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J. Anim. Sci. 2003. 81:432-440
© 2003 American Society of Animal Science

Lipogenic enzyme activities in different adipose depots of Pirenaican and Holstein bulls and heifers taking into account adipocyte size

P. Eguinoa*, S. Brocklehurst{dagger}, A. Arana*,1, J. A. Mendizabal*, R.G. Vernon{ddagger} and A. Purroy*

* Departamento Producción Agraria, Universidad Pública de Navarra, Campus de Arrosadía,31006 Pamplona, Spain and and {dagger} Biomathematics and Statistics Scotland and and {ddagger} Molecular Homeorhesis Group, Hannah Research Institute, Ayr KA6 5HL U.K.

1 Correspondence:
phone: 34-948-169116; fax: 34-948-169732; E-mail:
aarana{at}unavarra.es.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Implications
 Literature Cited
 
The effects of sex, genotype, and adipose depot on lipogenic enzyme activity have been investigated in Holstein and Pirenaican bulls and heifers, taking into account differences in adipocyte size. Fifteen Pirenaican bulls and 15 heifers and 15 Holstein bulls and 13 heifers were fattened until slaughter (12 to 13 mo old and 450 to 500 kg of body weight). During the fattening period, animals had ad libitum access to commercial concentrates and straw. The 10th rib was dissected to determine the fat content. Adipocyte size and activities of the following lipogenic enzymes were determined: glycerol 3-phosphate dehydrogenase, fatty acid synthase, nicotinamide adenine dinucleotide phosphate (NADP)-malate dehydrogenase, glucose 6-phosphate dehydrogenase, and NADP-isocitrate dehydrogenase, in the omental, perirenal, subcutaneous, and intermuscular adipose depots, respectively. Because adipocyte mean cell volume varied with sex, breed, and depot, regression analyses of loge activity per cell and loge cell volume were used to compare activities per unit volume. Sex, breed and depot had no effect (P > 0.05) on the gradients of regressions, which did not differ significantly from 1. Thus, activity per unit volume did not vary with cell size. Consequently, sex, breed, and depot effects on the regression analyses were equivalent to effects on activity per unit volume. Females had greater amounts of fat in the 10 th rib (P < 0.001), larger adipocytes (P < 0.001) and, in general, greater (P < 0.05) lipogenic activity per cell, even when adjusted for cell size, than males. These findings suggest that differences in adiposity between sexes are mainly due to females having a greater capacity for lipid synthesis, and hence, hypertrophy, than males. When adjusted for differences in carcass weight, Holsteins had larger adipocytes than Pirenaicans. The abdominal depots, omental and perirenal, had a greater adipocyte size (P < 0.001) and, in general, greater lipogenic enzyme activities per cell (P < 0.05) than the subcutaneous and intermuscular carcass depots. However, when activity per cell was adjusted for cell size, subcutaneous depots had greater fatty acid synthae, glucose 6-phosphate dehydrogenase, and NADP-malate dehydrogenase activities than omental and perirenal, indicating that other factors such as nutrient supply may restrict hypertrophy of carcass adipocytes.

Key Words: Adipocytes • Adipose Tissue • Cattle • Genotypes • Lipogenesis • Sex


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Implications
 Literature Cited
 
Adipose tissue develops in a number of depots throughout the body, some in the abdominal cavity (e.g., omental [OM], perirenal [PR]), some subcutaneously, and some in the musculature. Adipose tissue within the musculature, especially marbling fat, is important for meat quality in traditional markets, but most adipose tissue is not required for consumption and is discarded, creating an undesirable cost for the producer.

Adipose tissue mass accrues through both adipocyte hyperplasia and hypertrophy (Hood, 1982). Hyperplasia occurs mostly in young animals, whereas hypertrophy occurs predominantly in older animals (Vernon 1986; Robelin and Castiella, 1990). Hypertrophy is due to adipocytes accumulating triacylglycerols; these are formed by esterification of glycerol 3-phosphate and fatty acids. Glycerol 3-phosphate is synthesized from glucose in adipocytes, whereas fatty acids may be synthetized de novo within adipocytes or obtained from blood triacylglycerols.

The various adipose tissues do not develop at the same time, neither do they have the same rate of growth; thus, adipocyte size varies between depots. Furthermore, development of adiposity is influenced by sex and breed (Hood and Allen, 1975; Robelin, 1981, 1986; Vernon, 1986). Lipogenic activity varies with adipocyte size in cattle (Hood, 1982; Rule et al., 1987), and so may contribute to depot-specific differences in adipocyte volume. In the present study, the activity of five lipogenic enzymes was investigated for adipose tissue from four different depots in bulls and heifers of Pirenaican and Holstein breeds, slaughtered at market age and weight, taking into account differences in adipocyte size.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Implications
 Literature Cited
 
Animals and Feeding

Thirty Pirenaicans (15 males and 15 females) and 28 Holsteins (15 males and 13 females) were used. Pirenaica is a Spanish breed used for meat production; Holstein is the principal breed used for milk production, representing about half the total bovine population (60% in the European Union and 45% in Spain), with about 50% of beef meat also coming from this breed (European Commission, 1999). Pirenaican calves were weaned at 5 mo of age. Before weaning, calves consumed mother’s milk and precalving concentrates (89.3% DM, 6.53% ash, 18.00% CP, 5.58% crude fiber, 36.78% NDF, 8.68% modified ADF, 3.2% ether extract, 30% starch, 0.80% Ca, 0.60% P, 3.36 Mcal of ME/kg of DM). After weaning, animals had ad libitum access to commercial concentrate (89.8% DM, 6.90% ash, 16.28% CP, 5.86% CF, 34.20% NDF, 9.03% modified ADF, 5.4% ether extract, 32% starch, 0.69% Ca, 0.55% P, 3.28 Mcal of ME/kg of DM), and straw. Holstein calves were weaned at 2 d of age after taking colostrum, after which they were fed artificial milk and concentrates, as were Pirenaican calves until 4 mo of age. After this time, they were fed the same regimen as the weaned Pirenaican cattle. The care and use of the animals followed European guidelines (86/609/CEE and BOE 18/March/1998, Spain).

Slaughter and Sample Collection

Animals were slaughtered at 12 to 14 mo of age in an abattoir. Within 10 min after slaughter, 15-g fat tissue samples were taken from OM, PR, s.c., and i.m. depots. Five grams was frozen at -40°C for later analyses of lipogenic enzyme activities, 0.5 g was placed in glass test tubes containing 10 mL of Tirode solution (0.15 M NaCl; 6 mM KCl; 2 mM CaCl2; 6 mM glucose; 2 mM NaHCO3), pH 7.62, at 39°C for measurement of adipocyte size. The remaining tissue was frozen and used to determine lipid content. The OM sample was obtained from adipose tissue near the omasum, the PR sample from beside the left kidney, the s.c. sample from the region adjacent to the sternum, and the i.m. sample from the depot between the sternum and pectoral muscles. Forty eight hours after slaughter, the 10th rib was removed, weighed, and stored at 4°C for analysis. Fat content of the 10th rib was determined gravimetrically after dissection.

Adipocyte Size

Samples of fat for adipocyte sizing (0.5 g) were minced and fixed with 2% osmiun tetroxide solution by the technique of Hirsch and Gallian (1968). After fixation, the diameter of approximately 200 adipocytes from each depot was measured using an image analyzer program (Biocom, Photometric Image Analysis System, Les Ulis, France), as described by Mendizabal et al. (1997). Corresponding cell volumes were calculated from the diameters, and these values were used to calculate the average cell volume.

Enzyme assays

Adipose tissue samples from OM, PR, s.c., and i.m. depots for enzyme assays were homogenized in ice-cold STEG buffer (0.3 M sucrose, 30 mM trizma base, 1 mM EDTA, 1 mM glutathione, pH 7.4) in a 1:4 (wt/vol) ratio using a Sorvall Omni-Mixer homogenizer (OMNI Int., Waterbury, CT; 10 s at 50,000 rpm, three times); the tissue was kept ice-cold. Homogenates were filtered (20-µm pore diameter) to eliminate residual lipids and connective tissue. Homogenates were then centrifuged for 10 min at 4,500 x g. They were again filtered (0.45-µm pore diameter) and centrifuged for 10 min at 15,000 x g in a Heraeus Varifuge 20 RS (Heraeus Sepatech GmbH, Osterode, Germany). After the second centrifugation, the intermediate phase or supernatant (cytoplasmatic soluble fraction) was collected with a Pasteur pippette and filtered (0.45-µm cellulose filter). The final extracts were placed in 1.5-mL polypropylene tubes (Eppendorf AG, Hamburg, Germany; one tube per enzyme analysis) and frozen at -40°C until the day of enzyme assay. Preparatory studies showed that for the five enzymes assayed, activities of frozen and fresh extracts were similar.

The following lipogenic enzymes were assayed: Glycerol 3-phosphate dehydrogenase (G3PDH; EC 1.1.1.8) (Wise and Green, 1979), an enzyme involved in glycerol 3-phosphate synthesis from the glucose; fatty acid synthase (FAS; EC. 2.3.1.85) (Halestrap and Denton, 1973), involved in de novo fatty acid synthesis; and nicotinamide adenine dinucleotide phosphate-malate dehydrogenase (MD; EC 1.1.1.40) (Ochoa, 1955), glucose 6-phosphate dehydrogenase (G6PDH; EC 1.1.1.49) (Glock and McLean, 1953), and reduced nicotinamide adenine dinucleotide phosphate (NADPH)-isocitrate dehydrogenase (ICDH; EC 1.1.1.50) (Plaut, 1962), enzymes involved in reduced nicotinamide adenine dinucleotide phosphate supply for de novo fatty acid synthesis. Enzyme activities were determined by measuring the change in absorbance over time at 37°C. Reactions were linear with respect to time over the period of assay and proportional to the amount of extract added.

To calculate enzyme activity per 106 cells, the number of cells per gram of adipose tissue was calculated using the average volume of adipocytes, the lipid content of the tissue (determined by the Soxhlet method according to ISO 1443-1973), and the density of the lipid (0.915 g/mL).

Statistical Analyses

Analysis of variance was used to investigate the effects of sex, breed, and adipose depot on the data. To satisfy the conditions of normality and homogeneous variantes, it was necessary to loge transform all the variables, apart from the amount of adipose tissue, before applying the ANOVA. The following two models were applied: 1) for the amount of adipose tissue in the 10th rib, an ANOVA with fixed effects of sex and breed and the interaction between sex and breed and 2) for the loge of the adipocyte diameter, the loge of the volume, and the loge of the lipogenic enzyme activities per cell, an ANOVA with fixed effects of sex, breed, and depot and all two- and three-way interactions.

When main effects or interactions were P < 0.05, the exact nature of the differences between means was investigated using Tukey’s multiple comparison test.

There were highly significant effects of sex, breed, and depot on diameter and volume of adipocytes, and these effects were similar to those seen for the enzyme activities per cell. A further analysis was carried out to assess whether the effects seen in the activities per cell were due to different cell sizes or if there were genuine effects to be seen after taking into account cell size. This was done by using multiple linear regression analyses in which the loge of the lipogenic enzyme activities was modeled as a function of the loge of the adipocyte volume and of breed, sex, and depot and all interactions. Although interactions of all the factors with volume were investigated, they were omitted from the final models because they were not significant. Thus, this analysis included the possibility that the gradient, as well as the intercept, of the linear relationship between activity and volume (on the loge transformed scales) might vary with the factors and their interactions, but according to statistical tests, the gradient did not vary significantly for any of the five lipogenic enzyme activities. This means that analysis of the "group" effects (i.e., effects of breed, sex, depot, and all interactions thereof) are analyses of the differences between the intercepts of the lines, which are independent of volume. These group effects, therefore, will be essentially equivalent to group effects on the activity per unit volume.

Because activity per cell is calculated based on activity per unit volume together with cell volume, a significant relationship between activity per cell and cell volume with a gradient of 1 would be expected by definition, provided there is no relationship between activity per unit volume and cell volume. Because the gradients for all five enzymes were not significantly different from 1, this relationship was not of interest. The focus of the investigation was instead on the assessment of the "group" effect after allowing for any relationship that did exist, whatever its nature.

Genstat 5, release 4.1 (VSN International Ltd., Herts, U.K.), was used for all the statistical analyses, except for the Tukey test, for which Minitab 12 (Mitlab Inc., State College, PA) was used.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Implications
 Literature Cited
 
Mean age and hot carcass weight (HCW) at slaughter were 399 ± 14 d and 340.9 ± 7.6 kg for Pirenaican males, 371 ± 6 d and 247.6 ± 9.4 kg for Pirenaican females, 375 ± 4 d and 260.6 ± 5.4 kg for Holstein males, and 375 ± 5 d and 229.2 ± 6.5 kg for Holstein females. The HCW of heifers was less than that of bulls (P < 0.001).

Table 1Go shows the results and ANOVA for the amount of fat in the 10th rib, the diameter and volume of adipocytes, and the enzyme activities in various adipose depots for the two sexes of each breed. The 10th rib was chosen because in cattle, the rib composition (fat, muscle, bone) is representative of the whole carcass composition (Geay and Beranger, 1969).


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  		Table 1. Amount of adipose tissue on the 10th rib, diameter and volume of adipocytes and enzyme activity of glycerol 3-phosphate (G3PDH), fatty acid synthetase (FAS), nicotinamide adenine dinucleotide phosphate (NADP)-malate dehydrogenase (MD), glucose 6-phosphate dehydrogenase (G6PDH), and NADP-isocitrate dehydrogenase (ICDH) in different depots of Pirenaica (PIR) and Holstein (HOLS) males (M) and females (F)a
 
There was sex difference in the amount of adipose tissue around the 10th rib (Table 1Go), with a greater amount in females than in males (313.6 vs. 168.9 g, P < 0.001, representing 24.8 vs. 12.9% of rib weight, respectively, P < 0.001). When carcass weight was used as a covariable, females still had more adipose tissue in the 10th rib than did males (P = 0.004).

There were interactions between sex and breed (P = 0.01) and sex and depot (P < 0.001) for adipocyte volume (Table 1Go). Females had larger adipocytes than males, regardless of breed (P < 0.05), except for the i.m. depot, where no sex difference was apparent. Holstein females had larger adipocytes than Pirenaican females (P < 0.01), but there were no differences in adipocyte volume between males of the two breeds. The volume of adipocytes differed between adipose depots (P < 0.01): OM = PR > s.c. > i.m. This ranking was the same for both sexes, but differences were more accentuated in females than in males. As Pirenaican bulls were markedly heavier than other animals, results were reanalyzed including carcass weight as covariate. The sex x breed interaction was no longer significant, and Holsteins had larger adipocytes than Pirenaicans regardless of sex (P < 0.001). There was a significant effect of sex (P < 0.001), with females having larger adipocytes than males for all depots including i.m., whereas the ranking of depots by size was unchanged (OM = PR > s.c. > i.m.).

Enzyme Activities Per Cell

Enzyme activities per cell were analyzed initially without taking differences in cell size into account (Table 1Go). For G3PDH, there were no interactions (P> 0.05), but there were effects of sex, breed, and depot (Table 1Go). Females had a higher G3PDH activity than males (P < 0.001), and Holsteins had a higher (P = 0.004) activity than Pirenaicans. Differences (P < 0.001) between depots were observed, and G3PDH activity ranked: OM = PR > s.c. > i.m. (P < 0.01).

There was a significant (P < 0.001) interaction between sex and breed for FAS (Table 1Go). Holstein females had greater FAS activity than Holstein males (P = 0.003), whereas in Pirenaican animals there were no sex differences in FAS activity. There was greater FAS activity in adipocytes from male Pirenaican than male Holsteins (P = 0.004), but no difference (P > 0.05) in activity between genotypes was found with females. There was also a sex x depot interaction (P < 0.001) for FAS activity. For females FAS activity ranked: PR = OM > s.c. > i.m. (P < 0.001). Depot differences were smaller for males than females, and FAS activity ranked: PR = OM > i.m. (P < 0.001); s.c. activity was intermediate between OM and i.m., and did not differ significantly from either.

There was no effect (P > 0.05) of breed or any interactions between breed and sex or breed and depot for MD, but there was an interaction (P < 0.001) between sex and depot (Table 1Go). The MD activity of OM, PR, and s.c. depots was greater in adipocytes from females than males (P < 0.01), whereas for i.m., there was no sex difference in activity. No depot-specific differences in MD activity were detected in males, but in females, MD activity ranked OM = PR = s.c. > i.m. (P < 0.01). In contrast to MD, G6PDH showed a sex x breed interaction (P = 0.003); females had greater G6PDH activity than males (P < 0.001), and this sex difference was greater in Pirenaicans than in Holsteins. Furthermore, there was no breed effect in males, whereas G6PDH activity was greater in Pirenaican than Holstein females (P = 0.04). There was also a depot effect (P < 0.001) for G6PDH, with activity ranking OM > s.c. > i.m. (P < 0.01), and PR activity being intermediate between OM and SC activities and not differing significantly from either.

The ICDH activity showed sex x breed (P = 0.003) and sex x depot (P = 0.03) interactions. The ICDH activity was greater in adipocytes from females than males (P < 0.01), and this difference was greater in Pirenaicans than Holsteins. Activity of ICDH was greater in Holstein than Pirenaican males (P < 0.001), but no breed effect was apparent for females. The ICDH activity of females was greater than that of males for adipocytes from OM, PR, and s.c. depots (P < 0.001), but there was no sex difference in ICDH activity for i.m. adipocytes. For females, ICDH activity ranked OM > s.c. > i.m. (P < 0.05), whereas for males, OM > s.c. = i.m. (P < 0.01); for both males and females, ICDH activity of PR adipocytes was intermediate between that of OM and SC adipocytes and did not differ (P > 0.05) from either.

Analysis of Enzyme Activities Per Cell Taking into Account Cell Size

Becasue adipocyte mean cell volume varied significantly with sex, breed and depot, some differences in enzyme activity shown in Table 1Go may simply reflect differences in cell size. Regression analysis showed that all five enzyme activities increased (P < 0.001 in each case) with increasing adipocyte volume (Table 2Go). However, the gradients in all cases were not different (P > 0.05) from 1, and so this merely reflects the fact that, by definition, activity per cell is related to cell volume. This also indicates, however, that there is no significant relationship between activity per unit volume and cell size.


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  		Table 2. P-values for regression analyses fitted in the order adipocyte volume, S, B, D, S x B, S x D, B x Da
 
There were no interactions (P > 0.05) between cell volume and sex, or breed or depot; that is, plots of lines of best fit from the regression analyses comprised a series of parallel lines on the loge-transformed scales, so a significant difference between regressions indicates a difference in activity per cell (and thus per unit volume), independent of cell size. When values were adjusted for cell volume, not only was there no significant interaction between sex, breed, and depot, but now there was no interaction (P > 0.05) between sex and depot for any of the five enzymes.

The regression equations were used to derive the graphs of Figure 1Go, which show the relationship between activity per cell and adipocyte size for each enzyme. The graphs show that for a given cell size, activity can vary from threefold (FAS) to 17-fold (G6PDH), depending on depot, sex, and breed. For all five enzymes studied, there was either a sex effect (G3PDH, MD) (P < 0.001), with females having greater activity for a given cell size than males or a sex x breed interaction (FAS, P = 0.01; G6PDH, P < 0.001; ICDH, P < 0.001) (Table 2Go). For G6PDH, activities ranked: Pirenaican female > Holstein female > Holstein male = Pirenaican male, whereas for ICDH they ranked: Pirenaican female = Holstein male = Holstein female > Pirenaican male. Conversely, for FAS, activities ranked: Pirenaican male > Holstein male = Pirenaican female = Holstein female. Depot effects were found for FAS, MD and G6PDH, and there was a depot x breed interaction for G3PDH (Table 2Go). For MD, activities ranked: s.c. = i.m. > OM = PR. For FAS and G6PDH, activities ranked: s.c. > PR = OM and s.c. > OM = PR, respectively, with i.m. intermediate between SC and PR (FAS) and OM (G6PDH). The only depot effect with G3PDH was found in Holsteins, for which activity ranked: s.c. = PR = OM > i.m.. There were no depot effects with G3PDH in Pirenaicans or for ICDH in either breed (Table 2Go). Holsteins had a greater (P = 0.001) G3PDH activity in SC adipocytes than Pirenaicans, but there were no breed differences for the other depots.



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  		Figure 1. Plots of regression models for loge of enzyme activities: (a) glycerol 3-phosphate (G3PDH), b) fatty acid synthetase (FAS) c) malate dehydrogenase (MD) d) glucose 6-phosphate dehydrogenase (G6PDH), and e) NADP-isocitrate dehydrogenase (ICDH) with terms loge of adipocyte volume, breed, sex, depot, breed x depot, sex x breed included. OM = omental (———), PR = perirenal (– – –), s.c. = subcutaneous (— — —), i.m. = intermuscular (••••••). M-PIR = Pirenaican males ({square}), F-PIR = Pirenaican females ({circ}), M-HOLS = Holstein males ({blacksquare}), F-HOLS = Holstein females (•).

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Implications
 Literature Cited
 
Of the five lipogenic enzymes studied, G3PDH and FAS are important enzymes of fatty acid esterification and de novo synthesis, respectively, whereas G6PDH, MD, and ICDH are involved in the synthesis of NADPH for de novo fatty acid synthesis. A relationship between adipocyte size and lipogenesis (Hood, 1982; Rule et al., 1987; de la Hoz and Vernon, 1993), lipogenic enzyme activity (Vernon et al., 1987), lipoprotein lipase and acetyl CoA carboxylase gene expression (Barber et al., 2000), and catecholamine-stimulated lipolysis (Vernon and Finley, 1985; Vernon et al., 1995) has been found for adipocytes from ruminants. It is probable that large adipocytes are large because they have greater enzyme activity; however, for some enzymes, it is possible that the greater activity is the product rather than the cause of a large cell. To try to address these possibilities, the relationship between activity and adipocyte size was explored by regression analysis in the present study, which allowed comparisons of activities independent of cell size and which shows that the relationship between activity and cell size is complex, and that activity can vary with sex, breed, and depot independently of cell size.

Differences Between Sexes

Female cattle accumulate more fat than males (Berg and Butterfield, 1976; Robelin, 1978; Kempster, 1980 Kempster, 1981). Heifers have larger PR and OM adipocytes than bulls of the same BW (Beranger and Robelin, 1977; Robelin and Daericke, 1980; Robelin and Agabriel, 1986). The present study shows that s.c. and i.m. adipocytes are also larger in heifers than bulls. Thus, sex differences in adiposity are at least partly due to differences in hypertrophy. This is supported by the similarity between the ratios for the amount of fat in the 10th rib of males to females (1:1.6 and 1:2.0 for Pirenaicans and Holsteins, respectively) and ratios for adipocyte mean cell volumes of males to females (Table 3Go).


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  		Table 3. Ratios for amount of adipose tissue on 10th rib and adipocyte size of males and females of both breeds, and ratios of Pirenaican and Holstein of both sexesa
 
The larger adipocytes in females than in males were accompanied by higher activities of G3PDH, G6PDH, MD, and ICDH. In addition, FAS activity of female Holsteins (but not Pirenaicans) was greater than that of males. Sidu et al. (1973) and Mendizabal et al. (1997) also found greater G3PDH activity in adipocytes from ewes than rams, along with a greater amount of fat and adipocyte size in ewes. Furthermore, regression analysis showed that, for a given adipocyte size, G3PDH, MD and G6PDH activities per adipocyte, and also ICDH activity in adipocytes from Pirenaican, but not Holsteins, were greater for adipocytes from females compared with males, which could contribute to the greater rate of hypertrophy in females. When adjusted for cell size, FAS activity was greater in adipocytes from male than female Pirenaicans, whereas for Holsteins, neither FAS nor ICDH activity showed sex differences. The reason for FAS activity being greater in male than female Pirenaicans, whereas the activities of the dehydrogenases, which supply NADPH for use in the reaction catalysed by FAS, were greater in females than males, is not clear.

Differences Between Breeds

The breeds used in the present work differ in that Holstein cattle have been selected for milk production, whereas the Pirenaica is an indigenous Spanish breed improved for beef production. At commercial slaughter weight and age, the Pirenaicans were heavier than the Holsteins and there were no significant differences between breeds in the amount of adipose tissue present in the 10th rib. However, when carcass weight is used as a covariate, the Holsteins had a greater amount of adipose tissue than the Pirenaicans. These findings agree with the results obtained by Robelin (1978) in a comparison of Holsteins with French breeds selected for beef production (Charolaise and Limousin).

When differences in carcass weight were taken into account, Holsteins had larger adipocytes than Pirenaicans; however, the reason for the larger adipocytes in Holsteins is not clear. The G3PDH activity per cell was greater in adipocytes from Holsteins than Pirenaicans (Table 1Go), but when differences in cell volume are taken into account this breed difference was still apparent for SC, but not for other depots. Furthermore, when corrected for cell size, MD showed no breed differences, whereas for the other three enzymes there were sex-breed interactions, but activities of Holsteins were either the same as, or less than, those of Pirenaicans of the same sex. The one exception was ICDH, for which Hostein bulls had a greater activity than Pirenaicans.

Differences Between Depots

The hierarchy observed in the adipocyte size: OM > PR > s.c. > i.m., was also observed by Hood and Allen (1973), Allen (1976), Schiavetta et al. (1990), and Mendizabal et al. (1999a) in studies with different cattle breeds during the fattening period, and in mature animals (Mendizabal et al., 1999b). The greater size of abdominal adipocytes could be due to hyperplasia being largely complete in these abdominal depots at an earlier age, whereas substantial hyperplasia continues in carcass depots well into the fattening period (Vernon, 1986).

In general, this hierarchy in adipocyte size between depots was accompanied by a similar hierarchy in the lipogenic enzyme activities studied. A similar relationship between adipocyte size and lipogenic flux and also enzyme activities between different depots has been observed by other authors, such as Hood (1982) and Rule et al. (1987). Thus, depot differences in adipocyte size would seem to be explained by differences in lipogenic enzyme activities. However, this would seem to be an oversimplification. Although G3PDH activity seems to vary with cell size independent of depot of origin in Pirenaicans and also in Holsteins (except for i.m. cells) for at least three enzymes involved in fatty acid synthesis (FAS, G6PDH, MD), when differences in adipocyte size are taken into consideration, activities per cell of s.c. and i.m. origin were generally greater than that of OM and PR. Because s.c. and i.m. adipocytes do not seem to catch up in size with PR and OM adipocytes as animals mature, there must be some constraint on the lipogenic capacity of s.c. and i.m. adipocytes. There are several possibilities. Fatty acids for esterification can be derived from plasma lipids by the action of lipoprotein lipase (LPL). However, in fattening sheep, at least, LPL gene expression varies with adipocyte size independent of depot (Barber et al., 2000), suggesting that differences in the ability to use plasma lipid is unlikely to be the explanation. Blood flow to carcass depots is less than that to abdominal depots in ruminants (Vernon, 1980; Barnes et al., 1983; Gregory et al., 1986); hence, nutrient supply to s.c. and i.m. depots will be less than that to OM and PR. A greater capacity for fatty acid synthesis of s.c. and i.m. compared to OM and PR adipocytes may thus be a compensatory mechanism for a reduced nutrient supply to carcass depots. If, as suggested from sheep studies, there is no equivalent compensatory increase in LPL activity in carcass depots, this would imply a relatively greater importance of de novo fatty acid synthesis compared with LPL activity as a source of fatty acids for esterification in carcass depots compared to abdominal depot; this would be consistent with earlier findings that fatty acids of dietary origin have a greater tendency to be deposited in abdominal rather than carcass depots (Christie, 1978). A further possibility is a difference in the rate of lipolysis in adipocytes from carcass and abdominal depots. Catecholamine-stimulated lipolysis—but not basal lipolysis—assessed in vitro varies with adipocyte size, at least in sheep (Vernon and Finley, 1985; Vernon et al., 1995). The relationship between lipolytic rate and cell size for different depots has not been investigated, but in adult ewes, the rate of catecholamine-stimulated lipolysis of 600- to 800-pL s.c. adipocytes was the same as that of 1,700-pL OM adipocytes (Vernon et al., 1995), suggesting that s.c. adipocytes may have a greater rate of lipolysis for their size than OM adipocytes. Thus, the greater fatty acid synthetic activity of carcass adipocytes could also help to compensate for a greater rate of lipolysis in these cells relative to abdominal adipocytes.

The regulation of adipocyte volume is complex; not only does adipocyte size vary with sex, breed, and depot, but the mechanisms responsible for these differences also vary. Thus, females have larger adipocytes than males, and abdominal adipocytes (OM, PR) are larger than carcass adipocytes (s.c., i.m.). Furthermore, the larger adipocytes of females have, in general, greater lipogenic enzyme activities than the smaller adipocytes of males, even when allowance is made for differences in cell size, providing a probable cause for the greater size of the former. By contrast, although the larger abdominal adipocytes have, in general, greater lipogenic enzyme activities than the smaller adipocytes from carcass depots, when adjusted for differences in cell size, the smaller cells from the carcass depots have the greater lipogenic activity. This implies that other factors such as blood flow or lipolytic activity are likely to be important determinants of differences in adipocyte size between depots.


   Implications
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Implications
Literature Cited
 
Devising methods to manipulate the amounts of individual adipose tissue depots is important for improving carcass composition for consumers who want leaner meat, while still providing animals with a reserve of energy. The various adipose tissue depots do not develop at the same time or rate, so it is important to elucidate the factors responsible for these differences. The lipogenic capacity of adipose tissue is thought to be a key factor and is at least partly responsible for the greater amount of fat in females than in males. However, although abdominal adipocytes are larger than carcass adipocytes and have in general a greater lipogenic activity per cell, when adjusted for differences in cell size, it is apparent that the smaller carcass cells have greater activity. This suggests that other factors such as blood flow, and hence nutrient supply, or lipolysis are important in influencing depot differences in adiposity, and so are important potential targets for manipulation.

Received for publication November 27, 2001. Accepted for publication November 13, 2002.


   Literature Cited
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Implications
 Literature Cited
 


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