Effect of Myostatin Gene Knockout on Browning of White Fat and Related Gene Expression Levels in a Mouse Model of Type 2 Diabetes Mellitus

doi:10.12114/j.issn.1007-9572.2022.0178

Chinese General Practice ›› 2022, Vol. 25 ›› Issue (21): 2610-2617.DOI: 10.12114/j.issn.1007-9572.2022.0178

Special Issue: 内分泌代谢性疾病最新文章合集

• Original Research • Previous Articles Next Articles

Effect of Myostatin Gene Knockout on Browning of White Fat and Related Gene Expression Levels in a Mouse Model of Type 2 Diabetes Mellitus

1. Graduate School, Kyonggi University, Suwon 16227, South Korea
2. Editorial Department, Henan Provincial People's Hospital, Zhengzhou University People's Hospital, Henan University People's Hospital, Zhengzhou 450003, China
3. Department of Endocrinology, Henan Provincial People's Hospital, Zhengzhou University People's Hospital, Henan University People's Hospital, Zhengzhou 450003, China

Received:2022-01-03 Revised:2022-04-24 Published:2022-07-20 Online:2022-03-04
Contact: Yanfang WANG
About author:
CHENG J W, LIU Y Q, WANG Y F. Effect of myostatin gene knockout on browning of white fat and related gene expression levels in a mouse model of type 2 diabetes mellitus[J]. Chinese General Practice, 2022, 25 (21) : 2610-2617.

肌肉生长抑制素基因敲除对2型糖尿病小鼠白色脂肪棕色化及相关基因表达的影响研究

1．16227韩国京畿道水原市，韩国京畿大学研究生院
2．450003　河南省郑州市，河南省人民医院　郑州大学人民医院　河南大学人民医院编辑部
3．450003　河南省郑州市，河南省人民医院　郑州大学人民医院　河南大学人民医院内分泌科

通讯作者: 汪艳芳
作者简介:
程经纬，柳杨青，汪艳芳.肌肉生长抑制素基因敲除对2型糖尿病小鼠白色脂肪棕色化及相关基因表达的影响研究[J].中国全科医学，2022，25（21）：2610-2617. [www.chinagp.net] 作者贡献：程经纬负责实验设计与实施，数据收集与统计学分析，论文的构思、设计、撰写；柳杨青负责实验设计与实施，论文修订；汪艳芳负责研究选题，实验的监督管理，论文的质量控制及审校，对论文整体负责。
基金资助:
河南省医学科技攻关计划省部共建项目(SBGJ2018069)

Abstract

Abstract:

Background

The browning of white fat is a hot spot in metabolic disease research. Myostatin (Mstn) inhibits muscle growth, and has correlation with the growth and differentiation of adipocytes, but its effect on the browning of white fat in type 2 diabetes mellitus (T2DM) is still unclear.

Objective

To examine changes in expression levels of genes related to the browning of white fat following Mstn gene knockout to assess the effect of Mstn on the browning of white fat in a mouse model of T2DM.

Methods

This study was conducted from January 2019 to January 2020. Thirty-six male, SPF, C57BL/6N mice were selected. The 12 wild-type (WT) mice were equally randomized into WT group and WT+DM group, 12 heterozygous mice with Mstn gene knockout〔Mstn (+/-) 〕 were equally randomized into Mstn (+/-) group and Mstn (+/-) +DM group, 12 homozygous mice with Mstn gene knockout〔Mstn (-/-) 〕 were equally randomized into Mstn (-/-) group and Mstn (-/-) +DM group. The WT group, Mstn (+/-) group, Mstn (-/-) group received a normal diet, and the WT+DM group, Mstn (+/-) +DM group, Mstn (-/-) +DM group received a high-fat diet and a small dose of streptozotocin to construct T2DM models. When the intervention was finished in all groups, weight, body length, white fat and brown fat mass, and serum lipids〔triacylglycerol (TG) , total cholesterol (TC) , low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) measured using automatic biochemical analyzer, and free fatty acid (FFA) measured using ELISA〕 were collected. The Lee's index, white-brown fat ratio and fat mass index were calculated. The morphology of white fat and brown fat cells was observed using HE staining. The relative expression levels of peroxisome proliferator-activated receptor gamma (PPAR-γ) , peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) , uncoupling protein 1 (UCP1) and cluster of differentiation 137 (CD137) protein in white and brown fat were determined by Western-blotting.

Results

Mstn (-/-) group had lower Lee's index, white-brown fat ratio and levels of serum TG and TC compared with WT group (P<0.05) . Mstn (-/-) group had lower Lee's index, white-brown fat ratio and level of serum TG than Mstn (+/-) group (P<0.05) . Compared with WT group, WT+DM group had lower Lee's index and level of serum HDL-C, and higher white-brown fat ratio, fat mass index, serum TG, TC, LDL-C and FFA levels (P<0.05) . Mstn (-/-) +DM group had lower levels of Lee's index, white-brown fat ratio, fat mass index, serum TC, TG, LDL-C and FFA, but higher serum HDL-C than did WT+DM group (P<0.05) . Mstn (-/-) +DM group had lower levels of Lee's index, white-brown fat ratio and serum TC, but higher serum HDL-C than did Mstn (+/-) +DM group (P<0.05) . Compared with WT group, Mstn (-/-) group had higher relative expression levels of PPAR-γ, PGC-1α, UCP1 and CD137 protein in white and brown fat (P<0.05) . Mstn (-/-) group had higher relative expression levels of PPAR-γ, PGC-1α and CD137 protein in white and brown fat than did Mstn (+/-) group (P<0.05) . The relative expression levels of PPAR-γ, PGC-1α, UCP1 and CD137 protein in white and brown fat in WT+DM group were lower than those in WT group (P<0.05) . The relative expression levels of PPAR-γ, PGC-1α, UCP1 and CD137 protein in white and brown fat in Mstn (-/-) +DM group were higher than those in WT+DM group or Mstn (+/-) +DM group (P<0.05) .

Conclusion

The inhibition of Mstn gene expression may be against T2DM-induced obesity phenotypes such as white fat accumulation and lipid metabolism disorder, and up-regulate the expression levels of PPAR-γ, PGC-1α, UCP1 and CD137 genes, promoting the browning of white fat.

Key words: Diabetes mellitus, type 2, Myostatin, White fat browning, Adipocytes, white, Adipocytes, brown, Mice

摘要：

背景

白色脂肪棕色化是目前代谢性疾病研究的热点，肌肉生长抑制素（Mstn）不仅可负向调控肌肉生长，还与脂肪细胞生长、分化有关，但Mstn对2型糖尿病（T2DM）白色脂肪棕色化的影响尚不明确。

目的

观察Mstn基因敲除的T2DM小鼠白色脂肪棕色化相关基因表达变化，探究Mstn对T2DM小鼠白色脂肪棕色化的影响。

方法

2019年1月至2020年1月，将12只野生型（WT）、12只杂合型Mstn基因敲除〔Mstn（+/-）〕、12只纯合型Mstn基因敲除〔Mstn（-/-）〕雄性SPF级C57BL/6N小鼠随机各分为2组，每组6只，分别为WT组、WT+DM组，Mstn（+/-）组、Mstn（+/-）+DM组，Mstn（-/-）组、Mstn（-/-）+DM组，其中WT组、Mstn（+/-）组、Mstn（-/-）组给予普通饮食，WT+DM组、Mstn（+/-）+DM组、Mstn（-/-）+DM组给予高脂饮食+小剂量链脲佐菌素诱导构建T2DM模型。造模后测定各组小鼠体质量、体长、白色脂肪及棕色脂肪质量，计算Lee's指数、白棕比、脂肪指数；检测血清三酰甘油（TG）、总胆固醇（TC）、低密度脂蛋白胆固醇（LDL-C）、高密度脂蛋白胆固醇（HDL-C）水平；采用酶联免疫吸附试验（ELISA）检测血清游离脂肪酸（FFA）水平；苏木精-伊红（HE）染色法分析白色脂肪及棕色脂肪细胞形态；Western-blotting法测定白色脂肪及棕色脂肪过氧化物酶体增殖物激活受体γ（PPAR-γ）、过氧化物酶体增殖物激活受体γ辅激活因子1α（PGC-1α）、解偶联蛋白1（UCP1）、分化簇137（CD137）蛋白相对表达量。

结果

Mstn（-/-）组Lee's指数、白棕比、血清TG水平低于WT组、Mstn（+/-）组（P<0.05），血清TC水平低于WT组（P<0.05）。WT+DM组Lee's指数、血清HDL-C水平低于WT组，白棕比、脂肪指数、血清TG、TC、LDL-C及FFA水平高于WT组（P<0.05）。Mstn（-/-）+DM组Lee's指数、白棕比、血清TC水平低于WT+DM组、Mstn（+/-）+DM组，血清HDL-C水平高于WT+DM组、Mstn（+/-）+DM组（P<0.05）。Mstn（-/-）+DM组脂肪指数、血清TG、LDL-C及FFA水平低于WT+DM组（P<0.05）。Mstn（-/-）组白色及棕色脂肪PPAR-γ、PGC-1α、CD137蛋白相对表达量高于WT组、Mstn（+/-）组（P<0.05），白色及棕色脂肪UCP1蛋白相对表达量高于WT组（P<0.05）。WT+DM组白色及棕色脂肪PPAR-γ、PGC-1α、UCP1、CD137蛋白相对表达量低于WT组（P<0.05）。Mstn（-/-）+DM组白色及棕色脂肪PPAR-γ、PGC-1α、UCP1、CD137蛋白相对表达量高于WT+DM组、Mstn（+/-）+DM组（P<0.05）。

结论

抑制Mstn基因表达可拮抗T2DM引起的白色脂肪堆积、脂代谢紊乱等肥胖表型，上调PPAR-γ、PGC-1α、UCP1、CD137基因表达，促进白色脂肪棕色化。

关键词: 糖尿病，2型, 肌肉生长抑制素, 白色脂肪棕色化, 脂肪细胞，白色, 脂肪细胞，棕色, 小鼠

Jingwei CHENG,Yangqing LIU,Yanfang WANG. Effect of Myostatin Gene Knockout on Browning of White Fat and Related Gene Expression Levels in a Mouse Model of Type 2 Diabetes Mellitus[J]. Chinese General Practice, 2022, 25(21): 2610-2617. DOI: 10.12114/j.issn.1007-9572.2022.0178.
程经纬,柳杨青,汪艳芳. 肌肉生长抑制素基因敲除对2型糖尿病小鼠白色脂肪棕色化及相关基因表达的影响研究[J]. 中国全科医学, 2022, 25(21): 2610-2617. DOI: 10.12114/j.issn.1007-9572.2022.0178.

Figures/Tables 8

References 16

[1]	SHARMA M, MCFARLANE C, KAMBADUR R，et al. Myostatin：expanding horizons[J]. IUBMB Life，2015，67（8）：589-600. DOI：10.1002/iub.1392.
[2]	AKPAN I, GONCALVES M D, DHIR R，et al. The effects of a soluble activin type IIB receptor on obesity and insulin sensitivity[J]. Int J Obes （Lond），2009，33（11）：1265-1273. DOI：10.1038/ijo.2009.162.
[3]	SUN W X, DODSON M V, JIANG Z H，et al. Myostatin inhibits porcine intramuscular preadipocyte differentiation in vitro[J]. Domest Anim Endocrinol，2016，55：25-31. DOI：10.1016/j.domaniend.2015.10.005.
[4]	SHAN T Z, LIANG X R, BI P P，et al. Myostatin knockout drives browning of white adipose tissue through activating the AMPK-PGC1α-Fndc5 pathway in muscle[J]. FASEB J，2013，27（5）：1981-1989. DOI：10.1096/fj.12-225755.
[5]	BAATI N, FEILLET-COUDRAY C, FOURET G，et al. Myostatin deficiency is associated with lipidomic abnormalities in skeletal muscles[J]. Biochim Biophys Acta Mol Cell Biol Lipids，2017，1862（10 Pt A）：1044-1055. DOI：10.1016/j.bbalip.2017.06.017.
[6]	CAI C B, QIAN L L, JIANG S W，et al. Loss-of-function myostatin mutation increases insulin sensitivity and browning of white fat in Meishan pigs[J]. Oncotarget，2017，8（21）：34911-34922. DOI：10.18632/oncotarget.16822.
[7]	BARTELT A, HEEREN J. Adipose tissue browning and metabolic health[J]. Nat Rev Endocrinol，2014，10（1）：24-36. DOI：10.1038/nrendo.2013.204.
[8]	TANAKA-YACHI R, TAKAHASHI-MUTO C, ADACHI K，et al. Promoting effect of α-tocopherol on beige adipocyte differentiation in 3T3-L1 cells and rat white adipose tissue[J]. J Oleo Sci，2017，66 （2）：171-179. DOI：10.5650/jos.ess16137.
[9]	DINAS P C, VALENTE A, GRANZOTTO M，et al. Browning formation markers of subcutaneous adipose tissue in relation to resting energy expenditure，physical activity and diet in humans[J]. Horm Mol Biol Clin Investig，2017，31（1）：/j/hmbci.2017.31.issue-/j/hmbci.2017.31.issu1/hmbci-2017-0008/hmbci-2017-0008.xml. DOI：10.1515/hmbci-2017-0008.
[10]	EFREMOVA A, COLLELUORI G, THOMSKY M，et al. Biomarkers of browning in cold exposed Siberian adults[J]. Nutrients，2020，12（8）：E2162. DOI：10.3390/nu12082162.
[11]	KROON T, HARMS M, MAURER S，et al. PPARγ and PPARα synergize to induce robust browning of white fat in vivo[J]. Mol Metab，2020，36：100964. DOI：10.1016/j.molmet.2020.02.007.
[12]	RODRíGUEZ A, BECERRIL S, MÉNDEZ-GIMÉNEZ L，et al. Leptin administration activates irisin-induced myogenesis via nitric oxide-dependent mechanisms，but reduces its effect on subcutaneous fat browning in mice[J]. Int J Obes （Lond），2015，39（3）：397-407. DOI：10.1038/ijo.2014.166.
[13]	TABUCHI C, SUL H S. Signaling pathways regulating thermogenesis[J]. Front Endocrinol （Lausanne），2021，12：595020. DOI：10.3389/fendo.2021.595020.
[14]	SRIVASTAVA R K, MOLINER A, LEE E S，et al. CD137 negatively affects "browning" of white adipose tissue during cold exposure[J]. J Biol Chem，2020，295（7）：2034-2042. DOI：10.1074/jbc.AC119.011795.
[15]	GE X, SATHIAKUMAR D, LUA B J，et al. Myostatin signals through miR-34a to regulate Fndc5 expression and browning of white adipocytes[J]. Int J Obes （Lond），2017，41（1）：137-148. DOI：10.1038/ijo.2016.110.
[16]	KIM W K, CHOI H R, PARK S G，et al. Myostatin inhibits brown adipocyte differentiation via regulation of Smad3-mediated β-catenin stabilization[J]. Int J Biochem Cell Biol，2012，44 （2）：327-334. DOI：10.1016/j.biocel.2011.11.004.

组别	只数	体质量（g）	体长（cm）	Lee's指数	白棕比	脂肪指数
WT组	6	27.48±1.16	8.30±0.32	363.84±11.01	3.05±0.09	0.020 7±0.003 8
Mstn（+/-）组	6	29.26±1.55	8.58±0.15	359.19±10.80	2.24±0.12^a	0.017 7±0.004 1
Mstn（-/-）组	6	31.14±1.71^a	9.12±0.18^ab	345.07±12.80^ab	1.75±0.14^ab	0.014 9±0.001 5
WT+DM组	6	22.22±2.29^a	8.36±0.19	335.95±7.70^a	4.33±0.19^a	0.041 2±0.006 5^a
Mstn（+/-）+DM组	6	23.88±1.94	8.76±0.28	334.39±5.31	3.28±0.06^d	0.025 1±0.004 0^d
Mstn（-/-）+DM组	6	24.22±1.70^d	9.28±0.13^d	311.62±4.44^de	2.13±0.15^de	0.016 9±0.005 0^d
F值		19.510	16.621	21.199	259.435	24.230
P值		<0.001	<0.001	<0.001	<0.001	<0.001

组别	只数	体质量（g）	体长（cm）	Lee's指数	白棕比	脂肪指数
WT组	6	27.48±1.16	8.30±0.32	363.84±11.01	3.05±0.09	0.020 7±0.003 8
Mstn（+/-）组	6	29.26±1.55	8.58±0.15	359.19±10.80	2.24±0.12^a	0.017 7±0.004 1
Mstn（-/-）组	6	31.14±1.71^a	9.12±0.18^ab	345.07±12.80^ab	1.75±0.14^ab	0.014 9±0.001 5
WT+DM组	6	22.22±2.29^a	8.36±0.19	335.95±7.70^a	4.33±0.19^a	0.041 2±0.006 5^a
Mstn（+/-）+DM组	6	23.88±1.94	8.76±0.28	334.39±5.31	3.28±0.06^d	0.025 1±0.004 0^d
Mstn（-/-）+DM组	6	24.22±1.70^d	9.28±0.13^d	311.62±4.44^de	2.13±0.15^de	0.016 9±0.005 0^d
F值		19.510	16.621	21.199	259.435	24.230
P值		<0.001	<0.001	<0.001	<0.001	<0.001

组别	只数	TG	TC	LDL-C	HDL-C	FFA
WT组	6	1.17±0.06	2.84±0.23	0.36±0.02	0.66±0.04	0.62±0.01
Mstn（+/-）组	6	1.11±0.03	2.45±0.21^a	0.37±0.02	0.67±0.03	0.62±0.01
Mstn（-/-）组	6	1.02±0.06^ab	2.51±0.05^a	0.43±0.13	0.67±0.01	0.61±0.01
WT+DM组	6	1.62±0.06^a	5.13±0.20^a	0.50±0.03^a	0.38±0.01^a	0.78±0.02^a
Mstn（+/-）+DM组	6	1.44±0.04^d	4.27±0.18^d	0.49±0.02	0.44±0.02^d	0.72±0.01^d
Mstn（-/-）+DM组	6	1.38±0.02^d	3.81±0.18^de	0.43±0.02^d	0.50±0.01^de	0.70±0.01^d
F值		112.663	174.600	31.258	458.253	110.621
P值		<0.001	<0.001	<0.001	<0.001	<0.001

组别	只数	TG	TC	LDL-C	HDL-C	FFA
WT组	6	1.17±0.06	2.84±0.23	0.36±0.02	0.66±0.04	0.62±0.01
Mstn（+/-）组	6	1.11±0.03	2.45±0.21^a	0.37±0.02	0.67±0.03	0.62±0.01
Mstn（-/-）组	6	1.02±0.06^ab	2.51±0.05^a	0.43±0.13	0.67±0.01	0.61±0.01
WT+DM组	6	1.62±0.06^a	5.13±0.20^a	0.50±0.03^a	0.38±0.01^a	0.78±0.02^a
Mstn（+/-）+DM组	6	1.44±0.04^d	4.27±0.18^d	0.49±0.02	0.44±0.02^d	0.72±0.01^d
Mstn（-/-）+DM组	6	1.38±0.02^d	3.81±0.18^de	0.43±0.02^d	0.50±0.01^de	0.70±0.01^d
F值		112.663	174.600	31.258	458.253	110.621
P值		<0.001	<0.001	<0.001	<0.001	<0.001

组别	只数	PPAR-γ	PGC-1α	UCP1	CD137
WT组	6	0.879±0.016	0.705±0.012	0.236±0.020	0.081±0.002
Mstn（+/-）组	6	0.908±0.112	0.711±0.023	0.767±0.069^a	0.111±0.004^a
Mstn（-/-）组	6	1.108±0.026^ab	0.926±0.076^ab	0.800±0.049^a	0.129±0.003^ab
WT+DM组	6	0.170±0.005^a	0.189±0.012^a	0.073±0.001^a	0.025±0.002^a
Mstn（+/-）+DM组	6	0.190±0.010	0.173±0.010	0.095±0.010	0.060±0.002^d
Mstn（-/-）+DM组	6	0.348±0.009^de	0.311±0.008^de	0.165±0.008^de	0.104±0.008^de
F值		446.360	539.281	528.637	538.335
P值		<0.001	<0.001	<0.001	<0.001

Effect of Myostatin Gene Knockout on Browning of White Fat and Related Gene Expression Levels in a Mouse Model of Type 2 Diabetes Mellitus

肌肉生长抑制素基因敲除对2型糖尿病小鼠白色脂肪棕色化及相关基因表达的影响研究

RichHTML

PDF

Knowledge

Abstract

Cite this article

share this article

Figures/Tables 8

References 16

Related Articles 9

Recommended Articles

Metrics

Comments

组别	只数	PPAR-γ	PGC-1α	UCP1	CD137
WT组	6	1.359±0.060	0.929±0.008	0.658±0.038	0.079±0.002
Mstn（+/-）组	6	1.920±0.050^a	1.111±0.050^a	0.809±0.017^a	0.152±0.009^a
Mstn（-/-）组	6	2.052±0.040^ab	1.556±0.059^ab	0.830±0.030^a	0.273±0.016^ab
WT+DM组	6	0.301±0.014^a	0.187±0.011^a	0.092±0.007^a	0.062±0.003^a
Mstn（+/-）+DM组	6	0.455±0.027^d	0.251±0.020^d	0.129±0.006^d	0.130±0.006^d
Mstn（-/-）+DM组	6	1.218±0.085^de	0.645±0.014^de	0.288±0.010^de	0.168±0.004^de
F值		1 196.913	1 463.654	1 493.055	490.418
P值		<0.001	<0.001	<0.001	<0.001

[1]	Keng CHEN, Yiyao DENG, Shunlai SHANG, Qinggang LI, Xiangmei CHEN. Chinese Herbal Formula Shenyi Improves Kidney Injury and Inhibits the Activation of the Alternative Complement Pathway in a Mouse Model of Lupus Nephritis [J]. Chinese General Practice, 2022, 25(26): 3298-3307.
[2]	ZHANG Yushuang, YU Fuyang, WU Zhongbing, WANG Yiran, LI Jing. Analysis of Gut Flora in a Mouse Model of Esophageal Squamous Cell Carcinoma in Situ [J]. Chinese General Practice, 2022, 25(08): 945-951.
[3]	CHEN Shaohua，WANG Chinyun，WU Changgui，ZHAO Xiao，XU Hao，SHI Qi，LIANG Qianqian. Efficacy and Mechanism of Action of Danggui Niantong Decoction against Acute Gouty Arthritis in a Mouse Model：an Experimental Study　 [J]. Chinese General Practice, 2021, 24(24): 3116-3121.
[4]	YU Lei，YANG Yunshuang，LI Yanhui，LIU Xiaochen，DU Lei，WANG Sinian，ZHANG Rong. Protective Effect of Bushen Jiedu Prescription on Radiation-induced Thymus Injury in Mice　 [J]. Chinese General Practice, 2020, 23(9): 1164-1168.
[5]	CHANG Quan，ZHANG Nana，MA Dong. Bioinformatics Analysis of Myelin KLF5 Deficiency Inhibiting Abdominal Aortic Aneurysm Formation in Mice　 [J]. Chinese General Practice, 2020, 23(6): 692-698.
[6]	ZHANG Yan*，FAN Xiaoxiang，ZHANG Meiwu，ZHUANG Luhui，MAO Dafeng. Role of NLRC5 in the Activation of Hepatic Stellate Cells Induced by TGF-β1 and Reversal of Hepatic Fibrosis [J]. Chinese General Practice, 2020, 23(24): 3051-3059.
[7]	LI Hongtao，LIN Yusen，ZOU Xiaoling，LI Wenjuan，MENG Ping，ZHANG Tiantuo. Effects of CpG Oligodeoxynucleotides on Airway Remodeling in Combined Allergic Rhinitis and Asthma Syndrome　 [J]. Chinese General Practice, 2019, 22(26): 3191-3197.
[8]	XU Li，GE Qiongxiang，LIN Guoqiang. Effect of Mild Moxibustion on TRPV1 Expression in the Dorsal Root Ganglion in a Mouse Model of Pruritus Ani　 [J]. Chinese General Practice, 2018, 21(30): 3749-3752.
[9]	ZHANG Na1LIU Xuefang，JIANG Shihong，WANG Qi，TIAN Yange. Yifeizhuji Decoction Combined with Cyclophosphamide for Reducing the Tumor Volume and Improving the Immunologic Functions in the Lewis Lung Carcinoma Mouse Model　 [J]. Chinese General Practice, 2018, 21(30): 3753-3756.