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Plasma leptin in growing lambs as a potential predictor for carcass composition and daily gain
MEAT SCIENCE Meat Science 74 (2006) 600–604 www.elsevier.com/locate/meatsci
Plasma leptin in growing lambs as a potential predictor for carcass composition and daily gain M. Altmann a, H. Sauerwein b, E. von Borell a
a,*
Institute of Animal Breeding and Husbandry with Veterinary Clinic, Martin-Luther-University Halle-Wittenberg, D-06108 Halle, Adam-Kuckhoff-Str. 35, Germany b Institute of Animal Sciences, Bonn University, 53115 Bonn, Katzenburgweg 7-9, Germany Received 13 April 2006; received in revised form 8 May 2006; accepted 8 May 2006
Abstract Leptin is an adipocyte derived hormone and correlates highly to the extent of body fat tissue. The aim of this study was to determine if leptin could serve as an early predictor for carcass composition and final growth rate in lambs with special emphasis on size and cellularity of the different body fat depots. Thirty intact male ad libitum fed lambs were blood sampled at 20, 25, 30, 35, and 40 kg live weight. After slaughtering at 40 kg, lean and the visceral, subcutaneous and intermuscular fat were measured by dissection. The fat cell diameter was determined in subcutaneous and perirenal fat. Average daily gain from birth to slaughter correlated to leptin only at 30 and 35 kg live weight (r = 0.56 and 0.61, P < 0.01) and thus leptin cannot be regarded as a suitable early predictor for growth rate. That goes for the prediction of subcutaneous and intermuscular fat, too; because no relationships were detected between early leptin concentrations and the amount of these tissues. Leptin concentrations measured just before slaughter were related to all fat tissues except the pelvic and intramuscular fat. Among the visceral fat depots, omental fat expressed the highest correlations to leptin (r = 0.60, P < 0.001). Additionally, leptin concentrations at 35 and 40 kg live weight increased with increasing fat cell diameters (r = 0.38, P < 0.05 to r = 0.59, P < 0.001). This study indicates that leptin concentration measured in the slaughter weight range has the greatest potential to assess body fat content, whereas an earlier prediction does not seem to be feasible. Further studies should clarify if these results are reproducible for other breeds or species. 2006 Elsevier Ltd. All rights reserved. Keywords: Lambs; Leptin; Growth; Carcass composition
1. Introduction Subjective assessments or ultrasound measurements are two common methods for the in vivo evaluation of carcass merit for breeding purposes. In addition, the utilization of hormones as predictors for production characteristics has been discussed over several years. The value of an early assay based selection in the meat industry depends on region-specific breeding and marketing conditions. A model calculation for the selection of beef cattle in Australia showed that an early selection for IGF-I at weaning *
Corresponding author. Tel.: +49 345 5522332; fax: +49 345 5527106. E-mail address: [email protected] (E. von Borell). 0309-1740/$ - see front matter 2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.meatsci.2006.05.013
increased market profitability (Wood, Archer, & van der Werf, 2004). The IGF-I test has been commercially used since 2004 in Australia’s cattle performance testing (Graser, 2004). Leptin is another potential hormone for an assay based prediction of growth or carcass composition. It is released mainly from the adipose tissue and correlates with body fat. Geary et al. (2003) recommend leptin as an additional index for fat content in beef. The accuracy for the estimation of carcass fat in lambs by plasma leptin concentrations at the time of slaughter was similar to ultrasound fat thickness measurements (Altmann, Sauerwein, & von Borell, 2005). However, the relationships between leptin concentrations in early life and final production characteristics are poorly understood. Kawakita et al. (2001) considered
M. Altmann et al. / Meat Science 74 (2006) 600–604
the correlation between early leptin concentration and final backfat thickness as being too low for a prediction of adiposity in steers. The aim of the present study was to determine the relationship of leptin plasma concentrations during growth to carcass composition and daily gain of lambs with special emphasis on size and cellularity of several fat depots. 2. Materials and methods 2.1. Animals Intact male East Frisian Milk sheep (N = 15) and Blackheaded Mutton · East Frisian Milk sheep (N = 15) lambs were used in this study. The lambs were born in late winter and were naturally reared by their dams. They were grouphoused in deep litter pens and had ad libitum access to concentrates (10.2 MJ NE/kg) and hay. The lambs were weighed weekly. The blood-sampling period began at 20.0 (SEM 0.18) kg and ended at 40.5 kg (SEM 0.23) which corresponded to a mean age of 38.5 (SEM 1.7) and 97.1 (SEM 3.1) days, respectively. Weaning occurred on two fixed days when lambs averaged 60 days of age. The mean live weight at weaning was 27.6 kg. When the lambs reached the final weight of 40 kg they were feed deprived for 24 h immediately after the last blood collection to reduce the gut fill before slaughter. Perirenal, pelvic, intestinal, and omental fat was weighed and the sum of these tissues was designated as visceral fat. The left carcass side was dissected into lean, subcutaneous and intermuscular fat and bones. These tissue masses were multiplied by 2 for the calculation of the total carcass composition. Furthermore, the intramuscular fat content was analyzed in a sample of the M. longissimus dorsi between the 13th and 14th rib by n-hexane extraction (Soxtherm 406, Gerhardt Apperate GmbH & Co. KG, Bonn, Germany). Samples of subcutaneous and perirenal fat were collected and frozen at 20 C for the determination of the cell diameters.
601
2.3. Measurement of fat cell diameters The samples were sectioned in 20 lm thick slices on a Microtome-Cryostat and fixed in formol calcium for 5 min. The slices were stained with haemalaun and fat was removed by xylen for 30–60 s. The diameter was measured in 300 cells for each sample using a light microscope with a 4 · 10 objective (Labophot-2, NIKON, Japan) connected to a 3CCD color video camera (DXC 9100P, SONY, Japan). Fat cell diameters were analyzed with an image analysis program (Lucia-G, version 4.8, NIKON, Japan). 2.4. Data analysis Data from purebred and crossbred lambs were combined for the statistical analysis because genotype did not influence hormone concentrations and performance traits. Hormone concentrations in lambs of different live weight were analyzed by ANOVA and Duncan’s multiple comparison post hoc tests. The relationships between average daily gain, carcass composition and hormone concentrations were described by Pearson correlations and regression equations. 3. Results Lambs in this study realized a high level of average daily gain (Table 1) from birth to slaughter which is comparable to performance test results for East Frisian Milk sheep in Germany. The mass of the various fat tissues showed higher coefficients of variation (19.0–39.6%) than lean mass (7.1%). Leptin concentrations in plasma rise with increasing live weight during growth and reached the highest level at 40 kg live weight (Fig. 1). The effect of live weight was significant (P < 0.001). A negative relationship between leptin and average daily gain was observed at 30 and 35 kg live weight (Table 2).
2.2. Blood sampling and hormone assays Blood samples were drawn at 20, 25, 30, 35, and 40 kg live weight. Five blood samples were collected from each lamb between 9.00 h and 11.00 h at 30 min intervals and then centrifuged. In order to obtain a mean hormone concentration over 2 h, sera were pooled for each animal, aliquoted and stored at 20 C until analyzed. Hormone concentrations in pooled samples have been shown to adequately represent those obtained from serial samples (Tiwary, 1986). Plasma leptin concentrations were analyzed with a specific enzyme immunoassay that has been validated for use in several ungulate species, including ovine samples (Sauerwein, Heintges, Hennies, Selhorst, & Daxenberger, 2004). The intra-assay coefficient of variation was 6.3%, the inter-assay coefficient of variation was 13.9%, and the limit of detection was 0.3 ng/ml.
Table 1 Average daily gain and carcass composition of experimental lambs
Average daily gain, g/day Total dissectible fata, g Visceral fatb, g Intestinal fat, g Omental fat, g Perirenal fat, g Pelvic fat, g Carcass fat, g Intermuscular fat, g Subcutaneous fat, g Intramuscular fat, g/100 g of M. longissimus Lean (g) Perirenal fat cell diameter (lm) Subcutaneous fat cell diameter (lm) a b
Mean
SEM
CV (%)
373 3547 912 344 344 193 31 2635 1773 862 1.6 12033 57.0 66.1
9.4 129.2 33.7 11.9 16.5 9.9 2.2 106.7 67.5 42.3 0.1 155.4 1.2 1.7
13.9 19.9 20.2 19.0 26.3 28.1 39.6 22.2 20.9 26.8 32.7 7.1 11.6 13.8
The sum of visceral and carcass fat. The sum of intestinal, omental, perirenal, and pelvic fat.
602
M. Altmann et al. / Meat Science 74 (2006) 600–604
Leptin correlated to the visceral fat at all live weights except 20 kg, while the correlations to carcass fat were significant only at 40 kg live weight. Among the visceral fat depots, pelvic fat showed no correlations (P > 0.05) to leptin concentrations. The correlations to omental and perirenal fat tended to be stronger as live weight increased. No correlations were observed between plasma leptin and lean mass. The relationships between fat cell size and plasma leptin measured during growth are represented in Table 3. Leptin was correlated more strongly to the fat cell diameter in perirenal fat than to the subcutaneous fat. While no associations could be found between fat cell diameter and leptin measured in lambs of 20–30 live weight, the leptin concentrations at 35 and 40 kg live weight correlated to the mean fat cell diameter in a range of r = 0.38–0.59 (P < 0.05– 0.001). The correlations in Table 2 indicate, that total dissectible fat could be best predicted by leptin concentrations at
Table 3 Correlations between leptin concentrations during growth and fat cell diameters of subcutaneous and perirenal fat Growth period (kg live weight)
Perirenal fat
20 25 30 35 40 * ** ***
Subcutaneous fat
0.08 0.16 0.25 0.59*** 0.57***
0.05 0.01 0.02 0.38* 0.49**
P < 0.05. P < 0.01. P < 0.001.
40 kg live weight. The regression equation fitted for 40 kg live weight is represented in Fig. 2. The R2 value was 0.24. 4. Discussion The present study focused on the value of leptin as a potential predictor for final performance traits in growing
6.0
c 5500
5.0
b
Total dissectible fat, g
Leptin, ng/ml
5.5
4.5 4.0 3.5
a a a
3.0 2.5 20
25
30
35
40
5000 4500 4000 3500 3000
2000
Live weight, kg Fig. 1. Leptin concentrations during the growth period of 20–40 kg live weight (means and standard error of means, means with different letters differ significantly, P < 0.05).
y = 2183 + 258 * x; P < 0.01
2500
R2 = 0.24; RMSE = 629 4
3
5
6
7
8
9
Leptin, ng/ml Fig. 2. Relationship between leptin concentration at 40 kg live weight and total dissectible fat.
Table 2 Correlations between leptin measured at different live weights and average daily gain and tissue masses Growth period (kg live weight) 20 Average daily gain, g/day Total dissectible fata, g Visceral fatb,g Intestinal fat, g Omental fat, g Perirenal fat, g Pelvic fat, g Carcass fat, g Intermuscular fat, g (M. longissims) Subcutaneous fat, g Intramuscular fat, g/100 g of M. longissimus Lean, g a b * ** ***
The sum of visceral and carcass fat. The sum of intestinal, omental, perirenal, and pelvic fat. P < 0.05. P < 0.01. P < 0.001.
0.21 0.09 0.06 0.09 0.03 0.09 0.16 0.09 0.14 0.01 0.15 0.23
25 0.10 0.26 0.41* 0.36* 0.35* 0.36* 0.10 0.19 0.18 0.18 0.06 0.19
30 0.56** 0.22 0.43* 0.43* 0.38* 0.29 0.02 0.13 0.09 0.19 0.32 0.34
35 0.61** 0.26 0.54** 0.27 0.63*** 0.46** 0.06 0.14 0.08 0.23 0.31 0.00
40 0.19 0.49** 0.58** 0.40* 0.60*** 0.46** 0.11 0.41* 0.36* 0.45* 0.19 0.04
M. Altmann et al. / Meat Science 74 (2006) 600–604
lambs and examined the relationships of average daily gain and body composition to hormone concentrations measured at several times before slaughter. The study started at 20 and ended at 40 kg live weight. This growth phase corresponds to the length of lamb performance testing in Germany and 40 kg is a common live weight for slaughter lambs in some European countries. A hormonal indicator should be applicable at 20 kg live weight or earlier for preselection or at 40 kg live weight for the prediction of carcass value in lambs kept for breeding purposes. Plasma leptin did not change significantly between 20 and 30 kg live weight within the range in which weaning occurred. However, it increased at 35 kg until 40 kg. It is assumed that this elevation was caused by a progressive increase of fat accretion. Leymaster and Jenkins (1993) found in crossbred lambs on an intensive production system an accelerated deposition of kidney-pelvic and carcass fat with advanced live weight, corresponding with the weight range in the present study. To our knowledge, the relationship of early plasma leptin concentrations to carcass composition at slaughter was only reported in one study with cattle (Kawakita et al., 2001). The authors found in steers which were fed a diet of 16% crude protein a correlation of r = 0.59 between early leptin concentrations and backfat thickness whereas no correlation existed in steers fed with 12% crude protein. The negative correlation to average daily gain at 30 and 35 kg live weight in our study implies that slow growing lambs had high leptin concentrations. This could be explained by the inhibitory effect of leptin on feed intake and the stimulatory influence on energy expenditure (Clarke, Tilbrook, Turner, Doughton, & Goding, 2001; Henry et al., 1999; Morrison et al., 2001; Tokuda, Matsui, Ito, Torii, & Yano, 2000). No correlations to average daily gain were found at lower live weights (20 and 25 kg) and thus leptin cannot be regarded as a suitable predictor for early growth rate selection. That goes for the prediction of carcass fat, too; because no relationships were detected between early leptin concentrations and the amounts of these fat tissues. Only the visceral fat correlated to the leptin concentrations from 25 kg live weight on. All fat tissues except the pelvic and intramuscular fat were significantly related with leptin concentrations at slaughter. Furthermore, a great number of other studies in several species (Altmann, Sauerwein, & von Borell, in press; Berg et al., 2003; Daniel et al., 2002; Ehrhardt et al., 2000; Geary et al., 2003; Yamada, Kawakami, & Nakanishi, 2003) indicate that several body fat tissues are highly associated with blood leptin concentration measured at the same time as body fat. A correlation of 0.68 between leptin and body fatness was found in ewes that varied widely in the deposition of fat tissue (Delavaud et al., 2000). In a previous study, we observed correlations from 0.40 to 0.64 between blood leptin concentrations and the mass of several fat depots in lambs of 35 or 45 kg live weight, respectively (Altmann et al., 2005).
603
Plasma leptin concentration is influenced by the amount of fat reserves and the expression of leptin in the adipocytes (Ehrhardt, Slepetis, Bell, & Boisclair, 2001; Yuen et al., 2002). Leptin mRNA correlated positively to the adipocyte volume in humans (Oberkofler et al., 1997), rats (Villafuerte et al., 2000), mice (Guo, Halo, Leibel, & Zhang, 2004) and steers (Yang et al., 2003). Because fat cell diameter in the subcutaneous fat in our study was higher than in perirenal fat and the mass of subcutaneous fat was 4-fold higher than the perirenal fat mass, we expected a stronger correlation of plasma leptin to the subcutaneous fat mass than to the perirenal fat. However, this was not observed. Omental fat contributed to 9.7% of total fat, but to a much higher proportion of 36% to the variation of plasma leptin. This indicates a high expression of leptin in omental fat. Another study in steers showed no difference in correlations between leptin and visceral, subcutaneous, and intermuscular fat (Yamada et al., 2003). The correlations between leptin and body composition indicate that plasma leptin concentration at the end of the fattening period (40 kg live weight) is the most suitable predictor of body fat. It explained 24% of the variance in total dissectible fat. Similar results were obtained by Altmann et al. (2005) for the prediction of carcass fat by leptin concentrations measured before slaughter in lambs of 35 and 45 kg live weight. Our study confirmed the lack of a correlation between plasma leptin and lean mass as leptin is released from adipose tissue and can therefore not be considered as a predictor for lean tissue. 5. Implications Our data suggest that plasma leptin cannot be recommended as an early predictor of total fat in lambs. However, it has the potential to serve as a second stage selection criterion in breeding rams at the weight range of slaughter lambs. Further studies should clarify if these results are applicable to other breeds or species. References Altmann, M., Sauerwein, H., & von Borell, E. (2005). Relationship between plasma leptin concentrations and carcass composition in fattening mutton: a comparison with ultrasound results. Journal of Animal Physiology and Animal Nutrition, 89, 326–330. Altmann, M., Sauerwein, H., & von Borell, E. (in press). The relationships between leptin concentrations and body fat reserves in lambs are reduced by short-term fasting. Journal of Animal Physiology and Animal Nutrition. Berg, E. P., McFadin, E. L., Maddock, K. R., Goodwin, R. N., Baas, T. J., & Keisler, D. H. (2003). Serum concentrations of leptin in six genetic lines of swine and relationship with growth and carcass characteristics. Journal of Animal Science, 81, 167–171. Clarke, I. J., Tilbrook, A. J., Turner, A. I., Doughton, B. W., & Goding, J. W. (2001). Sex, fat and the tilt of the earth: effects of sex and season on the feeding response to centrally administered leptin in sheep. Endocrinology, 142, 2725–2727. Daniel, J. A., Whitlock, B. K., Baker, J. A., Steele, B., Morrison, C. D., Keisler, D. H., et al. (2002). Effect of body fat mass and nutritional
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status on 24-hour leptin profiles in ewes. Journal of Animal Science, 80, 1083–1089. Delavaud, C., Bocquier, F., Chillard, Y., Keisler, D. H., Gertler, A., & Kann, G. (2000). Plasma leptin determination in ruminants: effect of nutritional status and body fatness on plasma leptin concentration assessed by a specific RIA in sheep. Journal of Endocrinololgy, 165, 519–526. Ehrhardt, R. A., Slepetis, R. M., Bell, A. W., & Boisclair, Y. R. (2001). Maternal leptin is elevated during pregnancy in sheep. Domestic Animal Endocrinology, 21, 85–96. Ehrhardt, R. A., Slepetis, R. M., Siegel-Willott, J., Van Amburgh, M. E., Bell, A. W., & Boisclair, Y. R. (2000). Development of a specific radioimmonoassay to measure physiological changes of circulating leptin in cattle and sheep. Journal of Endocrinology, 166, 519–528. Geary, T. W., McFadin, E. L., MacNeil, M. D., Grings, E. E., Short, R. E., Funston, R. N., et al. (2003). Leptin as a predictor of carcass composition in beef cattle. Journal of Animal Science, 81, 1–8. Graser, H. (2004). IGF-I blood test improves NFI EBVs. Breedplan News, 14, 12. Guo, K. Y., Halo, P., Leibel, R. L., & Zhang, Y. (2004). Effects of obesity on the relationship of leptin mRNA expression and adipocyte size in anatomically distinct fat depots in mice. American Journal of Physiology: Regulatory, Integrative and Comparative Physiology, 287, 56–61. Henry, B. A., Goding, J. W., Alexander, W. S., Tilbrook, J. A., Canny, B. J., Duneshea, F., et al. (1999). Central administration of leptin to ovariectomized ewes inhibits food intake without affecting the secretion of hormones from the pituitary gland: evidence for a dissociation of effects on appetite and neuroendocrine function. Endocrinology, 140, 1175–1182. Kawakita, Y., Abe, H., Hodate, K., Iguchi, A., Kobayashi, M., Mori, T., et al. (2001). The relation between plasma leptin concentrations and carcass lipid contents in Japanese Black steers. Livestock Production Science, 73, 25–34. Leymaster, K. A., & Jenkins, T. G. (1993). Comparison of Texel- and Suffolk-sired crossbred lambs for survival, growth and compositional trait. Journal of Animal Science, 71, 859–869. Morrison, C. D., Daniel, J. A., Holmberg, B. J., Djiane, J., Raver, N., Gertler, A., et al. (2001). Central infusion of leptin into well-fed and undernourished ewe lambs: effects on feed intake and serum concen-
trations of growth hormone and luteinizing hormone. Journal of Endocrinology, 168, 317–324. Oberkofler, H., Beer, A., Breban, D., Hell, E., Krempler, F., & Patsch, W. (1997). Human obese gene expression: alternative splicing of mRNA and relation to adipose tissue localization. Obesity Surgery, 7, 7390–7396. Sauerwein, H., Heintges, U., Hennies, M., Selhorst, T., & Daxenberger, A. (2004). Growth hormone induced alterations of leptin serum concentrations in dairy cows as measured by a novel enzyme immunoassay. Livestock Production Science, 87, 189–195. Tiwary, C. M. (1986). An inexpensive method for assessing pituitary response to luteinizing hormone releasing hormones: analysis of serum gonadotropins in pooled samples. Hormone and Metabolic Research, 18, 616–620. Tokuda, T., Matsui, T., Ito, J., Torii, S., & Yano, H. (2000). The changes in body weight and plasma metabolite levels during leptin injection are caused by the reduction of food intake in sheep. Animal Science, 71, 343–348. Villafuerte, B. C., Fine, J. B., Bai, Y., Zhao, W., Fleming, S., & DiGirolamo, M. (2000). Expressions of leptin and insulin-like growth factor-I are highly correlated and region-specific in adipose tissue of growing rats. Obesity Research, 8, 646–655. Wood, B. J., Archer, J. A., & van der Werf, J. H. J. (2004). Response to selection in beef cattle using IGF-I as a selection criterion for residual feed intake under different Australian breeding objectives. Livestock Production Science, 91, 69–81. Yamada, T., Kawakami, S. I., & Nakanishi, N. T. (2003). The relationship between leptin concentrations and the distribution of body fat in crossbred steers. Animal Science Journal, 74, 95–100. Yang, S. H., Matsui, T., Kawachi, H., Yamada, T., Nakanishi, N., & Yano, H. (2003). Fat depot-specific differences in leptin mRNA expression and its relation to adipocyte size in steers. Animal Science Journal, 74, 17–21. Yuen, B. S., Owens, P. C., McFarlane, J. R., Symonds, M. E., Edwards, L. J., Kauter, K. G., et al. (2002). Circulating leptin concentrations are positively related to leptin messenger RNA expression in the adipose tissue of fetal sheep in the pregnant ewe fed at or below maintenance energy requirements during late gestation. Biology of Reproduction, 67, 911–916.
Plasma leptin in growing lambs as a potential predictor for carcass composition and daily gain M. Altmann a, H. Sauerwein b, E. von Borell a
a,*
Institute of Animal Breeding and Husbandry with Veterinary Clinic, Martin-Luther-University Halle-Wittenberg, D-06108 Halle, Adam-Kuckhoff-Str. 35, Germany b Institute of Animal Sciences, Bonn University, 53115 Bonn, Katzenburgweg 7-9, Germany Received 13 April 2006; received in revised form 8 May 2006; accepted 8 May 2006
Abstract Leptin is an adipocyte derived hormone and correlates highly to the extent of body fat tissue. The aim of this study was to determine if leptin could serve as an early predictor for carcass composition and final growth rate in lambs with special emphasis on size and cellularity of the different body fat depots. Thirty intact male ad libitum fed lambs were blood sampled at 20, 25, 30, 35, and 40 kg live weight. After slaughtering at 40 kg, lean and the visceral, subcutaneous and intermuscular fat were measured by dissection. The fat cell diameter was determined in subcutaneous and perirenal fat. Average daily gain from birth to slaughter correlated to leptin only at 30 and 35 kg live weight (r = 0.56 and 0.61, P < 0.01) and thus leptin cannot be regarded as a suitable early predictor for growth rate. That goes for the prediction of subcutaneous and intermuscular fat, too; because no relationships were detected between early leptin concentrations and the amount of these tissues. Leptin concentrations measured just before slaughter were related to all fat tissues except the pelvic and intramuscular fat. Among the visceral fat depots, omental fat expressed the highest correlations to leptin (r = 0.60, P < 0.001). Additionally, leptin concentrations at 35 and 40 kg live weight increased with increasing fat cell diameters (r = 0.38, P < 0.05 to r = 0.59, P < 0.001). This study indicates that leptin concentration measured in the slaughter weight range has the greatest potential to assess body fat content, whereas an earlier prediction does not seem to be feasible. Further studies should clarify if these results are reproducible for other breeds or species. 2006 Elsevier Ltd. All rights reserved. Keywords: Lambs; Leptin; Growth; Carcass composition
1. Introduction Subjective assessments or ultrasound measurements are two common methods for the in vivo evaluation of carcass merit for breeding purposes. In addition, the utilization of hormones as predictors for production characteristics has been discussed over several years. The value of an early assay based selection in the meat industry depends on region-specific breeding and marketing conditions. A model calculation for the selection of beef cattle in Australia showed that an early selection for IGF-I at weaning *
Corresponding author. Tel.: +49 345 5522332; fax: +49 345 5527106. E-mail address: [email protected] (E. von Borell). 0309-1740/$ - see front matter 2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.meatsci.2006.05.013
increased market profitability (Wood, Archer, & van der Werf, 2004). The IGF-I test has been commercially used since 2004 in Australia’s cattle performance testing (Graser, 2004). Leptin is another potential hormone for an assay based prediction of growth or carcass composition. It is released mainly from the adipose tissue and correlates with body fat. Geary et al. (2003) recommend leptin as an additional index for fat content in beef. The accuracy for the estimation of carcass fat in lambs by plasma leptin concentrations at the time of slaughter was similar to ultrasound fat thickness measurements (Altmann, Sauerwein, & von Borell, 2005). However, the relationships between leptin concentrations in early life and final production characteristics are poorly understood. Kawakita et al. (2001) considered
M. Altmann et al. / Meat Science 74 (2006) 600–604
the correlation between early leptin concentration and final backfat thickness as being too low for a prediction of adiposity in steers. The aim of the present study was to determine the relationship of leptin plasma concentrations during growth to carcass composition and daily gain of lambs with special emphasis on size and cellularity of several fat depots. 2. Materials and methods 2.1. Animals Intact male East Frisian Milk sheep (N = 15) and Blackheaded Mutton · East Frisian Milk sheep (N = 15) lambs were used in this study. The lambs were born in late winter and were naturally reared by their dams. They were grouphoused in deep litter pens and had ad libitum access to concentrates (10.2 MJ NE/kg) and hay. The lambs were weighed weekly. The blood-sampling period began at 20.0 (SEM 0.18) kg and ended at 40.5 kg (SEM 0.23) which corresponded to a mean age of 38.5 (SEM 1.7) and 97.1 (SEM 3.1) days, respectively. Weaning occurred on two fixed days when lambs averaged 60 days of age. The mean live weight at weaning was 27.6 kg. When the lambs reached the final weight of 40 kg they were feed deprived for 24 h immediately after the last blood collection to reduce the gut fill before slaughter. Perirenal, pelvic, intestinal, and omental fat was weighed and the sum of these tissues was designated as visceral fat. The left carcass side was dissected into lean, subcutaneous and intermuscular fat and bones. These tissue masses were multiplied by 2 for the calculation of the total carcass composition. Furthermore, the intramuscular fat content was analyzed in a sample of the M. longissimus dorsi between the 13th and 14th rib by n-hexane extraction (Soxtherm 406, Gerhardt Apperate GmbH & Co. KG, Bonn, Germany). Samples of subcutaneous and perirenal fat were collected and frozen at 20 C for the determination of the cell diameters.
601
2.3. Measurement of fat cell diameters The samples were sectioned in 20 lm thick slices on a Microtome-Cryostat and fixed in formol calcium for 5 min. The slices were stained with haemalaun and fat was removed by xylen for 30–60 s. The diameter was measured in 300 cells for each sample using a light microscope with a 4 · 10 objective (Labophot-2, NIKON, Japan) connected to a 3CCD color video camera (DXC 9100P, SONY, Japan). Fat cell diameters were analyzed with an image analysis program (Lucia-G, version 4.8, NIKON, Japan). 2.4. Data analysis Data from purebred and crossbred lambs were combined for the statistical analysis because genotype did not influence hormone concentrations and performance traits. Hormone concentrations in lambs of different live weight were analyzed by ANOVA and Duncan’s multiple comparison post hoc tests. The relationships between average daily gain, carcass composition and hormone concentrations were described by Pearson correlations and regression equations. 3. Results Lambs in this study realized a high level of average daily gain (Table 1) from birth to slaughter which is comparable to performance test results for East Frisian Milk sheep in Germany. The mass of the various fat tissues showed higher coefficients of variation (19.0–39.6%) than lean mass (7.1%). Leptin concentrations in plasma rise with increasing live weight during growth and reached the highest level at 40 kg live weight (Fig. 1). The effect of live weight was significant (P < 0.001). A negative relationship between leptin and average daily gain was observed at 30 and 35 kg live weight (Table 2).
2.2. Blood sampling and hormone assays Blood samples were drawn at 20, 25, 30, 35, and 40 kg live weight. Five blood samples were collected from each lamb between 9.00 h and 11.00 h at 30 min intervals and then centrifuged. In order to obtain a mean hormone concentration over 2 h, sera were pooled for each animal, aliquoted and stored at 20 C until analyzed. Hormone concentrations in pooled samples have been shown to adequately represent those obtained from serial samples (Tiwary, 1986). Plasma leptin concentrations were analyzed with a specific enzyme immunoassay that has been validated for use in several ungulate species, including ovine samples (Sauerwein, Heintges, Hennies, Selhorst, & Daxenberger, 2004). The intra-assay coefficient of variation was 6.3%, the inter-assay coefficient of variation was 13.9%, and the limit of detection was 0.3 ng/ml.
Table 1 Average daily gain and carcass composition of experimental lambs
Average daily gain, g/day Total dissectible fata, g Visceral fatb, g Intestinal fat, g Omental fat, g Perirenal fat, g Pelvic fat, g Carcass fat, g Intermuscular fat, g Subcutaneous fat, g Intramuscular fat, g/100 g of M. longissimus Lean (g) Perirenal fat cell diameter (lm) Subcutaneous fat cell diameter (lm) a b
Mean
SEM
CV (%)
373 3547 912 344 344 193 31 2635 1773 862 1.6 12033 57.0 66.1
9.4 129.2 33.7 11.9 16.5 9.9 2.2 106.7 67.5 42.3 0.1 155.4 1.2 1.7
13.9 19.9 20.2 19.0 26.3 28.1 39.6 22.2 20.9 26.8 32.7 7.1 11.6 13.8
The sum of visceral and carcass fat. The sum of intestinal, omental, perirenal, and pelvic fat.
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Leptin correlated to the visceral fat at all live weights except 20 kg, while the correlations to carcass fat were significant only at 40 kg live weight. Among the visceral fat depots, pelvic fat showed no correlations (P > 0.05) to leptin concentrations. The correlations to omental and perirenal fat tended to be stronger as live weight increased. No correlations were observed between plasma leptin and lean mass. The relationships between fat cell size and plasma leptin measured during growth are represented in Table 3. Leptin was correlated more strongly to the fat cell diameter in perirenal fat than to the subcutaneous fat. While no associations could be found between fat cell diameter and leptin measured in lambs of 20–30 live weight, the leptin concentrations at 35 and 40 kg live weight correlated to the mean fat cell diameter in a range of r = 0.38–0.59 (P < 0.05– 0.001). The correlations in Table 2 indicate, that total dissectible fat could be best predicted by leptin concentrations at
Table 3 Correlations between leptin concentrations during growth and fat cell diameters of subcutaneous and perirenal fat Growth period (kg live weight)
Perirenal fat
20 25 30 35 40 * ** ***
Subcutaneous fat
0.08 0.16 0.25 0.59*** 0.57***
0.05 0.01 0.02 0.38* 0.49**
P < 0.05. P < 0.01. P < 0.001.
40 kg live weight. The regression equation fitted for 40 kg live weight is represented in Fig. 2. The R2 value was 0.24. 4. Discussion The present study focused on the value of leptin as a potential predictor for final performance traits in growing
6.0
c 5500
5.0
b
Total dissectible fat, g
Leptin, ng/ml
5.5
4.5 4.0 3.5
a a a
3.0 2.5 20
25
30
35
40
5000 4500 4000 3500 3000
2000
Live weight, kg Fig. 1. Leptin concentrations during the growth period of 20–40 kg live weight (means and standard error of means, means with different letters differ significantly, P < 0.05).
y = 2183 + 258 * x; P < 0.01
2500
R2 = 0.24; RMSE = 629 4
3
5
6
7
8
9
Leptin, ng/ml Fig. 2. Relationship between leptin concentration at 40 kg live weight and total dissectible fat.
Table 2 Correlations between leptin measured at different live weights and average daily gain and tissue masses Growth period (kg live weight) 20 Average daily gain, g/day Total dissectible fata, g Visceral fatb,g Intestinal fat, g Omental fat, g Perirenal fat, g Pelvic fat, g Carcass fat, g Intermuscular fat, g (M. longissims) Subcutaneous fat, g Intramuscular fat, g/100 g of M. longissimus Lean, g a b * ** ***
The sum of visceral and carcass fat. The sum of intestinal, omental, perirenal, and pelvic fat. P < 0.05. P < 0.01. P < 0.001.
0.21 0.09 0.06 0.09 0.03 0.09 0.16 0.09 0.14 0.01 0.15 0.23
25 0.10 0.26 0.41* 0.36* 0.35* 0.36* 0.10 0.19 0.18 0.18 0.06 0.19
30 0.56** 0.22 0.43* 0.43* 0.38* 0.29 0.02 0.13 0.09 0.19 0.32 0.34
35 0.61** 0.26 0.54** 0.27 0.63*** 0.46** 0.06 0.14 0.08 0.23 0.31 0.00
40 0.19 0.49** 0.58** 0.40* 0.60*** 0.46** 0.11 0.41* 0.36* 0.45* 0.19 0.04
M. Altmann et al. / Meat Science 74 (2006) 600–604
lambs and examined the relationships of average daily gain and body composition to hormone concentrations measured at several times before slaughter. The study started at 20 and ended at 40 kg live weight. This growth phase corresponds to the length of lamb performance testing in Germany and 40 kg is a common live weight for slaughter lambs in some European countries. A hormonal indicator should be applicable at 20 kg live weight or earlier for preselection or at 40 kg live weight for the prediction of carcass value in lambs kept for breeding purposes. Plasma leptin did not change significantly between 20 and 30 kg live weight within the range in which weaning occurred. However, it increased at 35 kg until 40 kg. It is assumed that this elevation was caused by a progressive increase of fat accretion. Leymaster and Jenkins (1993) found in crossbred lambs on an intensive production system an accelerated deposition of kidney-pelvic and carcass fat with advanced live weight, corresponding with the weight range in the present study. To our knowledge, the relationship of early plasma leptin concentrations to carcass composition at slaughter was only reported in one study with cattle (Kawakita et al., 2001). The authors found in steers which were fed a diet of 16% crude protein a correlation of r = 0.59 between early leptin concentrations and backfat thickness whereas no correlation existed in steers fed with 12% crude protein. The negative correlation to average daily gain at 30 and 35 kg live weight in our study implies that slow growing lambs had high leptin concentrations. This could be explained by the inhibitory effect of leptin on feed intake and the stimulatory influence on energy expenditure (Clarke, Tilbrook, Turner, Doughton, & Goding, 2001; Henry et al., 1999; Morrison et al., 2001; Tokuda, Matsui, Ito, Torii, & Yano, 2000). No correlations to average daily gain were found at lower live weights (20 and 25 kg) and thus leptin cannot be regarded as a suitable predictor for early growth rate selection. That goes for the prediction of carcass fat, too; because no relationships were detected between early leptin concentrations and the amounts of these fat tissues. Only the visceral fat correlated to the leptin concentrations from 25 kg live weight on. All fat tissues except the pelvic and intramuscular fat were significantly related with leptin concentrations at slaughter. Furthermore, a great number of other studies in several species (Altmann, Sauerwein, & von Borell, in press; Berg et al., 2003; Daniel et al., 2002; Ehrhardt et al., 2000; Geary et al., 2003; Yamada, Kawakami, & Nakanishi, 2003) indicate that several body fat tissues are highly associated with blood leptin concentration measured at the same time as body fat. A correlation of 0.68 between leptin and body fatness was found in ewes that varied widely in the deposition of fat tissue (Delavaud et al., 2000). In a previous study, we observed correlations from 0.40 to 0.64 between blood leptin concentrations and the mass of several fat depots in lambs of 35 or 45 kg live weight, respectively (Altmann et al., 2005).
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Plasma leptin concentration is influenced by the amount of fat reserves and the expression of leptin in the adipocytes (Ehrhardt, Slepetis, Bell, & Boisclair, 2001; Yuen et al., 2002). Leptin mRNA correlated positively to the adipocyte volume in humans (Oberkofler et al., 1997), rats (Villafuerte et al., 2000), mice (Guo, Halo, Leibel, & Zhang, 2004) and steers (Yang et al., 2003). Because fat cell diameter in the subcutaneous fat in our study was higher than in perirenal fat and the mass of subcutaneous fat was 4-fold higher than the perirenal fat mass, we expected a stronger correlation of plasma leptin to the subcutaneous fat mass than to the perirenal fat. However, this was not observed. Omental fat contributed to 9.7% of total fat, but to a much higher proportion of 36% to the variation of plasma leptin. This indicates a high expression of leptin in omental fat. Another study in steers showed no difference in correlations between leptin and visceral, subcutaneous, and intermuscular fat (Yamada et al., 2003). The correlations between leptin and body composition indicate that plasma leptin concentration at the end of the fattening period (40 kg live weight) is the most suitable predictor of body fat. It explained 24% of the variance in total dissectible fat. Similar results were obtained by Altmann et al. (2005) for the prediction of carcass fat by leptin concentrations measured before slaughter in lambs of 35 and 45 kg live weight. Our study confirmed the lack of a correlation between plasma leptin and lean mass as leptin is released from adipose tissue and can therefore not be considered as a predictor for lean tissue. 5. Implications Our data suggest that plasma leptin cannot be recommended as an early predictor of total fat in lambs. However, it has the potential to serve as a second stage selection criterion in breeding rams at the weight range of slaughter lambs. Further studies should clarify if these results are applicable to other breeds or species. References Altmann, M., Sauerwein, H., & von Borell, E. (2005). Relationship between plasma leptin concentrations and carcass composition in fattening mutton: a comparison with ultrasound results. Journal of Animal Physiology and Animal Nutrition, 89, 326–330. Altmann, M., Sauerwein, H., & von Borell, E. (in press). The relationships between leptin concentrations and body fat reserves in lambs are reduced by short-term fasting. Journal of Animal Physiology and Animal Nutrition. Berg, E. P., McFadin, E. L., Maddock, K. R., Goodwin, R. N., Baas, T. J., & Keisler, D. H. (2003). Serum concentrations of leptin in six genetic lines of swine and relationship with growth and carcass characteristics. Journal of Animal Science, 81, 167–171. Clarke, I. J., Tilbrook, A. J., Turner, A. I., Doughton, B. W., & Goding, J. W. (2001). Sex, fat and the tilt of the earth: effects of sex and season on the feeding response to centrally administered leptin in sheep. Endocrinology, 142, 2725–2727. Daniel, J. A., Whitlock, B. K., Baker, J. A., Steele, B., Morrison, C. D., Keisler, D. H., et al. (2002). Effect of body fat mass and nutritional
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