Source: WASHINGTON STATE UNIVERSITY submitted to NRP
DECIPHERING MECHANISMS REGULATING INTRAMUSCULAR ADIPOGENESIS AND FIBROGENESIS IN BEEF CATTLE
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
0228757
Grant No.
(N/A)
Cumulative Award Amt.
(N/A)
Proposal No.
(N/A)
Multistate No.
(N/A)
Project Start Date
Mar 1, 2012
Project End Date
Feb 28, 2017
Grant Year
(N/A)
Program Code
[(N/A)]- (N/A)
Recipient Organization
WASHINGTON STATE UNIVERSITY
240 FRENCH ADMINISTRATION BLDG
PULLMAN,WA 99164-0001
Performing Department
Animal Science
Non Technical Summary
According to the past surveys of beef producers by National Cattlemen's Beef Association, marbling and tenderness were consistently identified as the top beef quality problems. Marbling becomes a top quality problem because of the selection for high lean growth. Fat is energy intensive but has low commercial value. Excessive fat accumulation in livestock leads to huge waste of feeds. Thus, enhancing lean growth while reducing fat accumulation dramatically improves the efficiency of animal production. Hence, for generations, animals are selected for their high lean/fat ratio; such selection results in overall reduction in fat accumulation, including intramuscular fat (marbling) which is critical for the palatability of meat. To improve the eating quality of meat, we have to increase beef marbling. Beef tenderness is mainly determined by two factors: myofibrils (the collection of contractile proteins within skeletal muscle fibers) and connective tissue. Myofibrillar effect on beef tenderness can be largely solved by proper aging of carcasses and stretching muscle fibers, however, the connective tissue which is responsible for background toughness is not. Connective tissue and its cross-linking resist protease hydrolysis and thus postmortem aging is not effective in tenderizing beef with high connective tissue content. Therefore, to improve tenderness, we need to reduce connective tissue content and its cross-linking in beef. The objectives of proposed studies are to elucidate mechanisms regulating the formation fat cells and fibronic cells. These molecular mechanisms identified will provide molecular targets for genetic selection or nutritional management to enhance formation of intramuscular fat cells and reduce connective tissue in beef cattle, improving beef quality. Enhancing knowledge of fat cell formation in beef cattle will also allow us to develop strategies to reduce excessive fat accumulation under skin and inside abdomen, elevating the production efficiency of beef cattle.
Animal Health Component
(N/A)
Research Effort Categories
Basic
(N/A)
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
3053310103050%
3083320103050%
Knowledge Area
305 - Animal Physiological Processes; 308 - Improved Animal Products (Before Harvest);

Subject Of Investigation
3320 - Meat, beef cattle; 3310 - Beef cattle, live animal;

Field Of Science
1030 - Cellular biology;
Goals / Objectives
According to the past surveys of beef producers by National Cattlemen's Beef Association, marbling and tenderness were consistently identified as the top beef quality problems. Skeletal muscle development mainly involves the formation of muscle fibers (myogenesis), but also the formation of intramuscular adipocytes (adipogenesis) and fibroblasts (fibrogenesis). Intramuscular adipogenesis forms sites for fat deposition and formation of marbling fat. Intramuscular fibrogenesis generates connective tissue responsible for the background toughness of meat. Very recent studies show that intramuscular adipocytes and fibroblasts are developed from common progenitor cells. Thus, enhancing adipogenic and inhibiting fibrogenic differentiation from progenitor cells will increase marbling and tenderness of meat, which necessitates our understanding of mechanisms regulating adipogenic and fibrogenic differentiation of intramuscular progenitor cells. We hypothesize that the lineage commitment of adipogenic/fibrogenic progenitor cells is regulated by the expression of a few critical transcription factors initiating adipogenesis or fibrogenesis, which leads to permanent epigenetic changes in these cells. To test our hypothesis, we will pursue two specific aims: 1. To identify epigenetic modifications committing progenitor cells to either adipogenic or fibrogenic lineage. Intramuscular progenitor cells separated from the same beef animal will be selected for their high and low potential for adipogenesis and fibrogenesis. The difference in epigenetic modifications of key genes will be compared. The working hypothesis here is that epigenetic modifications in the promoter of a few key transcription factors render progenitor cells committing to different lineages. 2. To test genomic DNA changes rendering progenitor cells with differential potential for adipogenic and fibrogenic commitment. Intramuscular progenitor cells will be separated from Angus and Wagyu cattle. The difference in adipogenesis and fibrogenesis will be compared and correlated with the expression of key transcription factors regulating lineage commitment. The working hypothesis here is that genetic changes in Wagyu cattle alter the expression of key transcription factors to favor adipogenic differentiation of progenitor cells. To date, our understanding of mechanisms regulating the lineage commitment of progenitor cells to either adipogenesis or fibrogenesis is very limited. In this hatch project, we will use two different approaches: 1) progenitor cells with the same genetic background but different epigenetic modifications, and 2) progenitor cells with different genetic background. The overall objective is to identify key transcription factors leading to the commitment of progenitor cells to either adipogenic or fibrogenic differentiation.
Project Methods
Aim 1. To identify epigenetic modifications committing progenitor cells to either adipogenic or fibrogenic lineage. Stromal vascular cells will be collected using our well established method, and progenitor cells will be separated. Isolated progenitor cells will be seeded at low density so that each primary cell can form a clone to obtain clonally derived cells. These cloned cells will be used to test their ability for adipogenesis. Three clones will high adipogenic capacity and low capacity per cattle will be selected for further experiments. Because these cells are separated from a single animal with identical genetic background, the difference in lineage commitment is solely due to epigenetic modifications. The difference in epigenetic modifications of key genes will be compared. The working hypothesis here is that epigenetic modifications in the promoters of a few key transcription factors render progenitor cells committing to different lineages. Aim 2. To test genomic DNA changes rendering progenitor cells with differential potential for adipogenic and fibrogenic commitment. Japanese Wagyu cattle have extremely high intramuscular fat (marbling). It is likely that such high marbling is due to genetic mutations which render adipogenic/fibrogenic progenitor cells with very high adipogenic capacity. As described for Aim 1 studies, intramuscular progenitor cells will be separated from Angus and Wagyu cattle. Both Japanese Wagyu and Angus cattle are available at WSU and the Meat Lab frequently slaughters. Therefore, we can obtain Wagyu and Angus muscle samples on a year-round basis. The difference in adipogenesis and fibrogenesis of progenitor cells will be compared and correlated with the expression of key transcription factors regulating lineage commitment. The working hypothesis here is that genetic changes in Wagyu cattle alter the expression of key transcription factors which enhance adipogenic differentiation of progenitor cells.

Progress 03/01/12 to 02/28/17

Outputs
Target Audience:Scientific community, and beef and other livestock producers Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Xing Fu, Corrine Harrisand Natasha Moffitt How have the results been disseminated to communities of interest?Journal articles and presentations in scientific conferences What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? We have been comparing the difference in adipogenic differentiation between Wagyu and Angus cattle and found that both adipogenic and fibrogenic differentiation in Wagyu cattle are enhanced but myogenic differentiation is attenuated in comparison with Angus cattle.

Publications

  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Du, M., S.P. Ford, and M. J. Zhu. (2017). Optimizing livestock production efficiency through maternal nutritional management and fetal developmental programming. Animal Frontiers, 7: 5-11.
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2017 Citation: Fu, X., Q. Yang, B. Wang, J. Zhao, M. Zhu, S. M. Parish, and M. Du. (2017). Reduced satellite cell density and myogenesis in Wagyu compared to Angus cattle as a possible explanation of its high marbling. Animal, In press.


Progress 10/01/15 to 09/30/16

Outputs
Target Audience:Scientific community, and beef and other livestock producers. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Corrine Harris, Bo Wang, Natasha Moffitt, and Xing Fu How have the results been disseminated to communities of interest?Journal articles and presentations in scientific conferences What do you plan to do during the next reporting period to accomplish the goals?We will continue to define factors regulating the divergence of myogenic and fibro/adipogenic lineages during early embryonic development, and the fibrogenic and adipogenic differentiation of progenitor cells during fetal development.

Impacts
What was accomplished under these goals? We are defining the mechanisms regulating adipogenic and fibrogenic differentiation of intramuscular progenitor cells. Our data show that adipogenic and fibrogenic processes are competitive, with strengthening adipogenesis inhibiting fibrogenesis and collagen deposition. We further found that Wagyu cattle have both enhanced adipogenesis and fibrogenesis, but impaired myogenesis, suggesting the strengthening of fibro/adipogenic lineage commitment is linked to the high marbling fat development in Wagyu cattle. These data support the notion that early divergence of myogenic and fibro/adipogenic lineages occurs during bovine cattle development.

Publications

  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Liang, X. W., Q. Y. Yang, L. P. Zhang, J. Maricelli, B. D. Rodgers, M. J. Zhu, and M. Du (2016). Maternal high-fat diet during lactation impairs thermogenic function of brown adipose tissue in offspring mice. Scientific Reports, 6: 34345.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Yang, Q., X. Liang, X. Sun, L. Zhang, X. Fu, C. J. Rogers, A. Berim, S. Zhang, S. Wang, B. Wang, M. Foretz, B. Viollet, D. R. Gang, B. D. Rodgers, M. Zhu, and M. Du. (2016). AMPK/a-ketoglutarate axis dynamically mediates DNA demethylation in the Prdm16 promoter and brown adipogenesis. Cell Metabolism, 24: 542-554.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Wang, B., Q. Yang, C. L. Harris, M. L. Nelson, J. R. Busboom, M. J. Zhu, and M. Du. (2016). Nutrigenomic regulation of adipose tissue development  role of retinoic acid. Meat Science, 120: 100-106.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Miao, Z. G., L. P. Zhang, X. Fu, Q. Y. Yang, M. J. Zhu, M. V. Dodson, and M. Du. (2016). Invited review: Mesenchymal progenitor cells in intramuscular connective tissue development. Animal, 10: 75-81.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Fu, X., M. J. Zhu, S. Zhang, F. Marc, V. Benoit, and M. Du. (2016). Obesity impairs skeletal muscle regeneration via inhibition of AMP-activated protein kinase. Diabetes. 65: 188-200.


Progress 10/01/14 to 09/30/15

Outputs
Target Audience:Scientific community, beef producers Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Ph.D. students: Xing Fu and Shenjuan Wei have finished their Ph.D. training. How have the results been disseminated to communities of interest?Through peer-reviewed publications, meetings and shortcourse presentations, knowledge is being disseminated to scientific community and beef producers. What do you plan to do during the next reporting period to accomplish the goals?We will continue to define mechanisms regulating adipogenic and fibrogenic differentiation of progenitor cells in beef cattle.

Impacts
What was accomplished under these goals? We continue to define mechanisms regulating intramuscular adipogenic and fibrogenic differentiation of progenitor cells. Using Wagyu cattle as a unique model for studying adipogenesis, we found that zfp423 has a key role in the initiation of adipogenic differentiation of progenitor cells in beef cattle. Currently, we are defining mechanisms regulating zfp423 expression in progenitor cells.

Publications

  • Type: Journal Articles Status: Accepted Year Published: 2016 Citation: Campos, C. F., M. S. Duarte, S. E. F. Guimaraes, L. L. Verardo, S. Wei, M. Du, Z. Jiang, W. G. Bergen, G. J. Hausman, M. Fernyhough-Culver, E. Albrecht, and M. V. Dodson. (2016). Review: Animal model and the current understanding of molecular dynamics of adipogenesis. Animal, In press.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Miao, Z. G., L. P. Zhang, X. Fu, Q. Y. Yang, M. J. Zhu, M. V. Dodson, and M. Du. (2016). Invited review: Mesenchymal progenitor cells in intramuscular connective tissue development. Animal, 10: 75-81.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Martins TS, ML Chizzotti, W Silva, LN Renno, LMP Sanglard, NVL Serao, FF Silva, SEF Guimaraes, MM Ladeira, MV Dodson, M Du, and MS Duarte. (2015). Molecular factors underlying the deposition of intramuscular fat and collagen in skeletal muscle of Nellore and Angus cattle. PLOS One, 0139943.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Wei, S., X. Fu, X. Liang, M. Zhu, Z. Jiang, S. M. Parish, M. V. Dodson, L. Zan, and M. Du. (2015). Enhanced mitogenesis in stromal vascular cells derived from subcutaneous adipose tissue of Wagyu compared with those of Angus cattle. Journal of Animal Science, 93: 1015-1024.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Du, M., W. Bo, M.J. Zhu. (2015). Fetal programming in meat production. Meat Science, 109: 40-47.


Progress 10/01/13 to 09/30/14

Outputs
Target Audience: Scientists and beef cattle producers Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Shengjuan Wei, Ph.D. student; Xing Fu, Ph.D. student; Carl Rogers, Ph.D. student. How have the results been disseminated to communities of interest? Peer-reviewed publications; abstracts and presentations in professional conferences and meetings. What do you plan to do during the next reporting period to accomplish the goals? We will be further defining mechanisms regulating intramuscular adipogenesis in beef cattle, focused on the expression of Zfp423 and PPARG, two transcription factors critical to the early and late stages of adipogenesis.

Impacts
What was accomplished under these goals? Japanese Wagyu cattle are well known for their extremely high marbling and lower subcutaneous adipose tissue compared with Angus cattle. However, mechanisms responsible for differences in adipose deposition are unknown. During the past year, we continued to study mechanisms regulating intramuscular adipogenesis in beef cattle. We found that Wagyu stromal vascular cells proliferate faster than Angus stromal vascular cells. In addition, we observed that the myogenesis was down regulated but adipogenesis and fibrogenesis were enhanced in Wagyu. Currently, we are further defining mechanisms regulating intramuscular adipogenesis in beef cattle, focusing on the expression of Zfp423 and PPARG, two transcription factors critical to the early and late stages of adipogenesis.

Publications

  • Type: Journal Articles Status: Under Review Year Published: 2014 Citation: Wei, S., X. Fu, X. Liang, M. Zhu, Z. Jiang, S. M. Parish, M.V. Dodson, L. Zan, and M. Du. Enhanced mitogenesis in stromal vascular cells derived from subcutaneous adipose tissue of Wagyu compared with those of Angus cattle
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Dodson, M.V., R.E. Allen, M. Du, W.G. Bergen, S.G. Velleman, S.P. Poulos, M. Fernyhough-Culver, M.B. Wheeler, S.K. Duckett, M.R.I. Young, B.H. Voy, Z. Jiang, and G.J. Hausman. (2015). Invited Review: Evolution of meat animal growth research during the past 50 years: Adipose and muscle stem cells. Journal of Animal Science. In press.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Hausman, G. J., U. Basu, M. Du, M. E. Fernyhough-Culver, and M. V. Dodson. (2014). Intermuscular and intramuscular adipose tissues: bad vs. good adipose tissues. Adipocytes, Accepted.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Dodson, M.V., M. Du, S. Wang, W. Bergen, M.E. Fernyhough-Culver, Urmila Basu, S.Y. Poulos and G.J. Hausman. (2014). Adipose depots differ in cellularity, adipokines produced, gene expression and cell systems. Adipocyte 3(4):2-7.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Meyer, A. M., B. W. Hess, S. I. Paisley, M. Du, and J. S. Caton. (2014). Small intestinal measures are correlated with feed efficiency in market weight cattle, despite minimal effects of maternal nutrition during early to mid gestation. Journal of Animal Science, 92, 3855-3867.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Duarte, M. S., M.P. Gionbelli, P.V.R. Paulino, N.V.L. Ser�o, C.S. Nascimento, M.E. Botelho, T.S. Martins, S.C.V. Filho, M.V. Dodson, S.E.F. Guimar�es, M. Du. (2014). Maternal overnutrition enhances mRNA expression of adipogenic markers and collagen deposition in skeletal muscle of beef cattle fetuses. Journal of Animal Science, 92: 3846-3854.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Du, M. (2014). Meat Science and Muscle Biology Symposium  Implants, muscle development and meat quality. Journal of Animal Science, 92: 1-2.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Wei, S., L. Zhang, X. Zhou, M. Du, Z. Jiang, G. J. Hausman, W. G. Bergen, L. Zan, M. V. Dodson. (2013). Emerging roles of zinc finger proteins in regulating adipogenesis. Cellular and Molecular Life Sciences, 70:4569-4584.


Progress 01/01/13 to 09/30/13

Outputs
Target Audience: Scientific community, beef cattle producers Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Shawn Harris, a PhD student, has been involved in this study. Marcio Duarte and Weisheng Juan, two visiting scientists, were trained through experiments related to this project. How have the results been disseminated to communities of interest? We have published peer-reviewed papers and also made presentations. What do you plan to do during the next reporting period to accomplish the goals? We will further define mechanisms leading to high marbling in Wagyu beef cattle, focusing on mechanisms leading to enhanced adipogenesis compared to Angus cattle.

Impacts
What was accomplished under these goals? Intramuscular fat and collagen content are major factors affecting beef quality, but mechanisms regulating intramuscular adipose and connective tissue deposition are far from clear. Japanese Wagyu cattle are well known for their extremely high marbling. We compared Wagyu and Angus cattle for their adipogenic capacity and found that both intramuscular fat content and collagen content are enhanced in Wagyu compared to Angus cattle, which was due to enhanced adipogenesis and fibrogenesis in Wagyu. These data provide mechanistic insights into the extremely high marbling in Wagyu cattle.

Publications

  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Duarte, M. S., P. V. R. Paulino, A. K. Das, S. Wei, N. V. L. Serao, X. Fu, S. M. Harris, M. V. Dodson, and M. Du. (2013). Enhancement of adipogenesis and fibrogenesis in skeletal muscle of Wagyu compared to Angus cattle. Journal of Animal Science, 91: 2938-2946.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Duarte, M.S., M.P.Gionbelli, P.V.R. Paulino, N.V.L. Ser�o, L.H.P. Silva, R. Mezzomo, M.V. Dodson, M. Du, J.R. Busboom, S.E.F. Guimar�es, S.C. Valadares Filho. (2013). Effects of pregnancy and feeding level on carcass and meat quality traits of Nellore cows. Meat Science, 94: 139-144.
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2014 Citation: Wei, S., L. Zhang, X. Zhou, M. Du, Z. Jiang, G. J. Hausman, W. G. Bergen, L. Zan, M. V. Dodson. 2013. Emerging roles of zinc finger proteins in regulating adipogenesis. Cellular and Molecular Life Sciences, In press.


Progress 01/01/12 to 12/31/12

Outputs
OUTPUTS: Intramuscular fat or marbling is critical for the palatability of beef. In mice, very recent studies show that adipocytes and fibroblasts share a common pool of progenitor cells, with Zinc finger protein 423 (Zfp423) as a key initiator of adipogenic differentiation. To evaluate the role of Zfp423 in intramuscular adipogenesis and marbling in beef cattle, we sampled beef muscle for separation of stromal vascular cells. These cells were immortalized with pCI neo-hEST2 and individual clones were selected by G418. A total of 288 clones (3 x 96 well plates) were isolated and induced to adipogenesis. The presence of adipocytes was assessed by Oil-Red-O staining. Three clones with high and low adipogenic potential, respectively were selected for further analyses. In addition, fibro/adipogenic progenitor cells were selected using a surface marker, platelet derived growth factor receptor (PDGFR) α. The expression of Zfp423 was much higher in high adipogenic cells, while transforming growth factor (TGF)-β was higher in low adipogenic cells. Following adipogenic differentiation, the expression of peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer binding protein α (C/EBPα) were much higher in high adipogenic cells. Over-expression of Zfp423 in stromal vascular cells and cloned low adipogenic cells dramatically increased their adipogenic differentiation, accompanied with the inhibition of TGF-β expression. Zfp423 knockdown in high adipogenic cells largely prevented their adipogenic differentiation. The differential regulation of Zfp423 and TGF-β between low and high adipogenic cells is associated with the DNA methylation in their promoters. In conclusion, data show that Zfp423 is a critical regulator of adipogenesis in stromal vascular cells of bovine muscle, and Zfp423 may provide a molecular target for enhancing intramuscular adipogenesis and marbling in beef cattle. PARTICIPANTS: Min Du, Yan Huang, Mike V. Dodson, Marcio Duarte TARGET AUDIENCES: Scientific community, beef producers PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Beef cattle production is a main component of agriculture in Washington. Enhancing beef production efficiency and quality will have tremendous impacts on beef cattle farming and the associated meat packing industry. The proposed research is innovative, because very few studies have been conducted on the adipogenic and fibrogenic differentiation of bovine animals. Identified mechanisms will help to formulate breeding and management strategies to enhance marbling and tenderness of beef produced. In addition, grass-fed beef is an important emerging segment of beef cattle production, with most grass-fed beef produced by small farms in rural areas. Our data will be especially important for this emerging section. Grass-fed beef contains higher ratios of unsaturated fatty acids, especially n-3 fatty acids, which are good for health, but the poor flavor of grass-fed beef is the major problem hampering its wide acceptance. The flavor of beef mainly comes from intramuscular fat (marbling), and grass-fed beef lacks marbling which weakens its flavor and reduces juiciness. Therefore, if we can effectively enhance marbling in grass-fed beef, we can significantly improve the eating quality of grass-fed beef, further expanding this segment of the beef industry. The objective of this project is to identify mechanisms regulating intramuscular adipogenesis in beef cattle. Knowledge generated from our proposed studies will be disseminated to beef producers through our extension and education programs such as short courses. Also, results will be disseminated to scientific community through presentations at professional society meetings including the American Society of Animal Science, and publications in peer reviewed journals.

Publications

  • Dodson M.V., M.Du, S.Velleman, D.C.McFarland, M.E.Fernyhough, S.Wei, M.Duarte, Z.Jiang, G.J.Hausman. 2012. Skeletal muscle and adipose cell precursors: Stem cells with a bright place in medicine, tissue engineering and reconstructive surgery. Stem Cells with Fat Transfer in Aesthetic Procedures: Science, Art and Clinical Techniques. Ed. M.A. Shiffman, A. Di Guiseppe and F. Bassetto. Berlin. Springer.
  • Du M., Y.Huang, A.Das, Q.Yang, M.Duarte, M.V.Dodson, M.Zhu. 2012. Manipulating mesenchymal progenitor cell differentiation to optimize performance and carcass value of beef cattle. Journal of Animal Science. doi:10.2527/jas.2012-5670
  • Du M. 2012. Zfp423 promotes adipogenic differentiation of bovine stromal vascular cells. PLoS One. 7:e47496.