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ANIMAL GROWTH, PHYSIOLOGY, AND REPRODUCTION |
* Institut National de la Recherche Agronomique (INRA), Unité Mixte de Recherches (UMR) 1079 Systèmes d’Elevage Nutrition Animale et Humaine, Saint-Gilles, 35590, France; and
High-Throughput Proteomics Platform OUEST-Genopole, 263 Avenue du Général Leclerc, Bâtiment 24, Campus de Beaulieu, Rennes, 35042, France
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Abstract |
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 Top Abstract INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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0.01), lipolysis (perilipin, P < 0.01), fatty acid oxidation (long-chain fatty-acyl CoA dehydrogenase, P < 0.01), and energy transfer (catalase, voltage-dependent anion channel 1, and electron-transfer flavoprotein, P < 0.05). In contrast, both prohibitin-1 and cell division cycle 42 homolog, with possible roles in cell growth, were up-regulated (P < 0.05) in i.m. adipocytes compared with other fat cells. Fewer differences were observed when adipocytes isolated from s.c., perirenal, and intermuscular fat tissues were compared, with a maximum of 17 spots differing significantly in abundance between perirenal and s.c. adipose tissues. The findings that proteins involved in both anabolic and energy-yielding catabolic pathways are downregulated in i.m. adipocytes compared with s.c., visceral, or intermuscular adipocytes, suggest that the metabolic activity of i.m. adipocytes is low. Thus, triggering adipogenesis rather than cell metabolism per se might be a valuable strategy to control lipid deposition in pig skeletal muscles.
Key Words: adipocyte • intramuscular fat • pig • proteome • regional difference
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INTRODUCTION |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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), there is a general consensus that i.m. fat content in contemporary lines of pigs is less than optimal for eating quality (Fernandez et al., 1999). Muscle fat content is influenced by age, sex, genotype, and environmental factors (Poulos and Hausman, 2005). These variations are mainly related to the number or the size of adipocytes clustered along myofiber fasciculi (Gondret and Lebret, 2002; Damon et al., 2006). To date, little is known about i.m. adipocyte features (Poulos and Hausman, 2005). In cattle (Miller et al., 1991) and pigs (Gardan et al., 2006), i.m. adipocytes are smaller than adipocytes originating from s.c. or perirenal fat pads. Other available data concern lipid metabolism. For example, the same authors also reported lower content of lipogenic enzymes and depressed lipolytic rate in i.m. fat cells than in s.c. adipocytes. Other adipocyte functions involving the regulation of body energy balance through the secretion of adipokines (Hauner, 2005) are poorly documented in i.m. adipocytes (Gardan et al., 2006). The recent finding that 10% of genes expressed in pig adipose tissue are of mitochondrial origin (Chen et al., 2006) might also transform our thinking about the physiological functions of adipocytes. Therefore, the identification of differentially expressed proteins between adipocytes originating from various locations might be helpful in defining the functions of adipocytes in muscle and in defining strategies to control meat lipid content independently of body fat depots. The objective of the present study was to evaluate the value of 2-dimensional (2D), fluorescent, differential gel electrophoresis (2D-DIGE) to identify proteins differentially expressed between adipocytes isolated from muscle and from various adipose tissues in the pig. |
MATERIALS AND METHODS |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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Animals and Adipose Cell Isolation
Six crossbred Pietrain x (Large White x Landrace) female pigs from different litters were weaned at 28 d. They had free access to a starter diet (19.5% CP, 6.6% fat, 2.54 Mcal/kg of NE, and 12.7 g/kg of digestible lysine, as-fed basis) from 15 d up to approximately 42 d, and to a piglet diet (18.5% CP, 2.5% fat, 2.29 Mcal/kg of NE, and 10.8 g/kg of digestible lysine, as-fed basis) until the age of approximately 77 d. Thereafter, growing animals had free access to a growing diet (17.8% CP, 4.0% fat, 2.29 Mcal/kg of NE, and 8.3 g/kg of digestible lysine, as-fed basis) and water until slaughter. All diets were purchased from Cooperl-Hunaudaye (Lamballe, France).
Pigs were killed after an overnight fast at 80 d of age (31.1 ± 0.1 kg) by exsanguination after electronarcosis. Trapezius skeletal muscles, intermuscular fat (INTER) located below the trapezius muscle, and samples of dorsal s.c. fat and of perirenal adipose tissue (PERI) were collected immediately after slaughter. Adipocytes were isolated by collagenase treatment of the isolated tissues (Etherton and Chung, 1981; Gardan et al., 2006) as follows. Minced tissues (60 g for muscle; 20 g for adipose tissues) were placed in Krebs-Ringer bicarbonate buffer (2 mL/g for muscle; 3 mL/g for s.c., INTER, and PERI) containing 3% BSA, 10 mM glucose, 1.3 mg/mL of collagenase A [Roche Applied Science, Meylan, France, (0.22 U/mg)], and antibiotics (6.25 U/mL of penicillin and 6.25 µg/mL of streptomycin; Invitrogen, Cergy-Pontoise, France). After digestion at 37°C for 45 min (muscle) to 1 h (s.c., INTER, and PERI), the material was filtered through 200-µm sterile, nylon, blutex mesh filters (Saati France, Sailly Saillisel, France) to separate adipose cells from tissue fragments. Adipocytes isolated from s.c., PERI, or INTER were allowed to float; they were rinsed 3 times with Dulbecco’s modified Eagle’s medium (5.5 mM glucose; Invitrogen) at 37°C by removing the infranatant with a plastic catheter attached to a syringe. The filtered muscle material was gently centrifuged at 100 x g for 1 min, and the resulting floating i.m. adipocytes were collected in Dulbecco’s modified Eagle’s medium. All cells were then washed twice in 10 mM Tris buffer (pH 7.0) containing 250 mM sucrose at 37°C.
An aliquot of the cell suspension (10 µL) was digitized by using a microscope (Leitz, Wetzlar, Germany) equipped with a CCD camera (CV-M90, JAI Corporation, Yokohama, Japan). Cells from each site and within each animal were kept separately. Individual diameters (µm) of adipocytes according to site and pig were then measured by using an Optimas 6.5 image analysis system (Media Cybernetics, Silver Spring, MD, http://www.mediacy.com).
Preparation of Protein Extracts
All cells (approximately 1 mL of cell suspension, corresponding to approximately 5 x 106 cells) were resuspended (vol/vol) in a lysis buffer containing 6 M urea (GE Healthcare, Orsay, France), 2 M thiourea (Sigma, Saint-Quentin Fallavier, France), 2% (wt/vol) of 3-3-cholamidopropyldimethylammoniol (GE Health-care), and 30 mM Tris (Sigma), pH 8.8. Cells were then sheared by 5 passages of a glass-glass homogenizer pestle. They were agitated by using magnetic stirrers for 30 min at room temperature before the homogenate was centrifuged at 10,000 x g for 20 min at 18°C. The supernatant below the fat cake was then collected. Lipids were further extracted by chloroform addition (vol/vol), followed by centrifugation at 3,000 x g for 10 min. Each cell homogenate was then centrifuged at 100,000 x g for 1 h at 18°C before the supernatant was collected. Proteins were enriched by using centrifugal filter devices (Centricon, Millipore, Billerica, MA), and the total protein concentration was determined by using Bradford protein reagents (Bio-Rad, Hercules, CA) and BSA as the standard (fraction V, Roche Diagnostics GmbH, Mannheim, Germany). Because of the low amounts of proteins in some samples, the analysis was carried out on 3 to 6 samples per site. Protein extracts were then stored at –75°C until used for electrophoresis.
Labeling
Proteins (50 µg) from each adipocyte extract were minimally labeled by incubation on ice with 400 pmol of the amine-reactive cyanine dyes Cy3 or Cy5 fluors (GE Healthcare) for 30 min in the dark. To create an internal standard, equal amounts of the 20 protein extracts were pooled and were then labeled with Cy2. The reaction was quenched by incubation for 10 min with 1 µL of 10 mM lysine on ice in the dark.
2D Electrophoresis
Preliminary experiments with unlabeled proteins were first performed to optimize the 2D electrophoresis from adipocytes. The protein extract (50 µg) was first diluted in 1 vol of lysis buffer (6 M urea, 2 M thiourea, 2% of 3-3-cholamidopropyldimethylammoniol, 30 mM Tris, 100 mMDL-dithiothreitol, pH 8.8), and 0.5% carrier ampholytes and DeStreack buffer (GE Healthcare) were added to make up the volume to 450 µL. First-dimension isoelectric focusing was carried out on an IPGphor system (GE Healthcare) by using precast immobilized pH gradient strips (pH 3 to 10 nonlinear, 24 cm; GE Healthcare). Strips were rehydrated overnight, and then low voltage (200 V) was applied in the initial step (1 h), followed by a gradual increase to 8,000 V and reaching a total focusing time of 62,000 Vh. The strips were then allowed to equilibrate with 2 incubations for 15 min each in a solution with 50 mM Tris HCl, pH 8.8, 6 M urea, 30% (vol/vol) glycerol, 2% (wt/vol) SDS, and 0.03% bromophenol blue (Sigma) as a dye, containing 65 mMDL-dithiothreitol for the first incubation or 250 mM iodoacetamide (GE Healthcare) for the second incubation at room temperature. Strips were applied to the top of 12% SDS-polyacrylamide gels, and run in a vertical Ettan DaltSix system (GE Healthcare). Gels were then silver-stained (Heukeshoven and Dernick, 1985), and representative images are shown in Figure 1.
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, various combinations, including dye-swap of labeled extracts to prevent dye-specific protein labeling, were run with the Cy2-labeled pool.
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Cyanine-labeled proteins were visualized after 2D-DIGE by scanning on a Typhoon 9400 Imager (GE Healthcare) in a fluorescence mode, as described previously (Rolland et al., 2007). All gels were scanned at a resolution of 200 µm (pixel size). Gel images converted to 16-bit TIF files were processed with DeCyder software (GE Healthcare), which allowed quantification, gel matching, and statistical analyses. Artifacts and mismatched spots were excluded from analyses. Pairwise comparison of 2 samples on one gel was performed by using the differential in-gel analysis module of the DeCyder software. The quantification is expressed as a spot ratio, comparing spot volumes on Cy3 or Cy5 images with corresponding spot volumes of the internal standard. Linking every sample to a common internal standard makes direct comparison of protein expression levels between multiple gels easier and more accurate (Alban et al., 2003). Ratios are inverse and are preceded by a minus sign for values less than 1. Protein-spot maps from comparable gels were matched by using the biological variation analysis (BVA) module of the DeCyder software. The differences in spot intensities (fold-change) between adipocytes of 2 anatomical locations were calculated with automatic and user-specific correction of background and artifactual variations by using the BVA feature. This module detects the consistency of differences between samples across all the gels, and applies statistics to associate a level of confidence for each of the differences. Log values are used so that the data points approach a normal distribution around zero to fulfill the requirements of subsequent statistical tests.
Protein Identification
After imaging and analysis, the gels were stored in 1% acetic acid at 4°C until spot excision. Spots of interest were excised from the gels by using an Ettan Spot Handling Workstation (GE Healthcare). After excision, the gels were silver-stained, as described previously, to visually confirm the accuracy of the excision. Excised spots were processed and digested with trypsin, as described previously (Rolland et al., 2007). Extraction was performed in 2 successive steps by adding 50% (vol/vol) acetonitrile and 0.1% trifluoroacetic acid. Digests were dried and dissolved with 2 mg/mL of 
-cyano-4-hydroxycinnamic acid in 70% acetonitrile and 0.1% trifluoroacetic acid before spotting them onto matrix-assisted laser desorption ionization (MALDI) targets (384 Scout MTP 600 µm AnchorChip, Bruker Daltonik, Bremen, Germany). Mass fingerprints were acquired by using a MALDI combined with a time-of-flight (TOF)/TOF mass spectrometer (Ultraflex, Bruker Daltonik) and processed with FlexAnalysis software (Bruker Daltonik). Autolysis products of trypsin were used for internal calibration. The monoisotopic masses of the tryptic peptides were used to query NCBI nonredundant sequence databases of all mammalian proteins by using the MASCOT search engine (http://www.matrixscience.com, last accessed February 26, 2007). Search conditions were as follows: initial rather open mass window of 70 ppm for an internal calibration and 200 ppm for an external calibration, one missed cleavage allowed modifications of cysteines by iodoacetamide, and methionine oxidation and N-terminal pyroglutamylation as variable modifications. To avoid incorrect identifications, at least 4 matched peptides per protein were required, and each matching was carefully checked manually by considering the MASCOT probabilistic score, the accuracy of the experimental to theoretical isoelectric point (pI), and the molecular weight (MW). The MASCOT baseline significant score is 68% of coverage of the entire amino acid sequence.
Statistical Analyses
Mean diameters of adipocytes were compared by ANOVA with the GLM procedure (SAS Inst. Inc., Cary, NC), with the fixed effect of anatomical site. Average abundance changes in 2D-DIGE were calculated from the differential in-gel analysis ratios, and Student’s t-test was used for statistics of spot intensities between pairs of adipocytes of 2 locations by using the BVA module. Differences were considered to be significant at P < 0.05. Only spots exhibiting at least a 2-fold change were then considered to be biologically different between 2 sites.
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RESULTS |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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Mean cell diameter was the least (P < 0.001) for i.m. adipocytes (Figure 2). Diameters of adipocytes derived from the 3 fat pads did not vary among regional locations.
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Analysis of the 10 images obtained after 2D-DIGE detected an average of 956 ± 19 spots per gel. Marked differences were found between i.m. adipocytes and other adipocytes. Altogether, 149 spots in i.m. adipocytes exhibited at least a 2-fold change (P < 0.05) in every comparison, with either a greater (96 spots) or a lower (53 spots) abundance than in the 3 other adipocyte types. These spots were then considered to be relevant for i.m. adipocyte features. The relative abundance of 17 spots differed (ratio exceeding |2|-fold difference, P < 0.05) when PERI adipocytes were compared with s.c. adipocytes. Only 7 spots differed between PERI and INTER adipocytes, and finally 6 spots differed in quantity between s.c. adipocytes and INTER fat cells.
Differentially Expressed Proteins
The 149 spots considered as characteristic of i.m. adipocytes and 26 nonredundant spots showing a variation in quantity between s.c., PERI, and INTER fat cells were selected for mass spectrometry analysis. However, only 52 spots (i.e., 30%) were identified, with a MASCOT score greater than 68 for the majority (89%) of them. Unfortunately, several proteins in the acidic region (5.7 < pH < 6.1) and with a predicted mass varying from 55.5 to 71 kDa were found to relate to porcine albumin (2 hits) or bovine albumin (30 hits, data not shown). The protein data descriptions for the other 20 proteins are listed in Table 2, including electron transfer flavoprotein subunit precursor and vinculin, with MASCOT scores slightly lower than the baseline significant score. They are categorized into 7 groups to facilitate interpretation.
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). Only 3 identified proteins were greater in abundance in i.m. adipocytes compared with fat cells derived from the 3 other fat regions. Both -actin and cell division cycle 42 homolog (Cdc42) in complex with guanine nucleotide dissociation inhibitors (GDI) were increased (P < 0.05) by at least 30-fold in i.m. adipocytes compared with other fat cells. Pro-hibitin-1 was 4-fold greater in quantity (P < 0.01) in the former compared with the latter cells (Figure 2
). The other identified proteins showed the lowest expression in i.m. adipocytes. They belonged to various cell pathways, such as fatty acid binding and lipolysis (perilipin; Figure 3), glycolysis (fructose-biphosphate aldolase, and 2 spots that are possibly different isoforms or posttranslationally modified forms of the same enolase protein), and the conversion or transfer of energy by cytoplasmic enzymes. For this latter pathway, one spot corresponded to malate dehydrogenase (MDH) and 2 spots corresponded to NAD phosphate-dependent isocitrate dehydrogenase (ICDH). Two enzymes involved in pathways that generate energy from fatty acids were also lower (P < 0.05) in abundance in i.m. adipocytes: the acyl-CoA dehydrogenase for long-chain fatty acids as the first mitochondrial flavin adenine dinucleotide-dependent enzyme (Figure 3), and succinyl-CoA:3-ke-toacid CoA transferase, an enzyme related to ketone body utilization. Finally, the abundance of 3 other mitochondrial proteins was also downregulated in i.m. adipocytes: the voltage-dependent anion channel 1, a membrane protein for the transport of ions and metabolites across the outer membrane, the catalase involved in antioxidative pathway, and electron transfer flavoprotein subunit precursor, involved in the regulation of the redox state.
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). Adipocytes derived from PERI location displayed the least abundance for MDH (P < 0.01) and the same trend (P = 0.02 to P = 0.18) was also observed for vimentin (i.e., an intermediate filament protein of the cytoskeleton around the lipid droplets). Conversely, lower (P < 0.03) expression of EH-domain-containing protein 2 was observed for s.c. fat cells compared with PERI and INTER adipocytes.
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DISCUSSION |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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), including a smaller mean diameter for isolated i.m. adipocytes compared with adipocytes stained on muscle histological cross-section (Gardan et al., 2006), we assumed that regional differences in protein abundance would be commensurate between digests and adipocytes in the tissues.
The identification of perilipin in adipocyte suspension was expected, because this abundant protein coats the large lipid droplet of differentiated adipocytes (Greenberg et al., 1991). The majority of the other proteins identified in the current study are known to be involved in energy production and conversion, supporting the notion that adipocytes are highly dependent on energy metabolism to store calories in the form of lipids (King and DiGirolamo, 1998). Activities of NAD phosphate+-ICDH and of cytoplasmic MDH (i.e., the sequence of this enzyme being different from that of the mitochondrial isoenzyme of the same species; Joh et al., 1987), are especially known to produce reducing equivalents for lipogenesis (Salway, 1994; Belfiore and Iannello, 1995). Furthermore, the presence of glycolytic enzymes, such as enolases and aldolase, has been also observed in differentiating 3T3-L1 adipocytes (Renes et al., 2005) and in mouse and human white adipose tissue homogenates (Sanchez et al., 2001; Corton et al., 2004). The presence of these enzymes is consistent with a significant involvement of the glycolytic metabolism in the first steps of the conversion of glucose during lipid accumulation in adipose cells (Temple et al., 2007). The finding of various mitochondrial proteins in adipocytes is also not really surprising, because mitochondrial genes represent approximately 10% of the highly expressed genes in porcine adipose tissue (Chen et al., 2006). In addition, mitochondriogenesis is generally coupled with adipogenesis (Wilson-Fritch et al., 2003). Finally, we identified proteins such as vimentin, enolase, and prohibitin, which are known to be expressed in adipocyte cytoplasm and which have also been reported into adipocyte-conditioned medium (Wang et al., 2004). Secretion signal peptides indicative of secretory proteins are present at least for vimentin (Chen et al., 2005) and prohibitin (Wang et al., 2004). Several other proteins were also detected; however, approximately 70% of the deregulated spots were not identified. This percentage of identification is close to the percentage reported by Renes et al. (2005), who identified 35% of the protein spots in 3T3-L1 differentiating fat cells. Information concerning proteins present in adipose tissue and related cells is limited. Although 1.3 million entries of pig sequences are in the public domain (Jiang and Rothschild, 2007), more than 50% of them are expressed sequence tags, and only 3 libraries were found for adipose tissue at the NCBI Uni-Gene Web site (Tuggle et al., 2007). High-abundance proteins might also have partially limited the detection of low-abundance proteins by MALDI-TOF spectrometry. Reducing sample complexity and dynamic range in protein abundance by subcellular fractionation of 3T3-L1 adipocytes, together with the use of nanoflow liquid chromatography-MS/MS/MS in a mass spectrometer with very high mass accuracy, has recently allowed the identification of more than 3,000 proteins in mouse differentiated adipocyte proteomes (Adachi et al., 2007). These approaches are difficult to apply to i.m. adipocytes because of their low numbers and the lack of availability of this recent state-of-the-art mass spectrometry. Finally, the prominent albumin reported here and in other proteomic studies involving adipose tissue homogenate (Sanchez et al., 2001) and adipocyte-conditioned media (Chen et al., 2005; Hausman et al., 2006) might have precluded the correct identification of other important proteins. Because the messenger coding for albumin has recently been reported to be expressed at high levels in pig adipocytes (Wang et al., 2006), the hypothesis of regional differences in albumin content between adipocytes cannot totally be ruled out.
Taken together, the current study shows some differences in protein abundance between s.c., PERI, and INTER fat locations. These results agree with other observations showing regionally distinct cell dynamic properties (Eguinoa et al., 2003; Lafontan and Berlan, 2003) and unique patterns of gene expression (Tchkonia et al., 2007) in preadipocytes or adipocytes isolated from s.c., perivisceral, omental, or intermuscular fat depots of different species. In the current study, lower abundance of EH-domain-containing protein 2 and greater content of vimentin, respectively, were observed in PERI adipocytes compared with s.c. adipocytes. Blüher and colleagues (2004) have shown that these 2 proteins are primarily affected by the presence or absence of insulin signaling in epidydimal fat of mice. Whether regional differences in abundance of these 2 proteins might be caused by differences in insulin action between s.c. and perirenal adipose tissues in pigs remains to be investigated.
More important, our study provides new evidence that the number of proteins differently expressed according to regional fat location is greater when adipocytes derived from skeletal muscle are considered. The present data, associated with our previous findings dealing with functional properties of adipocytes isolated from trapezius muscle (Gardan et al., 2006), further argue for a low lipogenic ability of i.m. adipocytes in pigs. Indeed, proteins associated with both the glycolytic pathway and the conversion of energy in the cytoplasm through MDH and ICDH were downregulated in i.m. adipocytes. We also provide new evidence for a low abundance of proteins associated with the catabolism of energy reserves in i.m. fat cells: perilipin and mitochondrial long-chain acyl-CoA dehydrogenase. β-Adrenergic receptor-stimulated lipolysis is almost totally lost in adipocytes lacking perilipin (Tansey et al., 2001). Therefore, low abundance of perilipin in i.m. adipocytes may account for the lower catecholamine-stimulated rate of lipolysis previously observed in i.m. adipocytes compared with s.c. pig adipocytes (Gardan et al., 2006). With respect to proteins upregulated in i.m. adipocytes, the identification of prohibitin-1, a protein also known to attenuate insulin-stimulated oxidation of glucose and fatty acids in adipose tissue (Vessal et al., 2006), is consistent with a possible reduction of the β-oxidative pathway in these cells. Koekemoer and Oelofsen (2001) demonstrated that pig adipocyte mitochondria have the appropriate enzymatic machinery to support the β-oxidative purpose. The differences in protein abundance reported here between i.m. adipocytes and other adipocytes might be explained by regional differences in adipocyte mean diameter, measured both on isolated cells (current study; Gardan et al., 2006) and histological cross-sections (Gardan et al., 2006). Indeed, differences in adipocyte cell size in mice are accompanied by alterations in protein expression at key steps in insulin-stimulated glucose uptake, triglyceride synthesis, lipolysis, and energy metabolism (Blüher et al., 2004). In addition, because i.m. adipose tissue is the latest developing adipose tissue in the pig (Hauser et al., 1997), there might be a delay in the temporal process of adipocyte differentiation in muscle compared with other body fat locations. For instance, perilipin gene expression in differentiating 3T3-L1 adipocytes (Arimura et al., 2004), and abundance of MDH, ICDH, long-chain fatty acyl-CoA dehydrogenase, and catalase proteins in white adipose tissue of rodents (Lanne et al., 2006) have been shown to be triggered by agonists of peroxisome proliferator-activated receptor Y, the master regulator of adipocyte differentiation in pigs (Fernyhough et al., 2007). Several other proteins exhibit temporal changes during adipocyte differentiation (Wang et al., 2004; Welsh et al., 2004). It is then possible that proteins participating in glucose and lipid metabolism may exhibit greater abundance, whereas other proteins known as negative growth regulators, such as prohibitin, may be at a lower level in i.m. adipocytes of older pigs. However, we have previously shown that regional differences between adipocytes observed at 80 d of age are also evident later in growth (Gardan et al., 2006). Finally, greater abundance in some proteins of i.m. adipocytes vs. other fat cells might be the cause, rather than the consequence, of inherent depot-specific adipocyte number and features. For instance, prohibitin-1 is described as negatively regulating cell proliferation (e.g., Smyth et al., 1993). In addition, a possible function of members of the evolutionarily conserved Rho family, including Cdc42 GTPases, has been suggested in modulating the switch between adipogenic and myogenic pathways (Sordella et al., 2003). Coculture systems of porcine myoblasts and preadipocytes might be useful in the future to test this hypothesis.
These results demonstrate that 2D electrophoresis-based proteome analysis is a potent tool that can be used to characterize i.m. adipocyte features. It was able to identify proteins in various metabolic and cellular pathways. Proteins with possible negative roles in cell growth are upregulated in muscle-derived fat cells. A better understanding of adipogenesis in skeletal muscle rather than of cell metabolism per se should then contribute to improved production efficiency and meat quality in the pork industry.
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Footnotes |
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2 Corresponding author: florence.gondret{at}rennes.inra.fr
Received for publication November 23, 2007. Accepted for publication February 15, 2008.
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LITERATURE CITED |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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