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© 2006 American Society for Nutrition J. Nutr. 136:2468-2474, October 2006

Biochemical, Molecular, and Genetic Mechanisms

SREBP1 and Thyroid Hormone Responsive Spot 14 (S14) Are Involved in the Regulation of Bovine Mammary Lipid Synthesis during Diet-Induced Milk Fat Depression and Treatment with CLA1,2

Kevin J. Harvatine and Dale E. Bauman*

Department of Animal Science, Cornell University, Ithaca, NY

* To whom correspondence should be addressed. E-mail: deb6{at}cornell.edu.


   ABSTRACT
TOP
 ABSTRACT
Introduction
 Materials and Methods
 Results
 Discussion
 LITERATURE CITED
 
Milk fat synthesis in dairy cows can be inhibited by unique fatty acid intermediates that are produced during rumen biohydrogenation. One of these inhibitory intermediates is trans-10, cis-12 conjugated linoleic acid (CLA), and this milk fat depression (MFD) involves a coordinated decrease in mammary expression of lipogenic enzymes. We investigated the sterol response element binding protein (SREBP) transcription factor system in the mammary tissue of cows during MFD, which was induced by a low forage, high oil (LF/HO) diet and trans-10, cis-12 CLA infusion. The LF/HO diet and CLA treatment decreased milk fat yield by 38 and 24%, respectively. Treatments causing MFD decreased expression of SREBP1 and the insulin responsive gene (INSIG) 1, consistent with decreased abundance of active SREBP1. The LF/HO diet also decreased expression of INSIG2 and SREBP cleavage activating protein. In addition, we identified the involvement of thyroid hormone responsive spot 14 (S14) in the regulation of mammary synthesis of milk fat. A broader role for S14 in the trans-10, cis-12 CLA-mediated decrease in fat synthesis was explored by mining publicly available microarray datasets, and we found that mouse adipose expression of S14 was decreased in response to CLA treatment. Overall, the decreased mammary expression of SREBP1, SREBP activation protein, and the coordinated reduction in SREBP1-responsive lipogenic enzymes provides strong support for a central role of SREBP1 in the regulation of milk fat synthesis. In addition, our results provide evidence for an involvement of S14 in mammary regulation of milk fat synthesis and a possible broader role for S14 in the reported antiobesity effects of CLA.


   Introduction
TOP
 ABSTRACT
 Introduction
Materials and Methods
 Results
 Discussion
 LITERATURE CITED
 
Mammalian milk fat concentration and composition are variable and responsive to nutritional factors. First described over a century ago, low-fat milk syndrome, commonly referred to as milk fat depression (MFD),3 is characterized by a marked decrease in the milk fat yield, typically observed in ruminants fed highly fermentable diets or diets that contain plant or fish oils. Recent investigations have established that the basis relates to an inhibition of mammary synthesis of milk fat by specific fatty acid intermediates produced during rumen biohydrogenation under certain dietary situations [see review by Bauman and Griinari (1)]. Trans-10, cis-12 conjugated linoleic acid (CLA) was the first of these unique biohydrogenation intermediates to be identified (2).

During MFD, transcription of mammary genes involved in milk fat synthesis are coordinately downregulated (1). Molecular mechanisms mediating this inhibition are not well understood, but a role for the sterol response element–binding protein (SREBP) family of transcription factors was proposed (3), based on their function as global regulators of expression for many genes involved in lipid synthesis (4). This was supported in studies with a bovine mammary epithelial cell line, where trans-10, cis-12 CLA decreased abundance of the nuclear active SREBP1 protein (5). An additional potential mechanism was disclosed when we identified thyroid hormone responsive spot 14 (S14) as a trans-10, cis-12 CLA responsive candidate gene in microarray analysis of bovine mammary epithelial cell cultures (unpublished data). Although its exact biochemical function is not known, S14 is a gene that encodes a nuclear protein that is closely associated with the regulation of fatty acid synthesis in lipogenic tissues (6).

Our objective was to investigate the expression of SREBP1 and S14 in the mammary tissue of lactating cows under 2 situations where milk fat synthesis is reduced, diet induced-MFD and administration of trans-10, cis-12 CLA. We found downregulation of SREBP1, SREBP1 regulatory proteins, and SREBP-regulated enzymes during milk fat depression. Moreover, our studies revealed a previously unrecognized involvement of S14 in the regulation of mammary synthesis of milk fat and a broader role in CLA-related regulation of lipid synthesis.


   Materials and Methods
TOP
 ABSTRACT
 Introduction
 Materials and Methods
Results
 Discussion
 LITERATURE CITED
 
    Animals and treatments. Experimental procedures were approved by the Cornell University Institutional Animal Care and Use Committee. Nine mid-lactation cows (193 ± 32 d postpartum; mean ± SD) from the Cornell University Teaching and Research Center were assigned randomly to replicated 3 x 3 Latin squares. Experimental periods were 14 d and specific treatments were control, short term administration of trans-10, cis-12 CLA, and feeding of a low forage, high oil (LF/HO) diet. During the control (CON) and CLA treatment periods, cows received a base diet that was formulated to meet or exceed nutrient requirements (Table 1). The LF/HO treatment cows received a diet that included 3.0% soybean oil and 1.5% fish oil. Diets were consumed ad libitum as a total mixed ration. Feed ingredients were sampled weekly, dried (55°C forced-air oven for 72 h), ground (Wiley mill with 1-mm screen; Arthur H. Thomas), and the nutrient composition was determined by wet chemistry procedures (7; Dairy One Cooperative).


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  		TABLE 1 Ingredient and nutrient composition of experimental diets

 
The CLA treatment involved a 3 d i.v. infusion of a CLA-Intralipid emulsion (days 12–14 of experimental period). Jugular catheters were installed on d 11 and the infusion supplied 10 g/d of trans-10, cis-12 CLA in equal doses infused every 6 h at a rate of 1 mL emulsion/min. The trans-10, cis-12 CLA methyl ester stock (BASF Corporation) contained 88.3% total CLA (98% trans-10, cis-12 isomer), 6.8% palmitic acid, 2.7% oleic acid, and 2.0% stearic acid. Previous investigations have shown that the methyl ester and free fatty acid forms of trans-10, cis-12 CLA are equally effective in inhibiting milk fat synthesis (8). Intralipid (Baxter Healthcare Corporation) emulsion contained 10% soybean oil and the CLA stock was added to give a final solution containing 10% CLA. Methyl esters of CLA were rapidly emulsified during homogenization.

    Milk sampling and analysis. Cows were milked 3 times/d and yields were recorded. Samples were taken at each milking on the last day and analyzed for fat and true protein, using a mid-infrared spectrophotometer (7; Dairy One Cooperative). Additional samples for fatty acid analysis were composited based on milk fat yield; lipid was extracted and transmethylated and methyl esters were quantified by GC according to Perfield et al. (9).

    Tissue biopsy. Mammary biopsies were taken 1–3 h after milking on d 14. Cows were given intravenous xylazine (15–25 mg) and a 15 mL lidocaine HCL subdermal block was administered above the incision site. A 0.5 cm incision was made in the skin at the midpoint of the rear quarter and the Magnum Biopsy Gun system (Bard Biopsy Systems) was used. Briefly, a 10 gauge cannula with trocar was inserted 5 cm into the mammary gland through the incision. The trocar was removed and a 12-gauge biopsy needle mounted in the biopsy gun was inserted through the cannula and fired. Tissue samples (30 mg tissue/biopsy) were rinsed with 0.9% saline solution, inspected to verify tissue homogeneity, and snap frozen in liquid nitrogen. Samples were stored at –80°C until RNA extraction. Multiple biopsies were routinely collected by reinserting the biopsy needle through the cannula. Immediately upon removal of the biopsy needle, the trocar was replaced and a purse string suture was placed around the cannula with #1 Nylon. The suture was tied as the cannula was removed and pressure applied to reduce collection of blood under the skin. The biopsy procedure resulted in minimal bleeding and milk appeared normal in 2–4 milkings following the biopsy; no intramammary infections or loss of production were encountered.

    RNA isolation and real-time PCR. Total RNA was isolated from ~30 mg mammary tissue from 1 biopsy using the RNeasy Lipid kit (Qiagen) and DNA contamination was removed by on-column DNase treatment (RNase-Free DNase Set; Qiagen). RNA concentration and integrity were determined by a 2100 BioAnalyzer (Agilent Technologies). Total RNA was reverse transcribed using SuperScript III First Strand Synthesis kit (Invitrogen) with random primers. Quantitative real-time reverse transcriptase PCR (qRT-PCR) assays were developed for genes of interest (Supplemental Table 1). Briefly, primers were designed on or spanning exon boundaries when possible using PrimerExpress v2.0 (Applied Biosystems), and optimal primer pairs were selected using Primer3 (10) and PerlPrimer (11). qRT-PCR included iTaq SYBR Green Supermix with ROX (Bio-Rad Laboratories) and 400 nmol/L of gene-specific forward and reverse primers (Invitrogen). cDNA (5–25 ng) was amplified using a 2-step program (95°C for 15 s and 60°C for 60 s) with an ABI PRISM 7000 sequence detection system (Applied Biosystems). Dissociation curves were generated at the end of amplification to verify presence of a single product. Sample message level was determined, relative to a dilution curve of pooled mammary cDNA (12).

    Data mining. Publicly available data from 2 microarray experiments were downloaded [NCBI GEO Dataset GSE1580 and 14]. Specifics of animals and experimental procedures were previously described (13,14). Briefly, House et al. (13) fed 9-wk-old mice (NCSU M16 line) diets containing 1% trans-10, cis-12 CLA or control (1% linoleic acid) for 14 d; total RNA was extracted from the epididymal adipose tissue of 70 mice, pooled into 4 groups per treatment, and analyzed on the Mouse Oligo microarray slide (G4121A; Agilent Technologies). Hargrave et al. (14) fed 12-wk-old mice (UNL MC line) diets containing 0 or 2% of a mixed isomer CLA (~50% trans-10, cis-12) for 16 d; adipose tissue RNA from 5 littermate pairs was analyzed on the Mouse 430A 2.0 GeneChip (Affymetrix).

    Statistical analysis. Data were analyzed using the fit model procedure of JMP (version 5.0, SAS Institute). The model to test treatment means included the random effect of cow and the fixed effects of period and treatment. Additionally, the geometric mean of 3 housekeeping genes (18S ribosomal subunit, ß-actin, and ß2-microglobulin) was calculated using GeNorm (15) and used as a covariant in the model (16). Data points with Studentized Residuals >2.5 were considered outliers and excluded from analysis. Few points were excluded in analysis and rarely more than 1 per response variable. Preplanned contrasts included the effect of CLA (CON vs. CLA) and the effect of LF/HO diet (CON vs. LF/HO). The relation between expression of individual genes (SREBP1 and S14) and lipogenic enzymes [fatty acid synthase (FASN) and lipoprotein lipase (LPL)] was tested with the above model by replacing treatment with the predictor gene of interest. Relations between expression of genes was declared significant at P < 0.01 and a trend at P < 0.05. For statistical analysis of S14 data from House et al. (13), the P-value and fold change for the S14 annotated probe was downloaded (GSE1580). For the data of Hargrave et al. (14), downloaded fluorescent intensities of the S14 annotated probeset were log (base 2) transformed and treatment means were compared using a t test.


   Results
TOP
 ABSTRACT
 Introduction
 Materials and Methods
 Results
Discussion
 LITERATURE CITED
 
    Milk fat synthesis and expression of lipogenic enzymes. The short term infusion of trans-10, cis-12 CLA decreased milk fat concentration and yield by 23 and 24%, respectively (Table 2). The LF/HO diet decreased milk fat concentration and yield to an even greater extent: 31 and 38%, respectively. Analysis of fatty acids revealed decreased secretion of both de novo and preformed fatty acids (Table 2). However, the decrease was greater for de novo synthesized fatty acids, resulting in a shift in milk fat composition to an increased proportion of long-chain fatty acids (Supplemental Table 2). Expression of key enzymes in mammary fat synthesis was analyzed by qRT-PCR to verify downregulation of lipogenic enzymes. Expression of FASN and LPL were downregulated for both treatments, whereas expression of stearoyl-CoA desaturase (SCD) only tended (P < 0.10) to be downregulated by the LF/HO treatment (Fig. 1A).


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  		TABLE 2 Effect of trans-10, cis-12 conjugated linoleic acid and a LF/HO diet on performance and milk fat synthesis of lactating cows

 

Figure 1
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  		Figure 1  Effects of trans-10, cis-12 CLA and a LF/HO diet on cows' mammary mRNA abundance of key lipogenic enzymes (FASN, LPL, and SCD) (A); key transcription factors (SREBP1 and S14) (B); and genes associated with SREBP1 processing (INSIG1, INSIG2, and SCAP) and transcriptional coactivators (PGC1{alpha}, and PGC1ß) (C). Values are means ± SE, n = 8 for CON and LF/HO and n = 9 for CLA. Values are expressed relative to CON. Asterisks indicate different from control: *P < 0.05, **P < 0.01, and ***P < 0.001.

 
    Expression of SREBP1 and SREBP1 regulatory proteins. Expression of SREBP1 was downregulated in mammary tissue during both treatments (Fig. 1B). Message abundance of several genes encoding SREBP processing proteins and transcriptional coactivators were analyzed. Insulin responsive gene (INSIG) 1 was decreased during trans-10, cis-12 CLA treatment and the LF/HO diet, and mammary expression of INSIG2, SREBP cleavage activating protein (SCAP), and peroxisome proliferative activated receptor, gamma, coactivator (PGC) 1{alpha} were moderately decreased during diet-induced MFD (Fig. 1C). However, expression of PGC1ß was not altered by treatments.

    Expression of S14 during milk fat depression. Expression of S14 was downregulated in mammary tissue during treatment with trans-10, cis-12 CLA, and LF/HO-induced MFD (Fig. 1B). Relatively little is known about S14 in bovine tissues, so we examined its tissue expression profile (Fig. 2). Adipose tissue and liver were the predominant sites of S14 expression. Expression of S14 in lactating mammary tissue was >2-fold greater than observed for nonlactating mammary tissue, and substantially greater (>75-fold) than lung. The tissue profile for midline 1 interacting protein 1 (MID1IP1), a gene with sequence homology to S14, was also examined, and it was expressed at similar levels in all tissues expect skeletal muscle, which had ~16-fold greater expression than mammary tissue (Fig. 2).


Figure 2
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  		Figure 2  Tissue profile of S14 and MID1IP1 in cows. With the exception of nonlactating mammary tissue, tissues came from cows in the mid- to late lactation. Values are plotted on a log axis; n = 6 for subcutaneous adipose tissue (AT), liver (Liv), and lactating mammary gland (Lact), and n = 3 for nonlactating mammary tissue (Nlact), uterus (Uter), lung, brain, skeletal muscle (Musc), and heart.

 
    Expression of S14 in mouse adipose tissue. Trans-10, cis-12 CLA is often referred to as having antiobesity effects because of its ability to reduce body fat accretion in several species (17). To see if the downregulation of S14 expression was more broadly applicable to CLA treatment, we mined publicly available microarray data. House et al. (13) and Hargrave et al. (14) treated mice with trans-10, cis-12 CLA and observed a substantial reduction in body fat concentration. We specifically extracted expression data for adipose S14 from their microarray data and found it was also markedly reduced (Fig. 3).


Figure 3
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  		Figure 3  Effects of trans-10, cis-12 CLA on mouse adipose mRNA abundance for S14, using online microarray data from studies by House et al. (13) and Hargrave et al. (14). Values are means, n = 4 for both experiments. Asterisks indicate different from control: *P < 0.05 and ***P < 0.001.

 

   Discussion
TOP
 ABSTRACT
 Introduction
 Materials and Methods
 Results
 Discussion
LITERATURE CITED
 
Diet-induced MFD is a naturally occurring situation in dairy production that involves an interrelation between rumen digestive processes and mammary synthesis of milk fat. MFD is observed for a range of diets and its basis relates to a shift in ruminal biohydrogenation and the production of unique fatty acid intermediates. Referred to as the "Biohydrogenation Theory" (1), the first specific ruminal biohydrogenation intermediate demonstrated to inhibit milk fat synthesis was trans-10, cis-12 CLA (2). We observed that milk fat yield decreased 38% with the LF/HO diet and 24% with the 3-d infusion of trans-10, cis-12 CLA. Consistent with previous research (1), we observed a reduction in the secretion of fatty acids originating from both de novo synthesis and uptake from circulation. Thus, many of the biochemical processes involved in the synthesis of milk fat must be downregulated.

The SREBP family of transcription factors function as global regulators of lipid metabolism (4). SREBP1c predominately regulates enzymes involved in fat synthesis and is expected to be the predominant transcript expressed in mammary tissue. However, because of the limited amount of available annotated sequence, our qRT-PCR assay does not distinguish between isoforms 1a and 1c; thus, we refer to them collectively as SREBP1. The full length SREBP protein is complexed with SCAP and anchored in the ER through association with a third protein, either INSIG1 or INSIG2 (4,19,20). SREBP is activated by dissociation of INSIG from the SREBP/SCAP complex, allowing translocation of the complex to the Golgi where 2 proteases act to release a nuclear fragment (nSREBP). In turn nSREBP translocates to the nucleus where it binds to sterol-regulatory elements (SRE) in the promoter/enhancer regions of target genes, recruits coactivators, and stimulates transcription (4).

SREBP1 is one of the predominant mechanisms of inhibition of fat synthesis by PUFA (21). We first examined the SREBP-regulatory system in bovine mammary epithelial cells (MAC-T cell line) and observed decreased abundance of nSREBP1 protein during trans-10, cis-12 CLA inhibition of fatty acid synthesis (5). In this investigation, we extend these results to in vivo and demonstrate decreased expression of SREBP1 and INSIG1 during both trans-10, cis-12 CLA treatment and diet-induced MFD. Expression of SREBP1 and INSIG1 correlates with the concentration of the nSREBP1 protein, as both SREBP1c and INSIG1 genes contain an SRE in their proximal promoter (22,23). INSIG1 is normally expressed at higher levels than INSIG2, has a faster turnover, and is more dynamic (20). SCAP and INSIG typically function in stoiciometric concentrations (20), and we observed SCAP and INSIG2 expressions were decreased during diet-induced MFD, but not by CLA treatment. This treatment difference may relate to the greater decrease in milk fat or the longer duration of MFD for the LF/HO diet.

The PGC-1 family of transcription coactivators are environmentally responsive factors regulating tissue metabolism (24). Specifically, PGC-1ß is increased in response to high fat intake and coactivates SREBP1 (25). In lactating rat mammary tissue, corn oil decreased expression of FASN, SREBP1c, and PGC-1ß, but increased expression of PGC-1{alpha}. In this study, expression of PGC-1ß was unaffected by treatment and PGC-1{alpha} was only slightly decreased by the LF/HO treatment. Thus, our results offer little or no support for altered transcription of the PGC family of coactivators in the regulation of milk fat synthesis.

The synthesis and secretion of milk fat by the mammary gland involves an integration of different biochemical processes, and characterization of lipogenic genes during MFD highlights the coordinated downregulation in the expression of key enzymes associated with these processes (Table 3). SREBP1 is highly expressed in the lactating bovine mammary gland (unpublished data) and, as illustrated in Table 3, many of these key enzymes are transcriptionally regulated by SREBP1 (28,29). In mice, SREBP1 is upregulated at the initiation of lactation (30), and disruption of the SREBP1c gene results in a 41% decrease in milk fat concentration (31). Interestingly, a maximum 50% reduction in milk fat synthesis is observed during diet-induced MFD (1) and during dose-response studies with exogenous trans-10, cis-12 CLA (8).


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  		TABLE 3 Summary of SREBP1-regulated lipogenic genes in bovine mammary tissue that demonstrate a coordinated reduction in expression during diet-induced MFD and with CLA treatment

 
Mammalian regulation typically includes redundant systems for amplification of signaling and regulation of biochemical processes. S14 has been implicated in the transcriptional regulation of lipogenic genes (6,32,33), and we identified S14 as a trans-10, cis-12 CLA responsive candidate gene in microarray analysis of bovine mammary cultures (unpublished data). In rodents, S14 was highly expressed in lipid-synthesizing tissues, including lactating mammary tissue (6). In our survey of bovine tissues, S14 was highly expressed in liver and adipose tissue and moderately expressed in mammary tissue. In addition, previous studies established that S14 expression was upregulated in mammary tissue during lactation in humans (34) and mice (30), and we also observed this for cows. MIG1IP1, a protein with sequence homology to S14, has been proposed as a lipogenic factor with possible redundancy to S14 function (35). However, in our study, the lack of treatment effects on mammary expression of MIG1IP1 (data not shown) and the tissue profile of MIG1IP1 offer little support for a role in regulation of lipogenesis in the lactating bovine mammary gland.

The exact biochemical function of S14 has not been established. Originally identified as a protein acutely responsive to thyroid hormone [see review by Cunningham et al. (6)], S14 is primarily a nuclear protein that forms homo- and heterodimers (6) and interacts with transcription factors (36). The S14 promoter also contains a SRE (37) and its expression is highly responsive to nSREBP1 (33). Perhaps the strongest evidence of S14 function comes from studies of rat heptocytes, where transfection with S14 antisense oligonucleotide prevented expression of lipogenic enzymes (32,38). In addition, mice with a partial S14 knockout have decreased milk fat concentration, due to decreased de novo fatty acid synthesis, although surprisingly, activities of mammary lipogenic enzymes were unaltered (35).

We found mammary expression of S14 was downregulated during diet-induced MFD and trans-10, cis-12 CLA treatment. This is the first study, to our knowledge, of the regulation of mammary expression and, to our knowledge, the first to examine the regulatory role of CLA. However, hepatic expression of S14 has been extensively investigated and was shown to be responsive to a range of metabolic hormones and dietary nutrients, including PUFA [see review by Cummingham et al. (6)]. Trans-10, cis-12 CLA reduces body fat accretion in several species (17), although the CLA dose in studies of its antiobesity affect is substantially greater than that required to reduce milk fat synthesis (0.5–2.0% of diet in rodents vs. 0.045% of diet in this study). Using microarray data from studies with mice (13,14), we found that CLA treatment resulted in a significant reduction in expression of S14 in adipose tissue. Thus, S14 may be more broadly implicated in the mechanism by which CLA is able to affect lipid metabolism.

Multivariate analysis in this study revealed a significant relation in bovine mammary tissue between expression of S14 and expression of FASN and LPL (R2 = 0.86 and 0.42, respectively; Supplemental Table 3). Altered expression of S14 has also been associated with other unique phenotypes involving regulation of fat synthesis. For example, abnormalities in the regulation of adipose S14 expression have been reported in obese subjects (39). Likewise, gene expression profiling identified differential expression of S14 in livers of chickens selected for growth (40), hepatic tissue of chickens selected for adiposity (41), and muscles of cattle that differ in marbling (42). Lastly, S14 is also a component of the lipogenic phenotype observed in aggressive breast cancers (34) and knockdown or overexpression of S14 results in corresponding effects on breast cancer cell growth (33). The anticarcinogenic affect of trans-10, cis-12 CLA has been well characterized for in vitro and in vivo models [see review by Ip et al. (43)]; based on our results, the possible role of S14 in the mechanism merits examination.

In conclusion, decreased expression of SREBP1 and proteins associated with SREBP1 activation during MFD, combined with the presence of SREBP response elements in lipogenic genes downregulated during MFD, provide strong evidence for SREBP1 as a central signaling pathway regulating fatty acid synthesis in bovine mammary glands. Furthermore, downregulation of S14 during diet-induced MFD and trans-10, cis-12 CLA treatment is consistent with a role for S14 in mammary fatty acid synthesis, possibly as a SREBP1 secondary cellular signal or a lipogenic factor.


   ACKNOWLEDGMENTS
 
The authors thank Debra Dwyer, Stephanie Thorn, James Perfield, II, Neil Mittelman, and Euridice Castañeda-Gutiérrez for their assistance in this experiment. In addition, we thank Dr. Patricia A. Johnson for assistance with the qRT PCR analyses, Baxter Healthcare for providing the Intralipid emulsion, and BASF for providing the purified trans-10, cis-12 CLA.


   FOOTNOTES
 
1 Research supported in part by National Research Initiative Competitive Grants Program, Cooperative State Research, Education, and Extension Service, United States Department of Agriculture (grant #2006-35206-16643), U.S. Department of Agriculture IFAFS (2001-52100-11211) and Cornell Agricultural Experiment Station. Back

2 Supplemental Tables 1–3 are available with the online posting of this paper at jn.nutrition.org. Back

3 Abbreviations used: CLA, conjugated linoleic acid; CON, control treatment; DM, dry matter; FA, fatty acids; FASN, fatty acid synthase; INSIG, insulin induced gene; LF/HO, low forage and high oil; LPL, lipoprotein lipase; MID1IP1, midline 1 interacting protein; MFD, milk fat depression; PGC, peroxisome proliferative activated receptor, gamma, coactivator; nSREBP, nuclear fragment of SREBP; qRT-PCR, quantitative real-time reverse transcriptase PCR; S14, thyroid hormone responsive spot 14; SCAP, SREBP cleavage-activating protein; SCD, stearoyl-CoA desaturase; SRE, SREBP response element; SREBP, sterol response element–binding protein. Back

Manuscript received 6 June 2006. Initial review completed 16 June 2006. Revision accepted 16 July 2006.


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 ABSTRACT
 Introduction
 Materials and Methods
 Results
 Discussion
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