Brief Summary of Minutes of Annual Meeting

Dr. Spurlock opened the meeting by introducing Drs. Kinder and Mirando. Dr. Kinder gave a brief administrative update, and stressed the importance of communicating the value of the group and meeting activity in annual reports and summaries. Dr. Mirando provided insight as to potential factors that might impact funding opportunities and NCCC097 activities. In particular, Dr. Mirando noted that the USDA and NIH were developing a joint project that would support the use of meat animal models for human applications. Dr. Sean Adams, USDA-Western Human Nutrition Research Center, was nominated and elected as the new meeting Chair. Project Directors, graduate students, and postdoctoral researchers provided research updates (summarized below).

(1) Growth, Development, & Functional Aspects of Adipose Tissue: a unique adipocyte transcript was discovered using differential gene expression studies in rodent brown adipose tissue (BAT). The gene was found to be robustly expressed in rodent and human white adipose tissue (WAT) relative to other tissues, and is up-regulated during adipocyte differentiation in the 3T3-L1 cell system. The open reading frame of this gene (now termed Tusc5) encodes a region with high similarity to proteins containing an interferon-inducible transmembrane domain which could signal a role in adipocyte growth and maturation. Mouse genetics studies implicate Tusc5 in adiposity phenotypes. Recent tissue expression studies have revealed a highly-unusual pattern of co-expression in sensory nerve ganglia which contain the cell bodies of nerves carrying information from the periphery to the brain. These initial results suggest an important role for Tusc5 in normal adipocyte maturation and function, and indicate that at the protein shares a functional role between fat cells and sensory nervous system cells. (2) Dietary Constituents Regulating Lipid Deposition or Lipid Composition: (2a) The effects of feeding a diet containing omega-3 (n-3) fatty acids during late gestation (G) or lactation (L) on sow milk composition were determined. Encapsulated n-3 fatty acids (Fertilium, United Feeds, Sheridan, IN) added to corn-SBM-based G and L diets resulted in a shift of the n-6/n-3 ratio from ~20 in control sows to 10-13 in n-3 supplemented sows. The addition of n-3 increased the total n-3 percent in milk (1.35%, vs. 1.12% in controls, p<0.0001) and significantly reduced the milk n-6/n-3 ratio from 17.49 to 14.56. n-3 addition to the L diet increased the level of EPA and DHA , regardless of whether sows consumed n-3 in the G diet. Litter weights were not affected by these interventions. Thus, even small amounts of n-3 in the maternal diet can influence milk composition. (2b) The influence of ractopamine (RAC) and dietary protein on carcass composition, performance, and meat quality were evaluated in crossbred gilts and sows. Dietary treatments included feeding crude protein (CP) at 16% (16CP), 18% (18CP), or 16% plus supplementation with Lys, Met, Thr to equal the 18% CP diet (16AA), with or without RAC at 10 ppm for 28 d. There was a trend toward higher ADG vs. the other groups for pigs fed 16AA, and these animals had significantly increased 28 d F:G vs. 16CP. RAC increased 28 d G:F, and UBF and ULM were increased in RAC pigs vs. controls. Tenth rib backfat (TRBF) was lower in 18CP vs. 16CP, with 16AA intermediate. RAC-fed pigs tended (p<0.08) to have lower TRBF, and had larger lean mass areas vs. controls. No meat quality differences were observed across dietary protein level or gender; LM from RAC-fed pigs was less red and less yellow, had higher 24 h pH, and higher NPPC marbling score. Overall, RAC supplementation may be an effective means of improving composition without compromising quality, regardless of dietary protein level. (3) Regulation of Energy Balance: Male Sprague Dawley rats and Zucker fatty rats were used to test how diets differing in macronutrient content influence a suite of hormones believed to participate in appetite regulation and energy balance. Rats were assigned for 5 wk to one of five diets: control, 75% of control calories, American Heart Association (AHA), Atkins, or high fat (HF). Weekly plasma samples were analyzed for ghrelin, leptin, insulin, and adiponectin, and 5 wk plasma also assayed for glucagon, oxyntomodulin, and metabolites. Macronutrient content in the diet and genetic background altered endocrine patterns. For instance, in obese Zuckers glucagon was significantly higher in the Atkins rats compared to HF-fed or calorie-restricted animals, and tended to be higher vs. control diet or AHA rats (p<0.1). In lean rats, glucagon was highest in the control conditions relative to other diets. These results support the idea that the endocrine response to dietary macronutrient can differ in obese vs. lean animals. (4) Dietary Constituents Regulating Lipid Deposition or Lipid Composition: A porcine-specific oligo gene array platform (provided by the Pig Genome Coordination Program at Iowa State University) was used to analyze differentially-expressed RNA transcripts in various pig tissues following 28 d RAC supplementation (60 ppm) (Study XP1) or after high-fat (40% HF) feeding for 14 d (Study XP2) in finishing pigs. Treatment-related differences in transcript intensities were evaluated using algorithms designed to account for multiple-comparisons in micro arrays (SAM software, Stanford University). In Study XP1, 550 genes were upregulated, and 569 downregulated, in WAT by RAC. Almost half of these genes lacked adequate sequence information to positively identify. RAC decreased expression of fatty acid synthase, stearoyl-CoA desaturase, acetyl-CoA carboxylase, and leptin, and increased some oxidative transcripts. In Study XP2, HF-feeding was found to alter expression of 1055, 847, and 1138 genes in liver, adipose, and muscle, respectively. In liver, FABP, fatty acid synthesis genes, and fatty acid oxidation genes were decreased. In adipose, expression of FABP, leptin, and TAG synthesis genes were increased by HF, and in muscle this treatment led to upregulation of CPT1, 3-hydroxyacyl-CoA dehydrogenase, and some genes related to oxidative phosphorylation. (5) Growth, Development, & Functional Aspects of Adipose Tissue: The impact of the lipid-soluble polybrominated diphenyl ethers (PBDEs) on adipose and mammary biology was determined. Since these chemicals are widely-dispersed in the environment, exposure may be relatively high and accumulation in fatty tissues may influence normal processes of adipose development and milk composition. PDBEs are lipophilic, continuously-released from consumer products (i.e., upholstery, computers, curtains), and persistent in the environment. They are believed to disrupt several endocrine pathways, and levels in biofluids such as milk could be important considering that magnification of PDBE concentrations occurs up the food chain. A study currently underway is examining the PDBE levels of breast milk in women whose exposure is assessed by a comprehensive questionnaire regarding their home and workplace environment, to understand potential contamination and whether this is correlated with possible environmental sources. A second study has determined that 4 wk exposure to PDBEs, given by oral gavage to rats (14 mg/kg in oil) led to significantly increased isoproterenol-stimulated lipolysis, and markedly reduced insulin-stimulated glucose uptake, in adipocytes isolated from treated rats vs. controls. These effects were not observed when control adipocytes were acutely exposed to PDBEs, or in rats dosed in vivo for 2 wk, however. (6) Growth, Development, & Functional Aspects of Adipose Tissue: The ontogeny of expression for twenty secreted proteins was evaluated in stromal-vascular (SV) cells and adipose tissue derived from 105 d fetuses and young pigs (aged 5, 90, 150, and 210 d), using microarray and RT-PCR technologies. Genes assayed included apolipoprotein A-1 (APO-A1), relaxin, RANTES, brain-derived neurotrophic factor (BDNF), IGF-binding protein 5 (IGFBP-5), several interferons, and a number of interleukins. Outer (OSQ) and inner (ISQ) subcutaneous adipose were evaluated. Microarray results indicated that, with the exception of connective tissue growth factor (CTGF) which was upregulated in OSQ with age, no genes encoding secreted molecules were changed with age in pig adipose tissue. However, real-time quantitative RT-PCR indicated significant changes in IL-1A, IL-1B, IL-6, RANTES, and several others with age. Therefore, the endocrine function of WAT changes in developing pigs, which could have functional effects on adipose growth, as well as metabolic function locally and in other tissues. (7) Dietary Constituents Regulating Lipid Deposition or Lipid Composition: (7a) The effects of supraphysiologic arachidonic acid (AA) exposure to the gut via supplementation of a milk-based formula to newborn piglets were determined. Specifically, the time-related effects of AA on fatty acid composition of enterocyte phospholipids and on the abundance of delta6- and delta5-desaturase mRNAs were tested. The diets contained either no PUFA (control), 5% eicosapentanoic acid (EPA), or 0.5%, 2.5%, 5% AA, and were fed for 4, 8, or 16 d starting at 1 d of age. At 16 d, enterocyte phospholipids containing AA were enriched significantly (i.e., by 355% in the 5% AA group) vs. controls or EPA treated, whereas in the latter phospholipids displayed a significant increase in EPA. Diet had no effect on growth, and desaturase mRNA did not differ with the exception of a reduction in the 2.5% AA group relative to control. (7b) The degree by which carnitine supplementation to sows increases tissue carnitine concentration in pig fetuses was tested. Pregnant gilts were fed control diets or diets supplemented with carnitine (88 mg/d) during the first 70 d gestation, and tissue carnitine concentration and carnitine palmitoyltransferase (CPT) kinetics determined in fetuses at 70 d. Maternal supplementation increased fetal liver and muscle tissue carnitine concentration significantly (by 58% and 33%, respectively) but did not affect kidney levels. CPT activity and kinetics were unaffected by changes in tissue carnitine concentration. (8) Dietary Constituents Regulating Lipid Deposition or Lipid Composition: Potential dietary fat-related shifts in bovine mammary tissue mRNA levels for genes encoding proteins relevant to lipid uptake and lipogenesis were evaluated (acetyl-CoA carboxylase, fatty acid synthase, lipoprotein lipase, stearoyl-CoA desaturase). The overall objective was to employ strategic nutritional inputs (fatty acid supplements) to affect mammary lipogenesis, which may influence adipose tissue accretion and thus modulate the postpartum interval for successful rebreeding. Primiparous beef cattle were fed either a low-fat control diet or a cracked high-linoleate safflower see supplement (LIN), and mammary lipogenic gene expression was evaluated both through direct measurement of mammary tissue samples derived at slaughter, and via mRNA isolation and analysis from the milk (exuded cells). Correlation of mRNA abundance comparing tissue and milk somatic cell RNAs was significant for all genes tested (Pearson correlation coefficients of 0.67-0.90; p<0.05), indicating that milk cell RNA is a valid surrogate for mammary tissue RNA for these genes. No significant treatment-related shifts in gene expression was observed. The milk mRNA protocol was used to test whether differences in body condition score (BCS) and dietary fat alter expression. Three year old beef cows were nutritionally managed to achieve a BCS of 4 or 6, and then given a low-fat control diet or one of two high-fat supplements (linoleate or oleate). Milk somatic cell RNA was analyzed at 30 d and 60 d of lactation. Lipid supplementation had no effect on gene expression; however, lower BCS was characterized by increased FAS and SCD expression, possibly reflective of a greater requirement for de novo mammary lipogenesis to compensate for the lower overall adipose to draw on for milk fat synthesis. There were significant effects of lactation duration on gene expression, with increased LPL and reduced FAS at 60 d vs. 30 d, which might reflect a shift toward higher demand for mammary uptake of circulating lipids and lower lipid synthesis later in lactation. (9) Growth, Development, & Functional Aspects of Adipose Tissue: A dynamic, mechanistic model of beef cattle growth and body composition is being developed, with a goal to provide a predictive model useful to industry (as a decision-aid tool) and researchers (to test hypotheses and interpret results). The current Davis Growth Model evolved from previous models and is based on the concepts of hyperplasia and hypertrophy to simulate the accretion of whole body protein and four fat depots (visceral, intermuscular, subcutaneous, and intramuscular). A meta-analysis of over 20 yr of literature has been performed to aid in model development. Reference animals growing from 120-600 kg have been established, with accompanying compositional data: early-moderate maturing and late maturing steers, both implanted and non-implanted. Preliminary fits using the model equations for fat distribution indicate little variation in parameter values for the various scenarios, indicating that the model responds well to variations in mature size and endocrine status. (10) Growth, Development, & Functional Aspects of Adipose Tissue: The autocrine/paracrine regulation of glucose transport and fatty acid oxidation by adiponectin has been studied in primary pig adipocytes and in the 3T3-L1 line. Adiponectin stimulates the expression of several genes readily identifiable with fatty acid oxidation, and in fact, stimulates palmitate oxidation in vitro. Additionally, the potential for hyperglycemic conditions to induce insulin resistance in adipocytes was investigated. Insulin resistance (decreased insulin-mediated 2-deoxy-D-glucose uptake) was induced by hyperglycemia, but adiponectin did not alleviate this condition. However, adiponectin did suppress the induction of reactive oxygen species (ROS) by hyperglycemia, an effect that was achieved in part by suppression of NADH oxidase and PKC-´. The regulation of ROS may be an key mechanism by which adiponectin suppresses the inflammation associated with hyperglycemia. (11) Growth, Development, & Functional Aspects of Adipose Tissue: The possibility that mature fat cells can return to a proliferation-competent state and to de-differentiate under certain conditions is being examined through the use of adipofibroblasts (progeny from de-differentiated adipocytes) in culture. Regulation of lipid metabolism, adipogenesis, and plasticity (ability to convert to other cell types) are being studied in this cell model to characterize the physiological characteristics of de-differentated fat cells.

Adams, S.H., C. Lei, C.M. Jodka, S.E. Nikoulina, J.A. Hoyt, B. Gedulin, C.M. Mack, and E.S. Kendall. 2006. PYY[3-36] administration decreases the respiratory quotient and reduces adiposity in diet-induced obese mice. J. Nutrition, 136: 195-201. Adams, V.L., C.D. Gilbert, H.J. Mersmann, and S.B. Smith. 2005. Conjugated linoleic acid depresses [3H]-thymidine incorporation into stromal-vascular cells of adipose tissue from postweanling pigs. Adipocytes 1:65-72. Azain, M. J., J. R. Broderson, and R. J. Martin. 2006. Effect of long-term somatotropin treatment on body composition and lifespan in aging obese Zucker rats. Experimental Biology and Medicine. 231: 76-83. Brewer, M.S., J. Novakofski and K. Freise.2006. Instrumental evaluation of pH effects on ability of pork chops to bloom. Meat Science 72( 4):596-602. Fernyhough, M.E., J.L. Vierck and M.V. Dodson. 2006. Assessing a non-traditional view of adipogenesis: adipocyte dedifferentiation  mountains or molehills? Cells, Tissues, Organs 182(3-4): [accepted 10 April] Jablonski, E. A., R. D. Jones, and M.J. Azain. 2006. Evaluation of pet food by-product as an alternative feedstuff in weanling pig diets. J. Anim. Sci. 84: 221-228. Novakofski, J. and M. S. Brewer. 2006. The Paradox of Toughening During The Aging of Tender Steaks, Journal of Food Science. In press. Schlegel, M.L., Bergen, W.G., Schroeder, A.L., VandeHaar, M.J., Rust, S.R. 2006. Use of bovine somatotropin for increased skeletal and lean tissue growth of Holstein steers. J. Anim. Sci. 84:1176-1187. Strat, A.L., T.J. Kokta, M.V. Dodson, A. Gertler, Z. Wu and R.A. Hill. 2005. Early signaling interactions between the insulin and leptin pathways in bovine myogenic cells. Biochemica et Biophysica ACTA (Molecular Cell Research) 1744(2):164-175. Wang, H.C., Y.K. Ko, H.J. Mersmann, C.L. Chen and S.T. Ding. 2006. The expression of genes related to adipocyte differentiation in pigs. J. Anim. Sci. 84:1059-1066.