in the pathogenesis of nonalcoholic steatohepatitis. Am. J. Physiol. Gastrointest Liver Physiol. 2007, 292, G518–G525. [Google Scholar] [CrossRef]
- Nakamura, A.; Yamamoto, K.; Takeda, R.; Yamada, R.; Kubo, A.; Morikawa, K.; Ando, S.; Shimazaki, T.; Izumi, T.; Umemura, M.; et al. The potential of soluble CD14 in discriminating nonalcoholic steatohepatitis from nonalcoholic fatty liver disease. Hepatol. Res. 2022, 52, 508–521. [Google Scholar] [CrossRef]
- Sabirin, F.; Lim, S.M.; Neoh, C.F.; Ramasamy, K. Hepatoprotection of Probiotics Against Non-Alcoholic Fatty Liver Disease in vivo: A Systematic Review. Front. Nutr. 2022, 9, 884374. [Google Scholar] [CrossRef]
- Zhou, X.; Wang, J.; Zhou, S.; Liao, J.; Ye, Z.; Mao, L. Efficacy of probiotics on nonalcoholic fatty liver disease: A meta-analysis. Medicine 2023, 102, e32734. [Google Scholar] [CrossRef] [PubMed]
- Mohammed, S.K.; Magdy, Y.M.; El-Waseef, D.A.; Nabih, E.S.; Hamouda, M.A.; El-Kharashi, O.A. Modulation of hippocampal TLR4/BDNF signal pathway using probiotics is a step closer towards treating cognitive impairment in NASH model. Physiol. Behav. 2020, 214, 112762. [Google Scholar] [CrossRef] [PubMed]
- Park, E.J.; Lee, Y.S.; Kim, S.M.; Park, G.S.; Lee, Y.H.; Jeong, D.Y.; Kang, J.; Lee, H.J. Beneficial Effects of Lactobacillus plantarum Strains on Non-Alcoholic Fatty Liver Disease in High Fat/High Fructose Diet-Fed Rats. Nutrients 2020, 12, 542. [Google Scholar] [CrossRef] [PubMed]
- Werawatganon, D.S.K.; Tumwasorn, S.; Klaikeaw, N.; Siriviriyakul, P. Lactobacillus plantarum attenuates oxidative stress and liver injury in rats with nonalcoholic steatohepatitis. Pharmacogn. Mag. 2018, 14, 471–476. [Google Scholar] [CrossRef]
- Chayanupatkul, M.; Somanawat, K.; Chuaypen, N.; Klaikeaw, N.; Wanpiyarat, N.; Siriviriyakul, P.; Tumwasorn, S.; Werawatganon, D. Probiotics and their beneficial effects on alcohol-induced liver injury in a rat model: The role of fecal microbiota. BMC Complement Med. Ther. 2022, 22, 168. [Google Scholar] [CrossRef] [PubMed]
- Boonma, P.; Spinler, J.K.; Venable, S.F.; Versalovic, J.; Tumwasorn, S. Lactobacillus rhamnosus L34 and Lactobacillus casei L39 suppress Clostridium difficile-induced IL-8 production by colonic epithelial cells. BMC Microbiol. 2014, 14, 177. [Google Scholar] [CrossRef] [PubMed]
- Panpetch, W.; Chancharoenthana, W.; Bootdee, K.; Nilgate, S.; Finkelman, M.; Tumwasorn, S.; Leelahavanichkul, A. Lactobacillus rhamnosus L34 Attenuates Gut Translocation-Induced Bacterial Sepsis in Murine Models of Leaky Gut. Infect. Immun. 2018, 86, 10–1128. [Google Scholar] [CrossRef] [PubMed]
- Witayavanitkul, N.; Werawatganon, D.; Chayanupatkul, M.; Klaikeaw, N.; Siriviriyakul, P. Genistein and exercise treatment reduced NASH related HDAC3, IL-13 and MMP-12 expressions in ovariectomized rats fed with high fat high fructose diet. J Tradit. Complement Med. 2021, 11, 503–512. [Google Scholar] [CrossRef]
- Brunt, E.M.; Janney, C.G.; Di Bisceglie, A.M.; Neuschwander-Tetri, B.A.; Bacon, B.R. Nonalcoholic steatohepatitis: A proposal for grading and staging the histological lesions. Am. J. Gastroenterol. 1999, 94, 2467–2474. [Google Scholar] [CrossRef]
- Deutsch, M.J.; Schriever, S.C.; Roscher, A.A.; Ensenauer, R. Digital image analysis approach for lipid droplet size quantitation of Oil Red O-stained cultured cells. Anal. Biochem. 2014, 445, 87–89. [Google Scholar] [CrossRef] [PubMed]
- Callahan, B.J.; McMurdie, P.J.; Rosen, M.J.; Han, A.W.; Johnson, A.J.; Holmes, S.P. DADA2: High-resolution sample inference from Illumina amplicon data. Nat. Methods. 2016, 13, 581–583. [Google Scholar] [CrossRef]
- Dhariwal, A.; Chong, J.; Habib, S.; King, I.L.; Agellon, L.B.; Xia, J. MicrobiomeAnalyst: A web-based tool for comprehensive statistical, visual and meta-analysis of microbiome data. Nucleic. Acids. Res. 2017, 45, W180–W188. [Google Scholar] [CrossRef] [PubMed]
- McMurdie, P.J.; Holmes, S. Waste not, want not: Why rarefying microbiome data is inadmissible. PLoS Comput. Biol. 2014, 10, e1003531. [Google Scholar] [CrossRef] [PubMed]
- Aderem, A.; Ulevitch, R.J. Toll-like receptors in the induction of the innate immune response. Nature 2000, 406, 782–787. [Google Scholar] [CrossRef] [PubMed]
- Solga, S.F.; Diehl, A.M. Non-alcoholic fatty liver disease: Lumen–liver interactions and possible role for probiotics. J. Hepatol. 2003, 38, 681–687. [Google Scholar] [CrossRef] [PubMed]
- Wang, H.; Mehal, W.; Nagy, L.E.; Rotman, Y. Immunological mechanisms and therapeutic targets of fatty liver diseases. Cell Mol. Immunol. 2021, 18, 73–91. [Google Scholar] [CrossRef] [PubMed]
- Sakai, Y.; Arie, H.; Ni, Y.; Zhuge, F.; Xu, L.; Chen, G.; Nagata, N.; Suzuki, T.; Kaneko, S.; Ota, T.; et al. Lactobacillus pentosus strain S-PT84 improves steatohepatitis by maintaining gut permeability. J. Endocrinol. 2020, 247, 169–181. [Google Scholar] [CrossRef]
- Khan, I.; Kang, S.C. Probiotic potential of nutritionally improved Lactobacillus plantarum DGK-17 isolated from Kimchi—A traditional Korean fermented food. Food. Control. 2016, 60, 88–94. [Google Scholar] [CrossRef]
- Tokatli, M.; Gulgor, G.; Bağder Elmacı, S.; İşleyen, N.; Ozçelik, F. In Vitro Properties of Potential Probiotic Indigenous Lactic Acid Bacteria Originating from Traditional Pickles. BioMed. Res. Int. 2015, 1, 1–8. [Google Scholar] [CrossRef]
- Cao, F.; Ding, Q.; Zhuge, H.; Lai, S.; Chang, K.; Le, C.; Yang, G.; Valencak, T.G.; Li, S.; Ren, D. Lactobacillus plantarum ZJUIDS14 alleviates non-alcoholic fatty liver disease in mice in association with modulation in the gut microbiota. Front. Nutr. 2022, 9, 1071284. [Google Scholar] [CrossRef] [PubMed]
- Kim, B.; Park, K.-Y.; Ji, Y.; Park, S.; Holzapfel, W.; Hyun, C.-K. Protective effects of Lactobacillus rhamnosus GG against dyslipidemia in high-fat diet-induced obese mice. Biochem. Biophys. Res. Commun. 2016, 473, 530–536. [Google Scholar] [CrossRef]
- Okubo, H.; Sakoda, H.; Kushiyama, A.; Fujishiro, M.; Nakatsu, Y.; Fukushima, T.; Matsunaga, Y.; Kamata, H.; Asahara, T.; Yoshida, Y.; et al. Lactobacillus casei strain Shirota protects against nonalcoholic steatohepatitis development in a rodent model. Am. J. Physiol. Gastrointest Liver Physiol. 2013, 305, G911–G918. [Google Scholar] [CrossRef]
- Wagnerberger, S.; Spruss, A.; Kanuri, G.; Stahl, C.; Schröder, M.; Vetter, W.; Bischoff, S.C.; Bergheim, I. Lactobacillus casei Shirota protects from fructose-induced liver steatosis: A mouse model. J. Nutr. Biochem. 2013, 24, 531–538. [Google Scholar] [CrossRef] [PubMed]
- Whittaker, R.H. Evolution and Measurement of Species Diversity. Taxon 1972, 21, 213–251. [Google Scholar] [CrossRef]
- Zhang, C.; Zhang, M.; Pang, X.; Zhao, Y.; Wang, L.; Zhao, L. Structural resilience of the gut microbiota in adult mice under high-fat dietary perturbations. ISME J. 2012, 6, 1848–1857. [Google Scholar] [CrossRef] [PubMed]
- Rivero-Gutiérrez, B.; Gámez-Belmonte, R.; Suárez, M.D.; Lavín, J.L.; Aransay, A.M.; Olivares, M.; Martínez-Augustin, O.; Sánchez de Medina, F.; Zarzuelo, A. A synbiotic composed of Lactobacillus fermentum CECT5716 and FOS prevents the development of fatty acid liver and glycemic alterations in rats fed a high fructose diet associated with changes in the microbiota. Mol. Nutr. Food Res. 2017, 61, 1600622. [Google Scholar] [CrossRef]
- Liu, L.; Fu, Q.; Li, T.; Shao, K.; Zhu, X.; Cong, Y.; Zhao, X. Gut microbiota and butyrate contribute to nonalcoholic fatty liver disease in premenopause due to estrogen deficiency. PLoS ONE 2022, 17, e0262855. [Google Scholar] [CrossRef]
- Parker, B.J.; Wearsch, P.A.; Veloo, A.C.M.; Rodriguez-Palacios, A. The Genus Alistipes: Gut Bacteria With Emerging Implications to Inflammation, Cancer, and Mental Health. Front. Immunol. 2020, 11, 906. [Google Scholar] [CrossRef]
- Zhang, X.; Coker, O.O.; Chu, E.S.; Fu, K.; Lau, H.C.H.; Wang, Y.X.; Chan, A.W.H.; Wei, H.; Yang, X.; Sung, J.J.Y.; et al. Dietary cholesterol drives fatty liver-associated liver cancer by modulating gut microbiota and metabolites. Gut 2021, 70, 761–774. [Google Scholar] [CrossRef]
- Jasirwan, C.O.M.; Lesmana, C.R.A.; Hasan, I.; Sulaiman, A.S.; Gani, R.A. The role of gut microbiota in non-alcoholic fatty liver disease: Pathways of mechanisms. Biosci. Microbiota Food Health 2019, 38, 81–88. [Google Scholar] [CrossRef] [PubMed]
- Kirsch, R.; Clarkson, V.; Verdonk, R.C.; Marais, A.D.; Shephard, E.G.; Ryffel, B.; de la MHall, P. Rodent nutritional model of steatohepatitis: Effects of endotoxin (lipopolysaccharide) and tumor necrosis factor alpha deficiency. J. Gastroenterol Hepatol. 2006, 21, 174–182. [Google Scholar] [CrossRef] [PubMed]
- Parks, B.W.; Nam, E.; Org, E.; Kostem, E.; Norheim, F.; Hui, S.T.; Pan, C.; Civelek, M.; Rau, C.D.; Bennett, B.J.; et al. Genetic control of obesity and gut microbiota composition in response to high-fat, high-sucrose diet in mice. Cell Metab. 2013, 17, 141–152. [Google Scholar] [CrossRef] [PubMed]
- Murphy, E.A.; Velazquez, K.T.; Herbert, K.M. Influence of high-fat diet on gut microbiota: A driving force for chronic disease risk. Curr. Opin. Clin. Nutr. Metab. Care 2015, 18, 515–520. [Google Scholar] [CrossRef]
- Zhou, D.; Pan, Q.; Shen, F.; Cao, H.X.; Ding, W.J.; Chen, Y.W.; Fan, J.G. Total fecal microbiota transplantation alleviates high-fat diet-induced steatohepatitis in mice via beneficial regulation of gut microbiota. Sci. Rep. 2017, 7, 1529. [Google Scholar] [CrossRef] [PubMed]
- Zhang, X.; Zhao, Y.; Zhang, M.; Pang, X.; Xu, J.; Kang, C.; Li, M.; Zhang, C.; Zhang, Z.; Zhang, Y.; et al. Structural changes of gut microbiota during berberine-mediated prevention of obesity and insulin resistance in high-fat diet-fed rats. PLoS ONE 2012, 7, e42529. [Google Scholar] [CrossRef]
- Tang, C.; Zhou, W.; Shan, M.; Lu, Z.; Lu, Y. Yogurt-derived Lactobacillus plantarum Q16 alleviated high-fat diet-induced non-alcoholic fatty liver disease in mice. Food Sci. Hum. Wellness 2022, 11, 1428–1439. [Google Scholar] [CrossRef]
- Wang, G.; Jiao, T.; Xu, Y.; Li, D.; Si, Q.; Hao, J.; Zhao, J.; Zhang, H.; Chen, W. Bifidobacterium adolescentis and Lactobacillus rhamnosus alleviate non-alcoholic fatty liver disease induced by a high-fat, high-cholesterol diet through modulation of different gut microbiota-dependent pathways. Food Funct. 2020, 11, 6115–6127. [Google Scholar] [CrossRef]
Group | n | Steatosis | Inflammation | Hepatocyte Ballooning | NASH Score | ||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
0 | 1 | 2 | 3 | 0 | 1 | 2 | 3 | 0 | 1 | 2 | |||
Control | 7 | 7 | - | - | - | 6 | 1 | - | - | 6 | 1 | - | 0 ± 0 |
NASH | 7 | - | - | 1 | 6 | - | - | 6 | 1 | - | - | 7 | 7 ± 0 * |
NASH + Single Strain | 7 | 1 | 2 | 3 | 1 | 3 | 4 | - | - | - | - | 7 | 5 ± 2 *,# |
NASH + Mixed Strains | 7 | - | 3 | 3 | 1 | 1 | 6 | - | - | - | 2 | 5 | 5 ± 2 *,# |
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Chayanupatkul, M.; Machchimapiro, P.; Chuaypen, N.; Wanpiyarat, N.; Tumwasorn, S.; Siriviriyakul, P.; Werawatganon, D. Single and Mixed Strains of Probiotics Reduced Hepatic Fat Accumulation and Inflammation and Altered Gut Microbiome in a Nonalcoholic Steatohepatitis Rat Model. Biomedicines 2024, 12, 1847. https://doi.org/10.3390/biomedicines12081847
Chayanupatkul M, Machchimapiro P, Chuaypen N, Wanpiyarat N, Tumwasorn S, Siriviriyakul P, Werawatganon D. Single and Mixed Strains of Probiotics Reduced Hepatic Fat Accumulation and Inflammation and Altered Gut Microbiome in a Nonalcoholic Steatohepatitis Rat Model. Biomedicines. 2024; 12(8):1847. https://doi.org/10.3390/biomedicines12081847
Chicago/Turabian StyleChayanupatkul, Maneerat, Panrawee Machchimapiro, Natthaya Chuaypen, Natcha Wanpiyarat, Somying Tumwasorn, Prasong Siriviriyakul, and Duangporn Werawatganon. 2024. "Single and Mixed Strains of Probiotics Reduced Hepatic Fat Accumulation and Inflammation and Altered Gut Microbiome in a Nonalcoholic Steatohepatitis Rat Model" Biomedicines 12, no. 8: 1847. https://doi.org/10.3390/biomedicines12081847