Sains Malaysiana 51(6)(2022): 1821-1832

http://doi.org/10.17576/jsm-2022-5106-18

Comparison of Non-Alcoholic Fatty Liver Disease (NAFLD) Model using Diet-Induced Nafld Mice with Genetically Modified Mice

(Perbandingan Model Penyakit Hati Berlemak (NAFLD) menggunakan Tikus Nafld Diet-Teraruh dengan Tikus Terubah Suai secara Genetik)

MOHD DANIAL MOHD EFENDY GOON^1,2, SHARANIZA AB RAHIM³, NORMALA ABD LATIP⁴, MARDIANA ABDUL AZIZ⁵, NORIZAL MOHD NOOR⁵, LEW SOOK WEIH⁶, MUSALMAH MAZLAN³ & SITI HAMIMAH SHEIKH ABDUL KADIR^1,2,3,*

¹Institute of Molecular Medicine Biotechnology, Faculty of Medicine, Universiti Teknologi MARA (UiTM), Cawangan Selangor, 47000 Sungai Buloh, Selangor Darul Ehsan, Malaysia

²Institute of Pathology, Laboratory and Forensic Medicine (I-PPerForM), Faculty of Medicine, Universiti Teknologi MARA (UiTM), Cawangan Selangor, 47000 Sungai Buloh, Selangor Darul Ehsan, Malaysia

³Department of Biochemistry, Faculty of Medicine, Universiti Teknologi MARA (UiTM), Cawangan Selangor, 47000 Sungai Buloh, Selangor Darul Ehsan, Malaysia

⁴Atta-ur-Rahman Institute for Natural Products Discovery (AuRIns), Faculty of Pharmacy, Universiti Teknologi MARA (UiTM), Cawangan Selangor, 42300 Puncak Alam, Selangor Darul Ehsan, Malaysia

⁵Department of Pathology, Faculty of Medicine, Universiti Teknologi MARA (UiTM), Cawangan Selangor, 47000 Sungai Buloh, Selangor Darul Ehsan, Malaysia

⁶Department of Pediatrics, Faculty of Medicine, Universiti Teknologi MARA (UiTM), Cawangan Selangor, 47000 Sungai Buloh, Selangor Darul Ehsan, Malaysia

Diserahkan: 11 Ogos 2021/Diterima: 5 November 2021

ABSTRACT

Prevalence of non-alcoholic fatty liver disease (NAFLD) is increasing steadily every year affecting all population both Western and Asian countries. The current treatments available for NAFLD are non-conclusive warranting newer effective pharmacological agents. Newly formulated agents require prior testing using animal models. However, in developing countries, these models are often costly. The possibility of using more affordable animal model in local settings should be investigated. In this study, ten Institute of Cancer Research (ICR) and seven B6.Cg-LepOb/J leptin-knockout (JAX) male mice were recruited. Five ICR and all JAX mice were subjected to high-fat diet (60% kcal fat) and remaining ICR mice were given standard diet (SD) for six weeks. Body weight and food intake were measured weekly while abdominal circumference, random blood glucose and liver span were measured at the end of the HFD study. Livers collected were subjected to histology assessment. Compared to ICR group, JAX group presented with significantly higher body weight (58 ± 0.72, p<0.05), larger body weight changes (16.57 ± 0.81, p<0.05), more HFD intake (197.14 ± 0.812, p<0.05) and larger abdominal circumference (11.79 ± 0.34: p<0.05). Liver from JAX group appeared with general steatosis and presentation of high-grade panacinar steatosis, low number of lobular inflammations and minimal fibrosis. Liver of ICR mice showed Zone 3 steatosis with high number of lobular inflammations without fibrosis. The NAFLD characteristics presented in JAX group suggested that B6.Cg-LepOb/J mice developed characteristics of NAFLD resembling human while ICR is suitable NAFLD model resembling human population resilient towards NAFLD.

Keywords: Animal model; B6.Cg-LepOb/J strain; high-fat diet; histology; Institute of Cancer Research strain; NAFLD

ABSTRAK

Kelaziman penyakit hati berlemak atau dikenali sebagai penyakit hati berlemak bukan alkohol (NAFLD) meningkat setiap tahun dalam kalangan populasi negara Barat dan Asia. Pada masa ini, rawatan bagi penyakit ini adalah tidak khusus, lalu mendorong kepada penyelidikan dalam mendapatkan agen rawatan yang berkesan. Rumusan agen rawatan yang baharu perlu dikaji menggunakan model haiwan. Penggunaan model haiwan dalam kalangan negara membangun membabitkan kos yang tinggi. Oleh itu, penyelidikan menggunakan model haiwan mengikut tetapan tempatan perlu dikaji. Dalam kajian ini, sepuluh ekor tikus jantan dan tujuh ekor tikus jantan B6.Cg-LepOb/J tanpa leptin (JAX) diambil daripada Institut Penyelidikan Kanser (ICR). Lima daripada tikus ICR dan kesemua tikus JAX diberi diet lemak tinggi (HFD) mengandungi 60% kcal daripada lemak selama enam minggu, dan selebihnya tikus ICR diberi diet piawai (SD). Berat badan dan pengambilan diet diukur setiap minggu manakala ukur lilit abdomen, bacaan glukosa darah rawak dan panjang hati diukur pada akhir tempoh kajian. Hati yang diperoleh daripada setiap tikus menjalani analisis histologi. Berbanding dengan kumpulan ICR, kumpulan JAX menunjukkan berat badan lebih berat (58 ± 0.72, p<0.05), perubahan berat badan lebih ketara (16.57 ± 0.81, p<0.05), pengambilan HFD lebih banyak (197.14 ± 0.812, p<0.05) dan ukur lilit abdomen lebih besar (11.79 ± 0.34: p<0.05). Analisis histologi kumpulan JAX menunjukkan kehadiran lemak (steatosis) dalam hati bertaraf tinggi, bilangan radang lobular dan parut yang rendah. Hati daripada kumpulan ICR menunjukkan gred 3 steatosis dan bilangan radang lobular yang tinggi. Ciri-ciri NAFLD yang ditonjolkan oleh kumpulan JAX menunjukkan model ini lebih mirip ke ciri-ciri NAFLD manusia manakala kumpulan ICR menunjukkan ciri-ciri ke arah manusia yang mempunyai kerintangan terhadap NAFLD.

Kata kunci: Diet lemak tinggi; histologi; model haiwan; NAFLD; strain B6.Cg-LepOb/J; strain Institut Penyelidikan Kanser

RUJUKAN

Anderson, E.L., Howe, L.D., Jones, H.E., Higgins, J.P., Lawlor, D.A. & Fraser, A. 2015. The prevalence of non-alcoholic fatty liver disease in children and adolescents: A systematic review and meta-analysis. PLoS ONE 10(10): e0140908.

Avtanski, D., Pavlov, V.A., Tracey, K.J. & Poretsky, L. 2019. Characterization of inflammation and insulin resistance in high‐fat diet‐induced male C57BL/6J mouse model of obesity. Animal Models and Experimental Medicine 2(4): 252-258.

B’chir, W., Dufour, C.R., Ouellet, C., Yan, M., Tam, I.S., Andrzejewski, S., Xia, H., Nabata, K., St-Pierre, J. & Giguère, V. 2018. Divergent role of estrogen-related receptor α in lipid-and fasting-induced hepatic steatosis in mice. Endocrinology 159(5): 2153-2164.

Besse-Patin, A., Léveillé, M., Oropeza, D., Nguyen, B.N., Prat, A. & Estall, J.L. 2017. Estrogen signals through peroxisome proliferator-activated Receptor− γ coactivator 1α to reduce oxidative damage associated with diet-induced fatty liver disease. Gastroenterology 152(1): 243-256.

Brix, A.E., Elgavish, A., Nagy, T.R., Gower, B.A., Rhead, W.J. & Wood, P.A. 2002. Evaluation of liver fatty acid oxidation in the leptin-deficient obese mouse. Molecular Genetics and Metabolism 75(3): 219-226.

Chalasani, N., Younossi, Z., Lavine, J.E., Charlton, M., Cusi, K., Rinella, M., Harrison, S.A., Brunt, E.M. & Sanyal, A.J. 2018. The diagnosis and management of nonalcoholic fatty liver disease: Practice guidance from the American Association for the Study of Liver Diseases. Hepatology 67(1): 328-357.

Chan, W.K., Tan, A.T.B., Vethakkan, S.R., Tah, P.C., Vijayananthan, A. & Goh, K.L. 2015. Low physical activity and energy dense Malaysian foods are associated with non-alcoholic fatty liver disease in centrally obese but not in non-centrally obese patients with diabetes mellitus. Asia Pacific Journal of Clinical Nutrition 24(2): 289-298.

Chang, Y., Jung, H.S., Cho, J., Zhang, Y., Yun, K.E., Lazo, M., Pastor-Barriuso, R., Ahn, J., Kim, C.W., Rampal, S. & Cainzos-Achirica, M. 2016. Metabolically healthy obesity and the development of nonalcoholic fatty liver disease. Official Journal of the American College of Gastroenterology 111(8): 1133-1140.

Chiba, T., Suzuki, S., Sato, Y., Itoh, T. & Umegaki, K. 2016. Evaluation of methionine content in a high-fat and choline-deficient diet on body weight gain and the development of non-alcoholic steatohepatitis in mice. PLoS ONE 11(10): e0164191.

Clapper, J.R., Hendricks, M.D., Gu, G., Wittmer, C., Dolman, C.S., Herich, J., Athanacio, J., Villescaz, C., Ghosh, S.S., Heilig, J.S. & Lowe, C. 2013. Diet-induced mouse model of fatty liver disease and nonalcoholic steatohepatitis reflecting clinical disease progression and methods of assessment. American Journal of Physiology-Gastrointestinal and Liver Physiology 305(7): G483-G495.

Domitrović, R., Jakovac, H., Tomac, J. & Šain, I. 2009. Liver fibrosis in mice induced by carbon tetrachloride and its reversion by luteolin. Toxicology and Applied Pharmacology 241(3): 311-321.

Duarte, J.A., Carvalho, F., Pearson, M., Horton, J.D., Browning, J.D., Jones, J.G. & Burgess, S.C. 2014. A high-fat diet suppresses de novo lipogenesis and desaturation but not elongation and triglyceride synthesis in mice [S]. Journal of Lipid Research 55(12): 2541-2553.

European Association for the Study of the Liver and European Association for the Study of Diabetes (EASD). 2016. EASL-EASD-EASO Clinical Practice Guidelines for the management of non-alcoholic fatty liver disease. Obesity Facts 9(2): 65-90.

Estes, C., Anstee, Q.M., Arias-Loste, M.T., Bantel, H., Bellentani, S., Caballeria, J., Colombo, M., Craxi, A., Crespo, J., Day, C.P., Eguchi, Y., Geier, A., Kondili,  L.A., Kroy, D.C., Lazarus, J.V., Loomba, R., Manns, M.P., Marchesini, G., Nakajima, A., Negro, F., Petta, S., Ratziu, V., Romero-Gomez, M., Sanyal, A., Schattenberg, J.M., Tacke, F., Tanaka, J., Trautwein, C., Lai, W., Zeuzem, S. & Razavi, H. 2018. Modeling NAFLD disease burden in China, France, Germany, Italy, Japan, Spain, United Kingdom, and United States for the period 2016-2030. Journal of Hepatology 69(4): 896-904.

Fakhry, T.K., Mhaskar, R., Schwitalla, T., Muradova, E., Gonzalvo, J.P. & Murr, M.M. 2019. Bariatric surgery improves nonalcoholic fatty liver disease: A contemporary systematic review and meta-analysis. Surgery for Obesity and Related Diseases 15(3): 502-511.

Fraulob, J.C., Ogg-Diamantino, R., Fernandes-Santos, C., Aguila, M.B. & Mandarim-de-Lacerda, C.A. 2010. A mouse model of metabolic syndrome: Insulin resistance, fatty liver and non-alcoholic fatty pancreas disease (NAFPD) in C57BL/6 mice fed a high fat diet. Journal of Clinical Biochemistry and Nutrition 1004080019-1004080019.

Gerbaix, M., Metz, L., Ringot, E. & Courteix, D. 2010. Visceral fat mass determination in rodent: Validation of dual-energy X-ray absorptiometry and anthropometric techniques in fat and lean rats. Lipids in Health and Disease 9(1): 1-9.

Guillaume, M., Riant, E., Fabre, A., Raymond‐Letron, I., Buscato, M., Davezac, M., Tramunt, B., Montagner, A., Smati, S., Zahreddine, R. & Palierne, G. 2019. Selective liver estrogen receptor α modulation prevents steatosis, diabetes, and obesity through the anorectic growth differentiation factor 15 hepatokine in mice. Hepatology Communications 3(7): 908-924.

Hebbard, L. & George, J. 2011. Animal models of nonalcoholic fatty liver disease. Nature Reviews Gastroenterology & Hepatology 8(1): 35-44.

Hung, C.K. & Bodenheimer, H.C. 2018. Current treatment of nonalcoholic fatty liver disease/nonalcoholic steatohepatitis. Clinics in Liver Disease 22(1): 175-187.

Kleiner, D.E., Brunt, E.M., Van Natta, M., Behling, C., Contos, M.J., Cummings, O.W., Ferrell, L.D., Liu, Y.C., Torbenson, M.S., Unalp-Arida, A. & Yeh, M. 2005. Design and validation of a histological scoring system for nonalcoholic fatty liver disease. Hepatology 41(6): 1313-1321.

Knuth, N.D., Johannsen, D.L., Tamboli, R.A., Marks‐Shulman, P.A., Huizenga, R., Chen, K.Y., Abumrad, N.N., Ravussin, E. & Hall, K.D. 2014. Metabolic adaptation following massive weight loss is related to the degree of energy imbalance and changes in circulating leptin. Obesity 22(12): 2563-2569.

Kristiansen, M.N.B., Veidal, S.S., Rigbolt, K.T.G., Tølbøl, K.S., Roth, J.D., Jelsing, J., Vrang, N. & Feigh, M. 2016. Obese diet-induced mouse models of nonalcoholic steatohepatitis-tracking disease by liver biopsy. World Journal of Hepatology 8(16): 673.

Kulkarni, N.M., Jaji, M.S., Shetty, P., Kurhe, Y.V., Chaudhary, S., Vijaykant, G., Raghul, J., Vishwakarma, S.L., Rajesh, B.N., Mookkan, J. & Krishnan, U.M. 2015. A novel animal model of metabolic syndrome with non-alcoholic fatty liver disease and skin inflammation. Pharmaceutical Biology 53(8): 1110-1117.

Leoni, S., Tovoli, F., Napoli, L., Serio, I., Ferri, S. & Bolondi, L. 2018. Current guidelines for the management of non-alcoholic fatty liver disease: A systematic review with comparative analysis. World Journal of Gastroenterology 24(30): 3361.

Li, J., Wu, H., Liu, Y. & Yang, L. 2020. High fat diet induced obesity model using four strains of mice: Kunming, C57BL/6, BALB/c and ICR. Experimental Animals 69(3): 326-335.

Liang, W., Menke, A.L., Driessen, A., Koek, G.H., Lindeman, J.H., Stoop, R., Havekes, L.M., Kleemann, R. & van den Hoek, A.M. 2014. Establishment of a general NAFLD scoring system for rodent models and comparison to human liver pathology. PLoS ONE 9(12): e115922.

Liu, Y., Meyer, C., Xu, C., Weng, H., Hellerbrand, C., ten Dijke, P. & Dooley, S. 2013. Animal   models of chronic liver diseases. American Journal of Physiology-Gastrointestinal and Liver Physiology 304(5): G449-G468.

Ma, L.L., Yuan, Y.Y., Zhao, M., Zhou, X.R., Jehangir, T., Wang, F.Y., Xi, Y. & Bu, S.Z. 2018. Mori Cortex extract ameliorates nonalcoholic fatty liver disease (NAFLD) and insulin resistance in high-fat-diet/streptozotocin-induced type 2 diabetes in rats. Chinese Journal of Natural Medicines 16(6): 411-417.

Machado, M.V., Michelotti, G.A., Xie, G., de Almeida, T.P., Boursier, J., Bohnic, B., Guy, C.D. & Diehl, A.M. 2015. Mouse models of diet-induced nonalcoholic steatohepatitis reproduce the heterogeneity of the human disease. PLoS ONE 10(5): e0127991.

Nagarajan, P., Kumar, M.J.M., Venkatesan, R., Majundar, S.S. & Juyal, R.C. 2012. Genetically modified mouse models for the study of nonalcoholic fatty liver disease. World Journal of Gastroenterology 18(11): 1141.

Obernier, J.A. & Baldwin, R.L. 2006. Establishing an appropriate period of acclimatization following transportation of laboratory animals. ILAR Journal 47(4): 364-369.

Palmisano, B.T., Zhu, L. & Stafford, J.M. 2017. Role of estrogens in the regulation of liver lipid metabolism. Advances in Experimental Medicine and Biology 1043: 227-256.

Pawar, S.V., Zanwar, V.G., Choksey, A.S., Mohite, A.R., Jain, S.S., Surude, R.G., Contractor, Q.Q., Rathi, P.M., Verma, R.U. & Varthakavi, P.K. 2017. Most overweight and obese Indian children have nonalcoholic fatty liver disease. Annals of Hepatology 15(6): 853-861.

Ricci, C., Baumgartner, J., Malan, L. & Smuts, C.M. 2020. Determining sample size adequacy for animal model studies in nutrition research: Limits and ethical challenges of ordinary power calculation procedures. International Journal of Food Sciences and Nutrition 71(2): 256-264.

Salsamendi, J., Pereira, K., Kang, K. & Fan, J. 2015. Minimally invasive percutaneous endovascular therapies in the management of complications of non-alcoholic fatty liver disease (NAFLD): A case report. Journal of Radiology Case Reports 9(9): 36.

Sanyal, A.J., Chalasani, N., Kowdley, K.V., McCullough, A., Diehl, A.M., Bass, N.M., Neuschwander-Tetri, B.A., Lavine, J.E., Tonascia, J., Unalp, A. & Van Natta, M. 2010. Pioglitazone, vitamin E, or placebo for nonalcoholic steatohepatitis. New England Journal of Medicine 362(18): 1675-1685.

Sayiner, M., Koenig, A., Henry, L. & Younossi, Z.M. 2016. Epidemiology of nonalcoholic fatty liver disease and nonalcoholic steatohepatitis in the United States and the rest of the world. Clinics in Liver Disease 20(2): 205-214.

Solinas, P., Isola, M., Lilliu, M.A., Conti, G., Civolani, A., Demelia, L., Loy, F. & Isola, R. 2014. Animal models are reliably mimicking human diseases? A morphological study that compares animal with human NAFLD. Microscopy Research and Technique 77(10): 790-796.

Sridharan, K., Sivaramakrishnan, G., Sequeira, R.P. & Elamin, A. 2018. Pharmacological interventions for non-alcoholic fatty liver disease: A systematic review and network meta-analysis. Postgraduate Medical Journal 94(1116): 556-565.

Van Herck, M.A., Vonghia, L. & Francque, S.M. 2017. Animal models of nonalcoholic fatty liver disease - A starter’s guide. Nutrients 9(10): 1072.

Wang, C., Tao, Q., Wang, X., Wang, X. & Zhang, X. 2016. Impact of high-fat diet on liver genes expression profiles in mice model of nonalcoholic fatty liver disease. Environmental Toxicology and Pharmacology 45: 52-62.

Wang, X.H., Li, C.Y., Muhammad, I. & Zhang, X.Y. 2016. Fatty acid composition in serum correlates with that in the liver and non-alcoholic fatty liver disease activity scores in mice fed a high-fat diet. Environmental Toxicology and Pharmacology 44: 140-150.

Wu, J. 2016. Utilization of animal models to investigate nonalcoholic steatohepatitis-associated hepatocellular carcinoma. Oncotarget 7(27): 42762.

Yahaghi, L., Ebrahim-Habibi, A., Hayati-Roodbari, N., Irani, S. & Yaghmaei, P. 2019. A simple method for inducing nonalcoholic steatohepatitis with fibrosis. Animal Models and Experimental Medicine 2(4): 282-290.

Yasmeen, R., Shen, Q., Lee, A., Leung, J.H., Kowdley, D., DiSilvestro, D.J., Xu, L., Yang, K., Maiseyeu, A., Bal, N.C. & Periasamy, M. 2018. Epiregulin induces leptin secretion and energy expenditure in high-fat diet-fed mice. Journal of Endocrinology 239(3): 377-388.

Younossi, Z.M., Marchesini, G., Pinto-Cortez, H. & Petta, S. 2019. Epidemiology of nonalcoholic fatty liver disease and nonalcoholic steatohepatitis: Implications for liver transplantation. Transplantation 103(1): 22-27.

Younossi, Z.M., Koenig, A.B., Abdelatif, D., Fazel, Y., Henry, L. & Wymer, M. 2016. Global epidemiology of nonalcoholic fatty liver disease-Meta-analytic assessment of prevalence, incidence, and outcomes. Hepatology 64(1): 73-84.

Zhang, Y.H., Ma, D.Q., Ding, D.P., Li, J., Chen, L.L., Ao, K.J. & Tian, Y.Y. 2018. S100A4 gene is crucial for methionine-choline-deficient diet-induced non-alcoholic fatty liver disease in mice. Yonsei Medical Journal 59(9): 1064-1071.

^*Pengarang untuk surat-menyurat; email: sitih587@uitm.edu.my