慢病毒介导shRNA干扰RBP4促进猪前体脂肪细胞成脂分化

doi:10.11843/j.issn.0366-6964.2013.06.016

畜牧兽医学报 ›› 2013, Vol. 44 ›› Issue (6): 937-943.doi: 10.11843/j.issn.0366-6964.2013.06.016

慢病毒介导shRNA干扰RBP4促进猪前体脂肪细胞成脂分化

程佳，蒲蕾，杨浩，史新娥，郑加盟，张振宇，杨公社^*

（西北农林科技大学动物科技学院动物脂肪沉积与肌肉发育实验室，杨凌 712100）

收稿日期:2012-11-16 出版日期:2013-06-23 发布日期:2013-06-23
通讯作者: 杨公社，教授，Tel:029-87092430，E-mail: gsyang999@hotmail.com
作者简介:程佳（1983-），女，内蒙古呼和浩特人，博士，主要从事动物生物技术研究，E-mail: chengjia@nwsuaf.edu.cn，Tel：029-87091017
基金资助:
国家973计划项目(2012CB124705)；高繁殖力转基因猪新品种培育(2011ZX08006-005)；西北农林科技大学“创新团队建设计划”项目资助

Lentivirus Mediated shRNA Interference Silencing RBP4 Promoted Differentiation of Porcine Preadipocytes

CHENG Jia, PU Lei, YANG Hao, SHI Xin-e, ZHENG Jia-meng, ZHANG Zhen-yu,YANG Gong-she^*

（Laboratory of Animal Fat Deposition and Muscle Development, College of Animal Science and Technology, Northwest A&F University, Yangling 712100, China)

Received:2012-11-16 Online:2013-06-23 Published:2013-06-23

摘要/Abstract

摘要：

为研究视黄醇结合蛋白4(retinol binding protein 4, RBP4)对猪前体脂肪细胞分化的影响，笔者构建了RBP4重组慢病毒干扰载体，包装并感染猪前体脂肪细胞，采用BODIPY和油红O染色以及real-time PCR等方法，从形态学和基因表达水平检测抑制RBP4表达对成脂分化的作用。研究结果显示，RBP4重组慢病毒干扰载体构建成功，病毒滴度达到 6.5×10⁷pfu·mL^-1，能显著抑制猪前体脂肪细胞中RBP4的表达水平（干扰效率在60%以上）。同时，干扰RBP4能促进猪前体脂肪细胞分化，增加成脂关键基因PPARγ、SREBP-1c和aP2的mRNA表达。本研究结果表明，RBP4对猪前体脂肪细胞分化有抑制作用，为进一步研究RBP4对猪前体脂肪细胞分化的作用机制奠定基础。

Abstract:

To investigate the role of RBP4 during the differentiation of porcine preadipocytes and the biological function of RBP4, we constructed porcine RBP4 lentivirus interfering vector, then infected the porcine preadipocytes. The changes in morphology and gene expression levels were studied to evaluate the effects of RBP4 expression on preadipocyte differentiation by BODIPY, Oil Red O staining and real-time PCR analysis. The results showed that RBP4 lentivirus interfering vector was constructed correctly, virus titer was 6.5×10⁷ pfu·mL^-1. RBP4 mRNA and protein expression was remarkably reduced by approximate 60% in porcine preadipocytes infected with shRBP4. Moreover, preadipocytes differentiation was promoted and the mRNA expression levels of PPARγ, SREBP-1c and aP2 were increased in adipocytes by knocking down of RBP4. Therefore, adipogenic differentiation of porcine preadipocytes are inhibited by RBP4, which would help the further exploration of the biological mechanism of RBP4 during the adipogenesis.

中图分类号:

S813.3
Q782

程佳，蒲蕾，杨浩，史新娥，郑加盟，张振宇，杨公社. 慢病毒介导shRNA干扰RBP4促进猪前体脂肪细胞成脂分化[J]. 畜牧兽医学报, 2013, 44(6): 937-943.

CHENG Jia, PU Lei, YANG Hao, SHI Xin-e, ZHENG Jia-meng, ZHANG Zhen-yu,YANG Gong-she. Lentivirus Mediated shRNA Interference Silencing RBP4 Promoted Differentiation of Porcine Preadipocytes[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2013, 44(6): 937-943.

0
/ / 推荐

导出引用管理器 EndNote|Reference Manager|ProCite|BibTeX|RefWorks

链接本文: https://www.xmsyxb.com/CN/10.11843/j.issn.0366-6964.2013.06.016

https://www.xmsyxb.com/CN/Y2013/V44/I6/937

参考文献

YANG Q, GRAHAM T E, MODY N, et al. Serum retinol binding protein 4 contributes to insulin resistance in obesity and type 2 diabetes［J］. Nature, 2005, 436(7049):356-362.

BROCH M, GOMEZ J M, AUGUET M T, et al. Association of retinol-binding protein-4 (RBP4) with lipid parameters in obese women［J］.Obes Surg, 2010, 20(9):1258-1264.

GRAHAM T E, YANG Q, BLÜHER M, et al. Retinol binding protein 4 and insulin resistance in lean, obese and diabetic subjects［J］. N Engl J Med, 2006, 354(24):2552-2563.

YAO-BORENGASSER A, VARMA V, BODLES A M, et al. Retinol binding protein 4 expression in humans relationship to insulin resistance, inflammation, and response to pioglitazone［J］. J Clin Endocrinol Metab, 2007, 92(7):2590-2597.

BAJZOVÁ M, KOVÁCIKOVÁ M, VÍTKOVÁ M, et al. Retinol-binding protein 4 expression in visceral and subcutaneous fat in human obesity［J］. Physiol Res, 2008, 57(6):927-934.

STEFAN N, HENNIGE A M, STAIGER H, et al. High circulating retinol-binding protein 4 is associated with elevated liver fat but not with total, subcutaneous, visceral, or intramyocellular fat in humans［J］. Diabetes Care, 2007, 30(5):1173-1178.

MUNKHTULGA L, NAGASHIMA S, NAKAYAMA K, et al. Regulatory SNP in the RBP4 gene modified the expression in adipocytes and associated with BMI［J］. Obesity (Silver Spring), 2010, 18(5):1006-1014.

LI X J, YANG H, LI G X, et al. Transcriptome profile analysis of porcine adipose tissue by high-throughput sequencing［J］. Anim Genet, 2012, 43(2):144-152.

张国华, 杨公社, 屈长青, 等. 猪前体脂肪细胞的分离培养［J］.细胞生物学杂志, 2005, 27(6): 693-696.

POULOS S P, HAUSMAN D B, HAUSMAN G J. The development and endocrine functions of adipose tissue［J］. Mol Cell Endocrinol, 2010, 323(1): 20-34.

HAJER G R, VAN HAEFTEN T W, VISSEREN F L. Adipose tissue dysfunction in obesity, diabetes, and vascular diseases［J］. Eur Heart J, 2008, 29(24): 2959-2971.

ANTUNA-PUENTE B, FEVE B, FELLAHI S, et al. Adipokines: the missing link between insulin resistance and obesity［J］. Diabetes Metab, 2008, 34(1):2-11.

KOTNIK P, FISCHER-POSOVSZKY P, WABITSCH M. RBP4: a controversial adipokine［J］. Eur J Endocrino, 2011, 165(5):703-711.

KLÖTING N, GRAHAM T E, BERNDT J, et al. Serum retinol-binding protein is more highly expressed in visceral than subcutaneous adipose tissue and is a marker of intra-abdominal fat mass［J］. Cell Metab, 2007, 6(1):79-87.

GIRY L M, VERHOEYEN E, SALMON P. Lentiviral vectors［J］. Methods Mol Biol, 2011,737: 183- 209.

COCKRELL A S, KAFRI T. Gene delivery by lentivirus vectors［J］. Mol Biotechnol, 2007, 36(3): 184-204.

王平, 熊燕, 杨公社, 等. 慢病毒载体介导的RNA干扰敲减Akt2表达抑制猪前体脂肪细胞分化［J］.中国生物化学与分子生物学报, 2012, 28(1):53-60.

LOWE C E, O＇RAHILLY S, ROCHFORD J J．Adipogenesis at a glance［J］．J Cell Sci, 2011, 124(16): 2681-2686.

DALL'OLIO S, FONTANESI L, TOGNAZZI L, et al. Genetic structure of candidate genes for litter size in Italian Large White pigs［J］. Vet Res Commun, 2010, 34 (Suppl 1):S203-S206.

刘远, 李良良, 莫小雨, 等. PRLR、RBP4基因多态性分布及其对大白猪产仔数的影响［J］. 中国畜牧兽医, 2010, 37(7):120-123.

慢病毒介导shRNA干扰RBP4促进猪前体脂肪细胞成脂分化

Lentivirus Mediated shRNA Interference Silencing RBP4 Promoted Differentiation of Porcine Preadipocytes

PDF

可视化

被引次数

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 0

编辑推荐

Metrics

本文评价