Journal of Animal Science and Technology

Korean Society of Animal Sciences and Technology (한국축산학회)
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Gene Expression of Candidate Genes Involved in Fat Metabolism During In vitro Adipogenic Differentiation of Bovine Mesenchymal Stem Cell

Bovine Mesenchymal Stem Cell의 지방분화를 이용한 지방대사관련 후보 유전자의 발현분석

  • Kim, Sung-Kon (Animal Genomics & Bioinformatics Division, National Institute of Animal Science, Rural Development Administration) ;
  • Kim, Nam-Kuk (Animal Genomics & Bioinformatics Division, National Institute of Animal Science, Rural Development Administration) ;
  • Yoon, Du-Hak (Animal Genomics & Bioinformatics Division, National Institute of Animal Science, Rural Development Administration) ;
  • Kim, Tae-Hun (Animal Genomics & Bioinformatics Division, National Institute of Animal Science, Rural Development Administration) ;
  • Yang, Boo-Keun (Dept. of Animal Resources Science, College of Animal Life Science, Kangwon National University) ;
  • Lee, Hyun-Jeong (Animal Genomics & Bioinformatics Division, National Institute of Animal Science, Rural Development Administration)
  • 김성곤 (농촌진흥청 국립축산과학원) ;
  • 김남국 (농촌진흥청 국립축산과학원) ;
  • 윤두학 (농촌진흥청 국립축산과학원) ;
  • 김태헌 (농촌진흥청 국립축산과학원) ;
  • 양부근 (강원대학교 동물생명과학대학 동물자원학과) ;
  • 이현정 (농촌진흥청 국립축산과학원)
  • Received : 2010.03.24
  • Accepted : 2010.08.04
  • Published : 2010.08.31
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Abstract

Adipogenesis has been one of the most intensely studied models of cellular differentiation. During adipogenesis, differential expression of many adipogenesis related genes lead to profound changes in cellular, morphological, and physiological characteristics of the differentiating cells. The aim of the present study was to examine the expression levels of adipogenic candidate genes, cAMP early repressor (ICER), nephroblastoma over-expressed protein (NOV), heat shock protein beta 1 (HSPB1) and succinate dehydrogenase (SDH), during adipogenesis of bovine mesenchymal stem cells (BMSC). The BMSC were cultured in DMEM / low glucose medium with adipogenic inducers for 6 days and the expression of various candidate genes which seemed related to adipogenesis were measured by real-time PCR. This study showed that the expression of peroxisome proliferator activated receptor ${\gamma}$(PPAR${\gamma}$) and fatty acid binding protein 4 (FABP4) genes as adipogenic indicators were increased to 3.11 and 3.11 folds on day 6 than on day 0, respectively (p<0.05). To determine whether candidate genes were related to adipogenesis, the expression levels of ICER, NOV, HSPB1, and SDH genes were measured during adipogenesis in BMSC. Our results showed that the expression level of ICER gene was significantly increased to 4.12 folds (0.01729 vs. 0.07138; p<0.05), whereas NOV, HSPB1, and SDH genes were decreased to 2.89, 3.18 and 2.36 folds, respectively, on day 6 when compared to day 0. These results suggest that these candidate genes have stimulatory or inhibitory effects on adipogenesis in BMSC, indicating that these genes may be directly or indirectly related to the adipogenic event of adipose precursor cells.

본 실험은 지방대사에 영향을 주는 후보 유전자의 발현양상을 조사하기 위하여 bovine mesenchymal stem cell을 이용하여 체외에서 지방분화를 유도하였고, 지방분화 경과에 따른 지방관련 후보유전자의 발현양상을 분석 하였다. Oil red-O 염색법을 이용하여 지방 분화양상을 조사한 결과, 지방 분화 단계별(0, 2, 4 및 6일)로 붉게 염색된 지방적이 증가되었음을 확인하였고, 지방대사의 indicator인 PPAR$\gamma$와 FABP4 유전자는 지방분화 0일과 비교하여 6일에는 각각 3.11배와 3.11배가 증가하였다. 또한 지방대사와 관련이 있다고 추측되는 ICER, NOV, HSPB1 및 SDH 유전자의 발현양상을 조사한 결과, 지방분화 동안 ICER 유전자는 계속적으로 증가하여 6일은 0일에 비해 4.12배의 유의적인 증가를 보인 반면, NOV, HSPB1 및 SDH 유전자의 발현양은 0일 이후 계속적으로 감소하여, 6일에서 각각 2.89배, 3.18배 및 2.36배의 감소를 나타냈다. 본 실험에서, 등심조직에서 근내지방도에서 차이를 나타냈던 ICER, NOV, HSPB1 및 SDH 유전자를 보다 구체적이고 직접적으로 세포에서 지방분화를 통해 연관성을 조사했을 때, 지방분화와 직접적으로 관련되어 있는 것으로 나타났으며, 지방 대사를 조절하는 유전자로 추정된다.

Keywords

References

  1. Benini, S., Perbal, B., Zambelli, D., Colombo, M. P., Manara, M. C., Serra, M., Parenza, M., Martinez, V., Picci, P. and Scotlandi, K. 2005. In Ewing’s sarcoma CCN3 (NOV) inhibits proliferation while promoting migration and invasion of the same cell type. Oncogene 24:4349-4361. https://doi.org/10.1038/sj.onc.1208620
  2. Bosnakovski, D., Mizuno, M., Kim, K. H., Taiagi, S., Okumura, M. and Fujinaga, T. 2005. Isolation and multilineage differentiation of bovine bone marrow mesenchymal stem cells. Cell Tissue Res. 319:243-253. https://doi.org/10.1007/s00441-004-1012-5
  3. Bronnikov, G., Houstek, J. and Nedergaard, J. 1992. ${\beta}$-Adrenergic, cAMP-mediated stimulation of proliferation of brown fat cells in primary culture. Mediation via ${\beta}1$ but not via ${\beta}3$ receptors. J. Biol. Chem. 267:2006-2013.
  4. Frohnert, B. I., Hui, T. Y. and Bernlohr, D. A. 1999. Identification of a functional peroxisome proliferator-responsive element in the murine fatty acid transport protein gene. J. Biol. Chem. 274:3970-3977. https://doi.org/10.1074/jbc.274.7.3970
  5. Garrido, C., Brucy, J. M., Fromentin, A., Hammann, A., Arrigo, A. P. and Solary, E. 1999. HSP27 inhibits cytochrome c-dependent activation of procaspase-9. FASEB J. 13:2061-2070. https://doi.org/10.1096/fasebj.13.14.2061
  6. Guo, X. and Liao, K. 2000. Analysis of gene expression profile during 3T3-L1 preadipocyte differentiation. Gene 251:45-53. https://doi.org/10.1016/S0378-1119(00)00192-X
  7. Hwa, V., Oh, Y. and Rosenfeld, R. G. 1999. The insulin-like growth factor-binding protein (IGFBP) superfamily. Endocr. Rev. 20:761-787. https://doi.org/10.1210/er.20.6.761
  8. Kazala, E. C., Petrak, J. L., Lozeman, F. J., Mir, P. S., Laroche, A., Deng, J. and Weselake, R. J. 2003. Hormone-sensitive lipase activity in relation to fat content of muscle in Wagyu hybrid cattle. Livest. Prod. Sci. 79:87-96. https://doi.org/10.1016/S0301-6226(02)00141-0
  9. Kim, H. S., Nagalla, S. R., Oh, Y., Wilson, E., Roberts, C. T. and Jr Rosenfeld, R. G. 1997. Identification of a family of low-affinity insulin-like growth factor binding proteins (IGFBPs): characterization of connective tissue growth factor as a member of the IGFBP superfamily. Proc. Nat. Acad. Sci. 94:12981-12986. https://doi.org/10.1073/pnas.94.24.12981
  10. Kim, N. K., Lee, S. H., Cho, Y. M., Son, E. S., Kim, K. Y., Lee, C. S., Yoon, D. H., Im, S. K., Oh, S. J. and Park, E. W. 2009. Proteome analysis of the m. Longissimus dorsi between fattening stages in Hanwoo steer. BMB reports 42:433-438. https://doi.org/10.5483/BMBRep.2009.42.7.433
  11. Lee, S. H., Park, E. W., Cho, Y. M., Kim, S. K., Lee, Jun. H., Jeon, J. T., Lee, C. S., Im, S. K., Oh, S. J., Thompson, J. M. and Yoon, D. H. 2007. Identification of Differentiatially Genes Related to Intramuscular Fat Development in the Early and Late Fattening Stages of Hanwoo Steers. J. of Biochem. and Mol. Biol. 40:757-764. https://doi.org/10.5483/BMBRep.2007.40.5.757
  12. Pandey, P., Farber, R., Nakazawa, A., Kumar, S., Bharti, A., Nalin, C., Weichselbaum, R., Kufe, D. and Kharbanda, S. 2000. Hsp27 functions as a negative regulator of cytochrome c-dependent activation of procaspase-3 Oncogene 19:1975-1981. https://doi.org/10.1038/sj.onc.1203531
  13. Pethick, D. W., D'Souza, D. N., Dunshea, F. R and Harper, G. S. 2005. Fat metabolism and regional distribution in ruminants and pig-influences of genetics and nutrition. Rec. Adv. Anim. Nutr. 15:39-45.
  14. Resen, E. D. 2005. The transcriptional basis of adipocyte development. Prostag. Leukotr. Ess. 73:31-34. https://doi.org/10.1016/j.plefa.2005.04.004
  15. Scheffler, I. E. 1998. Molecular genetics of succinate; quinine oxidoreductase in eukaryotes. Prog. Nucleic Acid. Res. Mol. Biol. 60:267-315. https://doi.org/10.1016/S0079-6603(08)60895-8
  16. Snaith, M. R., Natarajan, D., Taylor, L. B., Choi, C. P., Martinerie, C., Perbal, B., Schofield, P. N. and Boulter, C. A. 1996. Genomic structure and chromosomal mapping of the mouse nov gene. Genomics 38:425-428. https://doi.org/10.1006/geno.1996.0647
  17. Soret, J., Dambrine, G. and Perbla, B. 1989. Induction of nephroblastoma by myeloblastosis-associated virus type 1: state of proviral DNAs in tumor cells. J. Virol. 63:1803-1807.
  18. Tanabe, S., Sato, Y., Suzuki, T., Suzuki, K., Nagao, T. and Yanaguchi, T. 2008. Gene expression profiling of human mesenchymal stem cells for differentiation of novel markers in early- and late-stage cell culture. J. Biochem. 144:399-408. https://doi.org/10.1093/jb/mvn082
  19. Wolfrum, C., Ellinghaus, P., Fobker, M., Seedorf, U., Assmann, G., Borchers, T. and Spener, F. 1999. Phytanic acid is ligand and transcriptional activator of murine liver fatty acid binding protein. J. Lipid Res. 40:708-714.
  20. Yagi, K., Kondo, D., Okazaki, Y. and Kano, K. 2004. A novel preadipocyte cell line established from mouse adult mature adipocytes. Biochem. Biophy. Res. Commun. 321:967-74. https://doi.org/10.1016/j.bbrc.2004.07.055
  21. Zhou, Y. P., Marlen, K., Palma, J. F., Schweitzer, A., Reilly, L., Gregoire, F. M., Xu, G. G., Blume, J. E. and Honson, J. D. 2003. Overexpression of repressive cAMP response element modulators in high glucose and fatty acid-treated rat islets. A common mechanism for glucose toxicity and lipotoxicity?. J. Biol. Chem. 278:51316-51323. https://doi.org/10.1074/jbc.M307972200
  22. 김남국, 김성곤, 허강녕, 윤두학, 이창수, 임석기, 박응우. 2008. 한우 비육기간 중 중성지방 생합성 관련 유전자의 발현양상. 한국동물자원과학회지. 50:293-300. https://doi.org/10.5187/JAST.2008.50.3.293
  23. 서종배, 이윤석, 김재범. 2004. 지방조직과 지방세포 분화에 대한 연구. Bio-Medical Science. 2:17-46.
  24. 이승환, 박응우, 조용민, 김경훈, 오영균, 이지혜, 이창수, 오성종, 윤두학. 2006. 한우 비육 전.후기의 등심조직에 있어서 지방합성 유전자 발현. 한국동물자원과학회지. 48:345-352. https://doi.org/10.5187/JAST.2006.48.3.345
  25. 임희경, 최강덕, Baatartsogt Oyungerel, 최영숙, 정정수. 2008. Retinoic Acid가 돼지 지방전구세포의 분화와 유전자 발현에 미치는 영향. 한국동물자원과학회지. 50:475-484. https://doi.org/10.5187/JAST.2008.50.4.475
  26. 정유성, 김남국, 김건석, 김범수, 문희주, 김남영, 허강녕, 이승환, 박응우, 임석기. 2008. 한우 성장단계에 따른 한우 등심조직 내 경제형질관련 유전자의 발현양상 분석. 한국동물자원학회 춘계학술발표회 초록. NO.PA28087.

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