亮氨酸对黄牛皮下脂肪细胞棕色化的影响

doi:10.11843/j.issn.0366-6964.2023.08.015

Abstract

Abstract: This experiment was conducted to isolate subcutaneous preadipocytes from yellow cattle and study the effect of leucine on browning white adipocytes and lipid metabolism. Preadipocytes were isolated from subcutaneous adipose tissue of 4 3-year-old healthy male yellow cattle ((351.7±42.5) kg), mixed and added with different concentrations of leucine (0, 0.5, 1, 2, 4 and 8 mmol·L^-1) at the differentiation D 6-D 9 (n=6). RT-PCR was used to detect the expression level of key browning genes to select leucine concentrations that induce browning, and follow-up experiments were executed to set the control group (CON) and optimal concentration leucine addition group (Leu) (n=6). RT-PCR was used to detect the key mitochondrial biosynthesis genes expression, mtDNA copy number and genes expression related to lipid metabolism. The expression of key proteins of browning as well as pathway proteins were detected by Western Blot. Intracellular mitochondrial content and lipid droplet morphology were detected by Mito-tracker Green staining and Oil Red O staining, respectively.The results showed that subcutaneous preadipocytes were isolated and differentiated into mature adipocytes successfully. 4 mmol·L^-1 leucine significantly increased the relative mRNA expression of browning marker factors UCP1, PRDM16 and TMEM26 and key genes of mitochondrial biosynthesis SIRT1, TFAM and NRF1/2, and increased the protein expression of UCP1, CD137 and SIRT1 in adipocytes (P<0.01). Leucine significantly increased the mtDNA copy number and mitochondrial content (P<0.01), decreased the intracellular ATP content (P<0.05). Leucine induced the transformation of lipid droplets from large single-compartment to small multi-compartment and decreased intracellular triglyceride content (P<0.01). Leucine decreased the expression of lipid synthesis genes ACC and FAS, while promoting the expression of lipolysis genes ATGL and HSL and fatty acid β-oxidation genes PPARα, CPT1 and ACOX1 (P<0.01). Leucine also significantly upregulated protein expression levels of p-AMPKα/AMPKα and PGC1α (P<0.01). In conclusion, leucine promotes browning and mitochondrial biosynthesis in preadipocytes, promotes lipolysis and inhibits lipid synthesis, reduces lipid deposition, and upregulates the protein expression of p-AMPKα/AMPKα and PGC1α.

Key words: leucine, yellow cattle, primary adipocytes, browning, mitochondrial biosynthesis

CLC Number:

S823.81

GUO Yixin, WANG Zhisheng, HU Rui, WANG Junmei, WANG Sen, SHI Liyuan, ZHANG Xiaohong, ZOU Huawei, ZUO Jiaxue, PENG Quanhui, XUE Bai, WANG Lizhi. Effect of Leucine on Browning of Subcutaneous Adipocytes in Yellow Cattle[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3286-3298.

References 37

[1]	ZHANG Z Y, YANG D, XIANG J W, et al.Non-shivering thermogenesis signalling regulation and potential therapeutic applications of brown adipose tissue[J].Int J Biol Sci, 2021, 17(11):2853-2870.
[2]	张聪聪, 张馨心, 郭思玮, 等.维生素A对肉牛脂肪沉积及其调控的研究进展[J].中国畜牧兽医, 2021, 48(10):3652-3659.ZHANG C C, ZHANG X X, GUO S W, et al.Research progress of vitamin A on fat deposition and its regulation in beef cattle[J].China Animal Husbandry & Veterinary Medicine, 2021, 48(10):3652-3659.(in Chinese)
[3]	GU X, WANG L Y, LIU S Q, et al.Adipose tissue adipokines and lipokines:functions and regulatory mechanism in skeletal muscle development and homeostasis[J].Metabolism, 2023, 139:155379.
[4]	QIN X, PARK H G, ZHANG J Y, et al.Brown but not white adipose cells synthesize omega-3 docosahexaenoic acid in culture[J].Prostaglandins Leukot Essent Fatty Acids, 2016, 104:19-24.
[5]	WEI S J, ZHANG M M, ZHENG Y Y, et al.ZBTB16 overexpression enhances white adipogenesis and induces brown-like adipocyte formation of bovine white intramuscular preadipocytes[J].Cell Physiol Biochem, 2018, 48(6):2528-2538.
[6]	WU J, BOSTRÖM P, SPARKS L M, et al.Beige adipocytes are a distinct type of thermogenic fat cell in mouse and human[J]. Cell, 2012, 150(2):366-376.
[7]	MUKHERJEE S, YUN J W.Prednisone stimulates white adipocyte browning via β3-AR/p38 MAPK/ERK signaling pathway[J]. Life Sci, 2022, 288:120204.
[8]	BARGUT T C L, SOUZA-MELLO V, AGUILA M B, et al.Browning of white adipose tissue:lessons from experimental models[J]. Horm Mol Biol Clin Investig, 2017, 31(1).20160051.
[9]	焦俊.亮氨酸通过脂肪组织控制肥胖的机制研究[D].苏州:苏州大学, 2016.JIAO J.Effect of Leucine on controlling obesity and the Underlying Mechanism[D].Suzhou:Soochow University, 2016.(in Chinese)
[10]	ZHANG L Y, LI F N, GUO Q P, et al.Leucine supplementation:a novel strategy for modulating lipid metabolism and energy homeostasis[J].Nutrients, 2020, 12(5):1299.
[11]	DUAN Y H, LI F N, LIU H N, et al.Nutritional and regulatory roles of leucine in muscle growth and fat reduction[J].Front Biosci (Landmark Ed), 2015, 20(4):796-813.
[12]	MA Q Q, ZHOU X B, HU L L, et al.Leucine and isoleucine have similar effects on reducing lipid accumulation, improving insulin sensitivity and increasing the browning of WAT in high-fat diet-induced obese mice[J].Food Funct, 2020, 11(3):2279-2290.
[13]	郭红芳, 昝林森, 孙永刚.牛前体脂肪细胞的分离培养及诱导分化[J].西北农林科技大学学报:自然科学版, 2014, 42(2):1-6, 12.GUO H F, ZAN L S, SUN Y G.Primary culture and differentiation of bovine preadipocytes[J].Journal of Northwest A&F University:Natural Science Edition, 2014, 42(2):1-6, 12.(in Chinese)
[14]	王森, 师俊华, 王之盛, 等.牦牛不同部位前体脂肪细胞分离鉴定及分化关键基因表达研究[J].畜牧兽医学报, 2022, 53(3):755-765.WANG S, SHI J H, WANG Z S, et al.Isolation and Identification of preadipocytes from different parts of yak and expression of key genes for differentiation[J].Acta Veterinaria et Zootechnica Sinica, 2022, 53(3):755-765.(in Chinese)
[15]	VENEGAS V, WANG J, DIMMOCK D, et al.Real-time quantitative PCR analysis of mitochondrial DNA content[J].Curr Protoc Hum Genet, 2011, doi:10.1002/0471142905.hg1907s68.
[16]	余湘.牛原代前体脂肪细胞的分离及增殖、分化功能microRNA的筛选[D].长春:吉林大学, 2020.YU X.Isolation of primary bovine preadipocytes and screening of microRNAs associated with proliferation and adipogenesis[D]. Changchun:Jilin University, 2020.(in Chinese)
[17]	张萌萌.ZBTB16基因过表达对牛肌内前体脂肪细胞增殖与分化的影响研究[D].南京:南京农业大学, 2018.ZHANG M M.Effects of ZBTB16 overexpression on proliferation and differentiation of bovine intramuscular preadipocytes[D].Nanjing:Nanjing Agricultural University, 2018.(in Chinese)
[18]	YAMAMOTO T, DIAO Z C, MURAKAMI M, et al.Factors affecting the induction of uncoupling protein 1 in C2C12 myogenic cells[J].Cytokine, 2022, 157:155936.
[19]	KIM K, NAM K H, YI S A, et al.Ginsenoside Rg3 induces browning of 3T3-L1 adipocytes by activating AMPK signaling[J]. Nutrients, 2020, 12(2):427.
[20]	CHOI M, MUKHERJEE S, YUN J W.Trigonelline induces browning in 3T3-L1 white adipocytes[J].Phytother Res, 2021, 35(2):1113-1124.
[21]	SON Y, CHOI C, SONG C, et al.Development of CIDEA reporter mouse model and its application for screening thermogenic drugs[J].Sci Rep, 2021, 11(1):18429.
[22]	WANG G X, MEYER J G, CAI W K, et al.Regulation of UCP1 and mitochondrial metabolism in brown adipose tissue by reversible succinylation[J].Mol Cell, 2019, 74(4):844-857.e7.
[23]	JUNG T W, HWANG E J, PYUN D H, et al.3-hydroxymorphinan enhances mitochondrial biogenesis and adipocyte browning through AMPK-dependent pathway[J].Biochem Biophys Res Commun, 2021, 577:17-23.
[24]	KOWALD A, KIRKWOOD T B L.Evolution of the mitochondrial fusion-fission cycle and its role in aging[J].Proc Natl Acad Sci U S A, 2011, 108(25):10237-10242.
[25]	SUN X C, ZEMEL M B.Leucine modulation of mitochondrial mass and oxygen consumption in skeletal muscle cells and adipocytes[J].Nutr Metab (Lond), 2009, 6:26.
[26]	SCHNUCK J K, SUNDERLAND K L, GANNON N P, et al.Leucine stimulates PPARβ/δ-dependent mitochondrial biogenesis and oxidative metabolism with enhanced GLUT4 content and glucose uptake in myotubes[J].Biochimie, 2016, 128-129:1-7.
[27]	FAN H Y, DING R, LIU W C, et al.Heat shock protein 22 modulates NRF1/TFAM-dependent mitochondrial biogenesis and DRP1-sparked mitochondrial apoptosis through AMPK-PGC1α signaling pathway to alleviate the early brain injury of subarachnoid hemorrhage in rats[J].Redox Biol, 2021, 40:101856.
[28]	KIM H L, PARK J, PARK H, et al.Platycodon grandiflorum A. De Candolle Ethanolic extract inhibits adipogenic regulators in 3T3-L1 cells and induces mitochondrial biogenesis in primary brown preadipocytes[J].J Agric Food Chem, 2015, 63(35):7721-7730.
[29]	ZHANG H Q, CHEN S Y, WANG A S, et al.Sulforaphane induces adipocyte browning and promotes glucose and lipid utilization[J].Mol Nutr Food Res, 2016, 60(10):2185-2197.
[30]	CHOI J H, KIM S W, YU R N, et al.Monoterpene phenolic compound thymol promotes browning of 3T3-L1 adipocytes[J].Eur J Nutr, 2017, 56(7):2329-2341.
[31]	杨江涛.亮氨酸对3T3-L1脂肪细胞中脂肪分解和瘦素信号通路的影响[D].苏州:苏州大学, 2015.YANG J T.Effect of leucine on the fat metabolism and leptin signaling pathway in 3T3-L1 adipocyte[D].Suzhou:Soochow University, 2015.(in Chinese)
[32]	WHITEHEAD A, KRAUSE F N, MORAN A, et al.Brown and beige adipose tissue regulate systemic metabolism through a metabolite interorgan signaling axis[J].Nat Commun, 2021, 12(1):1905.
[33]	DU X J, OSORO E K, CHEN Q, et al.Pdcd4 promotes lipid deposition by attenuating PPARα-mediated fatty acid oxidation in hepatocytes[J].Mol Cell Endocrinol, 2022, 545:111562.
[34]	YANG Q Y, LIANG X W, SUN X F, et al.AMPK/α-ketoglutarate axis dynamically mediates DNA demethylation in the Prdm16 promoter and brown adipogenesis[J].Cell Metab, 2016, 24(4):542-554.
[35]	WU L Y, ZHANG L N, LI B H, et al.AMP-Activated Protein Kinase (AMPK) regulates energy metabolism through modulating thermogenesis in adipose tissue[J].Front Physiol, 2018, 9:122.
[36]	WANG S B, LIANG X W, YANG Q Y, et al.Resveratrol enhances brown adipocyte formation and function by activating AMP-activated protein kinase (AMPK) α1 in mice fed high-fat diet[J].Mol Nutr Food Res, 2017, 61(4):1600746.
[37]	MA T Y, HUANG X H, ZHENG H X, et al.SFRP2 improves mitochondrial dynamics and mitochondrial biogenesis, oxidative stress, and apoptosis in diabetic cardiomyopathy[J].Oxid Med Cell Longev, 2021, 2021:9265016.

Effect of Leucine on Browning of Subcutaneous Adipocytes in Yellow Cattle

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References 37

Related Articles 10

Recommended Articles

Metrics

Comments

[1]	YANG Xiaohua, LIU Fangfang, ZHANG Fenglin, YI Xin, CHEN Lin, SHU Gang, WANG Lina, ZHU Xiaotong, GAO Ping, JIANG Qingyan, WANG Songbo. Effects of Dietary Fish Oil on Estrous Cycles and Body Heat Production in Mice Fed High-fat Diets [J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(7): 1891-1902.
[2]	SHAO Jing, ZHANG Jiasu, YIN Baozhen, ZHANG Luomeng, XIA Guangjun. miR-17-3p Regulates Preadipocyte Differentiation by Targeting KCTD15 in Yanbian Yellow Cattle [J]. Acta Veterinaria et Zootechnica Sinica, 2020, 51(11): 2689-2698.
[3]	ZHANG Shou, JIN Guo-en, CHANG Lan,Lü Jun, SHEN Ming-hua, LIU Hui, GU Hai-yan, LEI Nai-hu. Comparison on Hypoxic Ventilatory Response and NO, NOS of Carotid Body between Yaks and Qaidam Yellow Cattle [J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2018, 49(10): 2276-2282.
[4]	CHANG Lan，ZHANG Shou，LEI Nai-hu，SHANG Guo-ma. Stereologic Study on Mitochondria and Electron Dense-cored Vesicles of Carotid Body Type Ⅰ Cells between Yak and Qaidam Yellow Cattle [J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2016, 47(7): 1474-1479.
[5]	CAO Chang-fu，SUN Mei-zhen，LU Yan，ZENG Qing-lin，LIU Wei，ZENG Zhen-ling. Pharmacokinetic of Marbofloxacin in Infancy Yellow Cattle after Single Intramuscular Administrations and ex vivo the Post-antibiotic Effects [J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2015, 46(11): 2078-2084.
[6]	WU Xiao-yan，WANG Zhi-sheng，ZOU Hua-wei. Different Protein Sources Dietary Affect the Quantities of Rumen Proteobacteria Adhensive to Solid Fractions in Xuanhan Yellow Cattle [J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2014, 45(6): 953-959.
[7]	YIN Duo;ZHU Yuxia;LIU Haixing;LI Zhongshu;FANG Nanzhu. Effects of Free Radical Absorptive Antioxidants on the Development of Somatic Cell Nuclear Transfer Yanbian Yellow Cattle Embryos [J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2012, 43(8): 1215-1221.
[8]	WANG Kun;ZHANG Yuan;SUN Dong-xiao. Analysis of Sequences of BoLA-DRB₃ Genes in Chinese Yellow Cattle [J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2008, 39(7): 853-857.
[9]	XIN Ya-ping;GAO Xue;GUO Ya-ning;WANG Hui-sheng;MA Yun;XU Shang-zhong;ZHANG Ying-han;ZAN Lin-sen;REN Hong-yan;CHEN Jin-bao. The Relationships of Blood Protein Polymorphism with Reproduction in Qinchuan Yellow Cattle [J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2006, 37(2): 193-198.
[10]	GAO Xue;XU Xiu-rong;XU Shang-zhong;ZHANG Ying-han. Genetic Polymorphisms of GH Genes in Chinese Yellow Cattle [J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2005, 36(10): 991-995.